1
Q
Signal molecules can take many forms. Name seven.
A
Signal molecules can be proteins, peptides, AAs, nucleotides, steroids, FA derivatives, or even dissolved gases (NO); typ rely on only a handful of basic comm styles: endocrine, paracrine, synaptic, and contact-dep.
2
Q
Signal molecules can be proteins, peptides, AAs, nucleotides, steroids, FA derivatives, or even dissolved gases. However, they rely on only a handful of basic comm styles, such as: _______, ________, ________, and _________.
A
Signal molecules can be proteins, peptides, AAs, nucleotides, steroids, FA derivatives, or even dissolved gases. However, they rely on only a handful of basic comm styles, such as: endocrine, paracrine, synaptic, and contact-dep.
3
Q
Diff types of cell-to-cell comm vary most critically in ______ and _______.
A
Diff types of cell-to-cell comm vary most critically in speed and selectivity. From slowest/least selective to fastest/most selective: endocrine, paracrine/autocrine, synaptic, contact-dep.
- Endocrine signals - lowest speed and selectivity: Endocrine glands prod hormones → secreted into bloodstream → widely distributed thru/o body (or plant’s sap).
- Paracrine signals - released into local ECF → act locally.
- Incl many signals that regulate inflam at site of infection or control cell proliferation in healing wounds.
- Autocrine signals - target same cell that produced signal; e.g. cancer cells promote own survival.
- Synaptic signals - xmtd electrically along nerve cell axon to terminals → transduced into chem signal in form of secreted nxmtrs (into synaptic cleft) → synapse on adj target cells.
- Long distance; but fast and v specific.
- Contact-dep signals - cell-surface signal directly binds receptor on adj cell, i.e. no secretion.
- Direct physical contact → most intimate and short-ranged form of comm.
- E.g. embryonic dev - allows adj cells to specialize.
4
Q
Name at least one example of a hormone, local mediator, nxmtr, and contact-dep signal molecule, incl its site of origin, chemical nature, and some of its effects.
A
- Hormone - e.g. EPI: derivative of tyrosine (tyr/Y) synthd in adrenal gland (on top of kidneys) → ↑ HR/BP and metabolism.
- Local mediator (para/autocrine) - e.g. platelet-derived growth factor (PDGF): protein synthd by various cells, incl blood platelets → stims many cell types to proliferate.
- Nxmtr - e.g. ACh: choline derivative synthd in nerve terminals → typ excitatory (EPSP), but can be inhibitory in some cell types.
- Contact-dep - e.g. delta: xmem protein synthd in prospective neurons and various other developing cell types → inhibits adj cells fr becoming specialized in same way as signaling cell.
5
Q
To remain a local stimulus, paracrine signals must be prevented fr straying too far fr points of origin. How might this could be accomplished?
A
Most paracrine signaling molecules are v short-lived after they are released fr a signaling cell: they are either degraded by EC enzymes or are rapidly taken up by adj target cells. In addition, some become attached to ECM and are thus prevented fr diffusing too far.
6
Q
Each cell responds to a ______ (limited/expansive) set of EC signals, dep on its ______ and ______.
A
Each cell responds to a limited set of EC signals, dep on its history and current state.
- Cells are v selective - based on specialized function and presence of appropriate receptors.
7
Q
Signals can alter a cell’s shape. What else can they effect?
A
Signal effects - can alter cell’s shape, movement, metabolism, gene expression, or combo of these.
8
Q
Signals typ conveyed into IC signals → IC relay → alter activity of _______ proteins wh have direct effect on behavior of cell.
A
Signals typ conveyed into IC signals → IC relay → alter activity of effector proteins wh have direct effect on behavior of cell.
- Diff types of cells respond to same signal in diff ways.
- E.g. ACh: ↓ firing rate of heart pacemaker cells, but ↑ secretion of saliva fr salivary glands; both types of cells have same ACh receptors; additionally, ACh binds to a diff receptor on skeletal muscle → cell contracts.
9
Q
T/F: The EC signal acts as the ‘message’, i.e. all cells respond to partic EC signal the same way.
A
False
IC relay differs b/w cells → diff types of cells respond to same signal in diff ways.
- E.g. ACh: ↓ firing rate of heart pacemaker cells, but ↑ secretion of saliva fr salivary glands; both types of cells have same ACh receptors; additionally, ACh binds to a diff receptor on skeletal muscle → cell contracts.
- EC signal alone is not the message; it’s how target cell receives/interprets the signal.
10
Q
T/F: Cells typ posses only one or a few diff types of receptors.
A
False
Cells typ possess many diff receptors → simult response → subtle and complex control via diff combos.
- Presence of one signal often modifies effects of another.
- One combo might enable cell to survive; another might drive it to differentiate in some specialized way; and another might cause it to divide.
- Absent any signals, most animal cells undergo apoptosis.
11
Q
Absent any signals, what is the response of most animal cells?
A
Absent any signals, most animal cells undergo apoptosis.
12
Q
EC signals vary in their speed of transmission and range of distribution. A cell’s response to these signals can also be fast or slow, dep on what needs to happen upon receival. Describe such diffs in fast and slow responses.
A
- Fast signals - affect activity of proteins already present inside target cell.
- E.g. ACh stims skeletal muscle to contract w/i milliseconds.
- Typ incl changes in cell movement, secretion, or metabolism.
- Slow signals - req changes in gene expr → prod of new proteins.
- Typ incl cell differentiation or cell growth/division.
13
Q
EC signals typ fall into two classes—EC or IC receptor signals—dep on what feature of the signal?
A
EC signal molecules typ fall into two classes, dep on pmem permeability.
- EC receptor signals - largest class; too large/hphilic to cross pmem of target cell → bind EC receptor proteins → generate 1+ IC signal molecules.
- IC receptor signals - small/hphobic enough to pass thru pmem → bind IC receptors in cytosol/nucleus or activate IC enzymes directly; incl steroid hormones (e.g. cortisol, estradiol, testosterone) & thyroid hormones (e.g. thyroxine), and nitric oxide (NO).
14
Q
Describe how nuclear receptors function.
A
Nuclear receptors - incl BOTH cytosolic and nuclear receptors; activated by hormone (IC receptor signal) binding → act as xcr regulators (in nucleus).
- Mechanism overview: cell unstimulated, w nuclear receptors typ inactive → hormone binds IC receptor → IC receptor changes into active conform wh can promote/inhibit xcr of specific target genes.
- E.g. cortisol: prod by adrenal gland in response to stress → crosses pmem to bind/activate cytosolic receptor protein → receptor-hormone complex xprtd into nucleus via nuclear pores → active receptor can bind specific regulatory seqs in DNA and activate/repress xcr of specific target genes.
- Other hormone receptors are already bound to DNA, even in absence of hormone.
15
Q
T/F: Some hormone receptors are bound to DNA, even in absence of hormone.
A
True
Some hormone receptors are bound to DNA, even in absence of hormone.
- E.g. Hormone crosses pmem → binds cytosolic receptor → releases IC signal → enters nucleus thru nuclear pores → binds DNA-bound hormone receptor → regulates xcr.
16
Q
T/F: Ea hormone binds diff nuclear receptor, and ea receptor acts at a diff set of regulatory sites in DNA.
A
True
Ea hormone binds diff nuclear receptor, and ea receptor acts at a diff set of regulatory sites in DNA.
- A given hormone typ regulates diff sets of genes in diff cell types → evoke diff physio responses in diff target cells.
17
Q
Testosterone shapes formation of external genitalia and influences fetal brain dev. At puberty, it triggers dev of male secondary sexual characteristics. Some v rare individuals are genetically male (have both an X and Y chromo) but lack testosterone receptors as result of a mutation. Describe how such an individual would develop?
A
They would develope as females—same path that genitalia and brain would develope if neither male nor female hormones were produced.
18
Q
Some dissolved gases can cross pmem and activate IC enzymes directly. One such gas, nitric oxide (NO), typ targets guanylyl cyclase in smooth muscles. Describe this mechanism of action, such as in the relaxation of endothelial cells that line all blood vessels.
A
Smooth muscle (blood vessel) relaxation mechanism: nerve endings secrete ACh → ACh binds EC receptors on endothelial cells → activates NO synthase (NOS) inside endothelial cell → NOS catalyzes synth of NO fr arginine (arg/R) → NO rapidly diffuses out of endothelial cell, thru basal lamina, and into surrounding smooth muscle cell → NO binds/activates guanylyl cyclase inside smooth muscle cell → catalyzes synth of cGMP fr GTP → cGMP causes smooth muscle cell to relax → blood vessel dilates.
- Nitric Oxide (NO) gas - synthd fr arginine (arg/R); diffuses readily fr site of synth into adj cells → quickly converted into nitrates/nitrites (w half-life of ~5-10 sec) by reacting w oxygen and water outside cell.
- Guanylyl cyclase - NO diffuses thru mem → binds/activates guanylyl cyclase → catalyzes synth of cGMP fr GTP → cGMP acts as small IC signal → final effect of NO signal chain (e.g. blood vessel relaxation).
- E.g. Viagra: blocks enzyme that degrades cGMP → prolongs NO signal.
- cGMP is v similar in struc/mechanism of cAMP, a much more commonly used IC signal.
- Endothelial cells - flattened cells that line every blood vessel.
19
Q
The vast majority of signals are too large/hphilic to diffuse thru pmem, instead bind to EC receptors on pmem and generate new IC signals. Describe several crucial functions that IC signals perform.
A
IC relay until interaction w effector proteins (metabolic enzymes, cytoskeletal protein, or xcr regulators) → cell response, e.g. altered metabolism, cell shape/movement, or gene expr (on/off).
IC signals perform many crucial functions:
- Relay signal onward → help spread it thru cell.
- Amplify signal → ↑ sensitivity to EC signal, i.e. few EC signals reqd to evoke large IC response.
- Detect signals fr 1+ IC signal pathway → integrate them before relaying signal onward.
- Distribute signal to 1+ effector protein → creates branches in info flow and evoke complex response.
20
Q
Some proteins in IC path may be held in close proximity by _________ → activated at specific location w/i cell and w greater speed/eff/selectivity.
A
Some proteins in IC path may be held in close proximity by scaffold proteins → activated at specific location w/i cell and w greater speed/eff/selectivity.
21
Q
IC signal pathway is typ subject to feedback regulation → complex responses. Wh type of feedback can result in switch-like responses? Oscillating responses?
A
IC signal pathway is typ subject to feedback regulation → complex responses.
- Positive feedback can generate all-or-none, switch-like responses.
- Negative feedback can generate oscillating responses.
22
Q
In principle, how might an IC signal amplify the signal as it relays it onward?
A
The IC signal protein could be an enzyme that produces a large # of other small IC signals (e.g. cAMP or cGMP). Or, it could be an enzyme that modifies a large # of IC target proteins (e.g. by phosphorylation).
23
Q
Some IC signal proteins act as molecular switches: receive signal → toggle fr inactive to active state → stim/suppress other proteins in pathway → persist in active state until some other process inactivates/switches them off again. Why is switching off so critical, and what are two classes of molecular switch proteins?
A
Switching off is critical - every activated protein in pathway must be reset to original, unstim’d state → enables signal pathway to recover and xmt additional signals.
Two classes of molecular switch proteins:
- In/activated by phosphorylation - much larger class; protein kinase covalently attaches P group onto switch protein; protein phosphatase hydrolyzes P group fr switch protein.
- GTP-binding proteins - smaller class; active when GTP bound, inactive when GDP bound.
24
Q
Many switch proteins are themselves protein kinases. Describe how this helps amplify, distribute, and regulate a signal.
A
Many switch proteins are themselves protein kinases → often organized into phosphorylation cascades: one protein kinase, activated by phosphorylation, phosphorylates next protein kinase in the seq, etc. → signal xmtd onward and, in the process, amplifies, distributes, and regulates it.
- Recall: two classes of swtich proteins - those in/activated by phosphorylation or GTP-binding.
- Activity of switch protein deps on balance b/w protein kinase/phosphatase activity.
- Two main types of protein kinases in IC signal path: ser/thr kinases (most common) and tyr kinases.
25
GTP-binding proteins are the smaller of the two classes of switch proteins (other being those in/activated by de/phos). These proteins are activated by GTP binding → stims intrinsic GTP-hydrolyzing (GTPase) activity → shut themselves off by hydrolyzing GTP (Pi released, GDP remains bound).
What are the two main types of GTP-binding proteins in IC signal paths?
***Trimeric* GTP-binding proteins (Trimeric GTPases; "G proteins")** - relay messages fr GPCRs.
***Monomeric* GTP-binding proteins (Monomeric GTPases)** - aided by two sets of regulatory proteins:
* **Guanine ntide exchange factors** (**GEFs**) *activate* switch proteins by promoting exchange of GDP for GTP.
* **GTPase-activating proteins** (**GAPs**) - *inactivate* switch proteins by promoting GTP hydrolysis.
* Both involved w enzyme-coupled receptors (in/activation of Ras)
26
What three main classes do most cell-surface (EC) receptors fall into?
**Ion-channel-coupled receptors** (**ICCRs; also: xmtr-gated ion channels**) - change permeability of pmem to selected ions → alter mem-pot and, if conditions are right, prod elec current.
**G-protein-coupled receptors** (**GPCRs**) - activate mem-bound, *trimeric* GTP-binding proteins (G proteins) → activate (or inhibit) an enzyme/ion channel in pmem → initiate IC signal cascade.
**Enzyme-coupled receptors** (**ECRs**) - either act as enzymes or assoc w enzymes inside cell; when stimulated, enzymes can activate wide variety of IC signal pathways.
27
T/F: EC signals can have more than one receptor, including receptors fr diff classes, e.g. ion-channel-, G-protein-, or enzyme-coupled receptors.
True
**EC signals can have 1+ receptor (even a diff class) → # of diff types of receptors \> # of assoc EC signals**.
* E.g. ACh: acts on skeletal muscle cells via an ICCR, whereas in heart cells it acts thru a GPCR → receptors prod diff IC signals → skeletal muscle cell contraction ↑ whereas heart contraction ↓.
* Variability offers convenient target for drugs.
28
Ion-channel-coupled receptors function in the simplest and most direct way by converting _____ signals into _____ ones.
Ion-channel-coupled receptors function in the simplest and most direct way by converting **chemical** signals into **electrical** ones.
* Recall: xmtr-gated ion channels transduce a chem signal—a pulse of secreted nxmtrs—directly into an electrical signal—a change in mem-pot.
* **Mechanism**: nxmtr binds ICCR/ion channel → receptor changes conform → ion channel opens → ion influx/efflux → change in mem-pot (w/i milliseconds) → may trigger AP or make it easier/harder for other nxmtrs to do so.
* Opening of Ca2+ channels has additional imp effects, as changes in IC [Ca2+] can profoundly alter activities of many Ca2+- responsive proteins.
29
The signaling mechanisms used by a steroid-hormone-type nuclear receptor and by an ICCR are relatively simple as they have few components. Can they lead to an amplification of the initial signal, and, if so, how?
* For steroid-hormone receptor, a one-to-one complex of steroid and receptor binds to DNA to activate or inactivate gene xcr; thus, *no amplification b/w ligand binding and xcr regulation*. **Amplification occurs later**, bc gene xcr gives rise to many mRNAs, ea wh is translated to give many copies of protein it encodes.
* For ICCR, a single ion channel will let thru thousands of ions in the time it remains open; this serves as the amplification step in this type of signaling system.
30
T/F: GPCRs form largest family of cell-surface receptors.
True
GPCRs form largest family of cell-surface receptors.
* GPCRs mediate responses to enormous diversity of EC signals, incl hormones, local mediators, and nxmtrs.
* Diversity of function → attractive target for drugs; ~1/3 of all drugs used today work thru GPCRs.
31
T/F: All GPCRs are seven-pass xmem receptor proteins w IC G-protein bound to cytoplasmic domain.
True
All GPCRs are seven-pass xmem receptor proteins w IC G-protein bound to cytoplasmic domain.
* Receptors for small signals (EPI/ACh) - ligand typ binds *deep w/i plane of mem* to a pocket formed by AAs fr several xmem segments.
* Receptors for larger protein signals - typ have large EC domain that, t/g w some xmem segments, bind the protein ligand.
33
All G-proteins are trimeric, i.e. composed of three protein subunits (\_\_, \_\_, and \_\_), w __ and __ subunits _________ (non/covalently) tethered/anchored to pmem by short lipid tails. In unstim/inactivated state (w/o EC signal bound to GPCR) → α subunit/G protein are both ______ (in/active), i.e. α subunit has ___ (GDP/GTP) bound, and G protein is idle.
All composed of three protein subunits (α, β, and γ), w α and γ subunits **covalently** tethered/anchored to pmem by short lipid tails. In unstim/inactivated state (w/o EC signal bound to GPCR) → α subunit/G protein are both **inactive**, i.e. α subunit has **GDP** bound, and G protein is idle.
* In inactive state, GPCR and G protein may assoc as preformed complex, i.e. not always entirely sep entities.
* Several varieties of G proteins, ea specific to partic receptor/set of receptors and for partic set of target enzymes or ion channels in pmem, but all have similar general struc/mechanism of operation.
34
In stim/activated state (EC signal bound to GPCR) → receptor activates G proteins by encouraging __ subunit to expel its ___ and pick up \_\_\_.
In stim/activated state (EC signal bound to GPCR) → receptor activates G proteins by encouraging **α** subunit to expel its **GDP** and pick up **GTP**.
* GPCR activated (conform change) → α subunit ↓ affinity for GDP and ↑ affinity for GTP (exchanges GDP for GTP) → triggers conform change that *activates both* α subunit (clutching its GTP) *and* βγ complex → activated α subunit and βγ complex both dissoc fr GPCR → can then ea interact directly w target proteins in pmem → relay signal to other IC destinations.
* Note: IC [GTP] \>\> IC [GDP], so ↓ α subunit's affinity for GDP quickly results in GTP binding.
35
Stimulation of GPCRs activates _______ subunits.
Stimulation of GPCRs activates **G-protein** subunits.
36
The receptor stays active as long as the EC signal is bound → can catalyze activation of many G proteins. Similarly, the longer the *target protein* remains bound to α (and βγ) subunit → more prolonged relayed signal. What causes the target protein to dissoc and inactivate the α subunit? How is the βγ-complex inactivated?
**α subunit inactivates itself by hydrolyzing its bound GTP → mediates duration of signals relayed by *both* α and βγ subunits**.
* Normally, α subunit has **intrinsic GTPase activity** → hydrolyzes its bound GTP (to GDP) w/i seconds → inactivates α subunit → α subunit dissocs fr target protein and *reassoc* w βγ complex (assuming it dissoc during initial activation) → reform inactive G-protein → inactive G-protein now ready to bind another activated GPCR.
* Note: βγ complex can bind diff target proteins than α subunit, but inactive α subunit binds more favorably w βγ complex, thus βγ complex releases its target protein to reform inactivated G protein.
39
GPCRs remain active for as long as the EC signal is bound. What does this suggest about its enzymatic activity?
The receptor stays active as long as the EC signal is bound → **can catalyze activation of *many* G proteins.**
40
Mammalian heart rate (HR) is controlled by two sets of nerves: one ↑ HR, other ↓ HR. The nerves that signal ↓ HR do so by releasing ACh. Describe how Gi (inhibits adenylyl cyclase) directly couples receptor activation to opening of K+ channels in pmem of heart pacemaker cells.
* ACh binds specific GPCR on heart 'pacemaker' cells → activation of specific G protein—Gi, inhibits adenylyl cyclase.
* Activated **βγ-complex** binds IC face/opens K+ channel in pmem → K+ efflux → inhibitory against APs, i.e. heart cell is harder to activate → HR ↓.
* Original signal terminated → **α subunit** inactivates via intrinsic GTPase (hydrolysis of bound GTP) → Gi returns to inactive state and unbinds/closes K+ channel.
41
Many G proteins activate mem-bound enzymes that produce small "second" messenger molecules, wh is less rapid but more complex than directly binding channels. What are the two most freq target enzymes for G proteins?
Two most freq target enzymes for G proteins are **adenylyl cyclase** and **phospholipase C**.
* **Adenylyl cyclase** - synths cyclic AMP (cAMP).
* **plipase C** - synths inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG); IP3, in turn, promotes accumulation of cytosolic Ca2+.
* cAMP, IP3, DAG, and Ca2+ are all small/second messengers; "first" messenger is EC signal.
* Once activated, enzymes generate large qty of second messengers → rapidly diffuse away fr source → amplify and spread IC signal → bind specific IC signaling proteins and influence their activity.
43
Most commonly, activated G-protein α subunit switches on (excites/stims) adenylyl cyclase (hence, Gs) → dramatic and sudden ↑ in synth cAMP fr ATP, wh is always present in cell. Describe how cAMP is synthesized and degraded.
Adenylyl cyclase catalyzes cAMP fr ATP and phosphodiesterase (PDE) degrades cAMP back to AMP.
* cAMP is formed fr ATP by a cyclization rxn that removes two P groups fr ATP and joins the 'free' end of remaining P group to sugar part of AMP (shown in red).
* PDE rapidly breaks this new bond to reform AMP.
* E.g. one way that caffeine acts as a stimulant is by inhibiting cAMP PDE in nervous sys → blocks cAMP degradation → IC [cAMP] remains high.
44
T/F: cAMP PDE is continuously active inside cell.
True
cAMP PDE is continuously active inside cell → eliminates cAMP v quickly → IC [cAMP] can change rapidly in response to EC signals, rising/falling tenfold in matter of seconds .
45
T/F: cAMP is water-soluble.
True
cAMP is water-soluble → can carry signal thru/o cell → interact w proteins in cytosol, nucleus, or on other organelles (mem proteins).
46
EC signal binds/activates GPCR → activates Gs → (adenylyl cyclase synths cAMP fr ATP) → IC [cAMP] ↑. From here, cAMP exerts most of its effects by activating \_\_\_\_\_\_\_\_\_\_\_\_\_.
**cAMP exerts most of its effects by activating** ***cAMP-dep protein kinase*** (**PKA**; '**A**' bc deps on c**A**MP binding).
* PKA is typ held *inactive* in a complex w a regulatory protein.
* **…*cAMP binds PKA's* *regulatory* (*inhibitory*) *protein* → PKA changes conform/activates → PKA catalyzes phos** **of partic ser's/thr's on specific IC proteins → affect activity****.**
* Recall: protein kinases (PK\_'s) are one of two classes of switch proteins (other being GTP-binding proteins); most common type of PK's are those that phos ser/thr.
* In diff cell types, diff sets of proteins are available to be phos'd → effects of cAMP vary by target cell.
* Note: always remember that biochem rxns deal in energetic favorability and probabilities, i.e. the dramatic and sudden ↑ IC [cAMP] makes binding PKA's regulatory protein signif more likely.
47
Bacterial toxins like cholera and pertussis (whooping cough) cause disease by altering the activity of G proteins. Describe how modifying the α subunit of partic G proteins can lead to such diseases.
Both cholera and pertussis toxins cause disease by continuously stimulating adenylyl cyclase, but do so through diff mechanisms:
* Cholera - bac multiplies in human intestine → prod cholera toxin (protein) → protein enters cells that line intestine and modifies α subunit of partic G protein (**Gs - *stims* adenylyl cyclase**) → **prevents GTPase activity** (Gs can't hydrolyze its bound GTP), i.e. ***Gs* persists in *active* state** → **continuously *stims* adenylyl cyclase** → prolonged and excessive outflow of Cl– and water into gut → catastrophic diarrhea and dehydration → often leads to death unless urgent steps taken to replace lost ions/water.
* Whooping cough (pertussis) - disease-causing bac colonizes the lung → prods pertussis toxin (protein) → alters α subunit of partic G protein (**Gi - *inhibits* adenylyl cyclase**) → **prevents Gi fr releasing bound GDP**, i.e. **Gi persists in *inactive* state** → no inhibition of **adenylyl cyclase** (i.e. continously stimulated, same as w Gs) → stims coughing.
48
The 'fight or flight' response (activation of sympathetic division of autonomic nervous sys) causes adrenal glands (atop kidneys) to synth/secrete EPI into bloodstream → EPI binds adrenergic receptors (a class of GPCRs) that are present on many types of cells → effects vary b/w cell types, but all help prepare body for sudden action.
Describe the remainder of adrenergic pathway in skeletal muscle cells.
EPI binds/activates adrenergic receptors (GPCRs) on skeletal muscle cells → activates α subunit of Gs → (adenylyl cyclase synths cAMP fr ATP) → ↑ IC [cAMP] → cAMP binds PKA's regulatory (inhibitory) protein → activated **PKA** catalyzes phos of ser/thr on target proteins/enzymes → **simult** **stim of glycogen breakdown** and **inhibition of glycogen synth** → amplifies/maximizes 'free' blood glucose available for anticipated muscular activity.
* PKA activation has **two simult effects**: phos/activates phosphorylase kinase (another enzyme) → activates glycogen phosphorylase (yet another enzyme) → breaks down glycogen *and* inhibits glycogen synth.
* **These rxns occur rapidly bc they don't involve changes in gene xcr or new protein synth**.
49
Effects of ↑ [cAMP] occur more rapidly if they don't req changes in gene xcr or new protein synth (as in skeletal muscle). In slower response pathways, PKA typ p-lates xcr regulators → activate xcr of selected genes; e.g. ↑ [cAMP] in certain neurons in brain control prod of proteins involved in learning/LTM. Describe the xcr regulation pathway via PKA.
EC signal binds/activates GPCR → activates α subunit of Gs → (adenylyl cyclase synths cAMP fr ATP) → ↑ IC [cAMP] → cAMP binds PKA's regulatory/inhibitory protein → PKA changes conform/activates → **PKA moves into nucleus and p-lates specific xcr regulators → stims xcr of whole set of target genes**.
* Controls many processes in cells, fr hormone synth in endocrine cells to prod of proteins involved in long-term memory in brain.
50
Normally, GPCRs activate G proteins by ↓ strength of GDP binding to G protein → rapid dissoc of bound GDP and replaced w GTP, bc GTP is present in cytosol in much higher concen than GDP. What conseqs would result fr a mutation in the α subunit of a G protein that caused its affinity for GDP to ↓ w/o signif changing its affinity for GTP? Compare the effects of this mutation w effects of cholera toxin.
* Mutant G protein would be almost continuously activated, bc GDP would dissoc spont → allows GTP to bind even in absence of an activated GPCR. The consequences for the cell would therefore be similar to those caused by cholera toxin, wh modifies the α subunit of Gs so that it cannot hydrolyze GTP to shut itself off.
* In contrast to cholera toxin, h/e, mutant G protein would not stay permanently activated: it would switch itself off normally, but then it would instantly become activated again as GDP dissociated and GTP re-bound.
51
The inositol plipid pathway operates in almost all euk cells and can regulate many diff effector proteins; involves a partic G protein (\_\_), wh activates mem-bound __________ (instead of adenylyl cyclase) → prods \_\_\_\_\_\_\_\_\_, \_\_\_\_\_\_\_\_\_\_, and \_\_\_\_.
Inositol plipid pathway - operates in almost all euk cells and can regulate many diff effector proteins; involves a partic G protein (**Gq**), wh activates mem-bound **phospholipase C** (instead of adenylyl cyclase) → prods **diacylglycerol** (**DAG**), **inositol 1,4,5-triphosphate** (**IP3**), and **Ca2+** (small/2nd messengers).
52
Target proteins of G-protein subunits are either \_\_\_\_\_\_or _________ in pmem.
Target proteins of G-protein subunits are either **enzymes** or **ion channels** in pmem.
* ~20 diff types of mammalian G proteins, ea activated by partic set of receptors and effect/activate partic set of target proteins.
* EC signal binds specific GPCR → effects activities of a specific subset of possible target proteins in pmem → specific/approp response for that partic signal and cell type.
53
Inositol 1,4,5-triphosphate (IP3) is a water-soluble sugar phosphate that is released into _______ (EC space/cytosol) after ________ hydrolyzes/cleaves it from an inositol plipid in the pmem → binds/opens ____ channels in _____ (plasma/ER mem) → ion _____ (influx/efflux).
Inositol 1,4,5-triphosphate (IP3) is a water-soluble sugar phosphate that is released into **cytosol** after **plipase C** hydrolyzes/cleaves it from an inositol plipid in the pmem → binds/opens **Ca2+** channels in ***ER* mem** → Ca2+ efflux (fr ER lumen into cytosol).
* Recall: Inositol plipid is embedded in cytosolic leaf of pmem w IP3 attached to its cytosolic head. Activated plipase C hydrolyzes the inositol plipid to form free cytosolic IP3 and mem-bound diacylglycerol (DAG).
54
Diacylglycerol (DAG) is the lipid that remains embedded in the cytosolic leaf of pmem after activated plipase C hydrolyzes/cleaves off IP3 → DAG helps recruit ___________ to pmem fr cytosol, wh becomes fully activated upon ___ binding.
Diacylglycerol (DAG) is the lipid that remains embedded in the cytosolic leaf of pmem after activated plipase C hydrolyzes/cleaves off IP3 → DAG helps recruit **protein kinase C (PKC)** to pmem fr cytosol, wh becomes fully activated upon **Ca2+** binding (hence, PK**C**) → PKC p-lates cell-specific set of IC proteins.
* PKC operates on same principle as PKA (cAMP-dep protein kinase), but p-lates diff proteins.
55
Describe the inositol plipid pathway.
EC signal binds/activates GPCR → activates α ***and* βγ** subunits of **Gq** → α *and* βγ subunits *both* activate **plipase C** → hydrolyzes/cleaves inositol plipid in cytosolic monolayer of pmem to form two small messengers → (IP3 releases into cytosol; DAG remains embedded in cytosolic leaf of pmem) → **activates two signaling pathways**:
* **IP3** diffuses thru cytosol → **binds/opens special Ca2+ channels in ER mem** → large Ca2+ efflux fr ER into cytosol (bc large echem grad) → **↑ cytosolic [Ca2+]** →…
* … → **DAG**—still embedded in cytosolic leaf of pmem—**recruits PKC** → **Ca2+ binds and fully activates PKC** → PKC interacts w other target proteins, further propagating the signal.
56
T/F: activation of both α and βγ subunits of Gq is required to activate phospholipase C in the inositol plipid pathway.
True
activation of both α and βγ subunits of Gq is required to activate phospholipase C in the inositol plipid pathway.
57
Excitatory signals transiently open Ca2+ channels in either the plasma or ER mem → Ca2+ efflux (fr ECF to cytosol, OR fr ER lumen to cytosol) → triggers conform changes in cytosolic Ca2+-responsive proteins, such as calmodulin. Describe how calmodulin interacts w CaM-kinases.
**Calmodulin** - most common Ca2+-responsive protein; present in cytosol of all euk cells, incl plants, fungi, and protozoa.
* Structure - dumbbell shape: long α helix conn two globular heads, ea w two Ca2+-binding domains.
* **Ca2+ binds/activates calmodulin** → conform change (α helix 'jackknifes' to surround target) → enables interaction w wide range of target proteins, e.g. CaM-kinases.
* **Ca2+/calmodulin-dep protein kinases** (**CaM-kinases**) - partic imp class of calmodulin targets; activated by binding to calmodulin complexed w Ca2+ → p-late selected proteins.
* E.g. mammalian brain - neuron-specific CaM-kinase is activated by pulses of Ca2+ that occur during neural activity; abundant at synapses → imp role in some forms of learning/memory.
* The same Ca2+ pumps that typ maintain echem grad also help terminate Ca2+ signal.
59
Photoreceptors are unusual in that they ______ (de/hyperpol) in absence of stimuli and ______ (de/hyperpol) in light.
Photoreceptors are unusual in that they **depol** in absence of stimuli (scotopic conditions; darkness) and **hyperpol** in light (photopic conditions), i.e. rods/cones are switched ON (activated) in unstimulated state
60
Rod cells are composed of a nucleus w a synaptic region on one side and inner and outer segment on the other. The outer segment is composed of discs of photoreceptive membrane embedded w rhodopsin—a type of GPCR activated by light, and wh interacts w the G protein transducin (Gt). Transducin (Gt) is also present in cones, but diff α subunit and assoc w cone-opsin instead of rhodopsin. cGMP PDE is activated by transducin and converts cGMP into ordinary GMP, much the same as cAMP PDE degrades cAMP. Recall that in the absence of stimuli (light), the rod cell is *depolarized*.
Describe the signaling pathway thru rhodopsin in response to light.
Rod cell in inactive state (depol'd) in absence of light (scotopic conditions) → light signal (photon) strikes/activates rhodopsin (GPCR) → activates α subunit of transducin (Gt) → activates PDE → (PDE degrades cGMP to GMP) → sharp ↓ IC [cGMP] → cGMP-gated Na+ channels close → cell hyperpols → v-gated Ca2+ channels close → ↓ rate of glutamate (nxmtr) release → (excites ON-bipolar cells → signal xmtd to ganglion, optic nerve, thalamus, and occipital lobe).
In absence of light signal (scotopic conditions), cGMP is continuously synthd in photoreceptor cytosol → cGMP binds/keeps open cation channels in photoreceptor pmem, e.g. cGMP-gated Na+ channels and v-gated Ca2+ channels → ↑ rate of glutamate release → excites OFF-bipolar cells → no AP sent to brain.
61
The light-induced signaling cascade in rod (photoreceptor) cells greatly amplifies the light signal. In low light (scotopic) conditions, amplification is enormous: as few as a dozen photons absorbed in entire retina will cause a perceptible signal to delivered to brain. How is the signaling cascade amplified or adapted in high light (photopic) conditions?
**In high light (photopic) conditions, signaling cascade adapts by stepping down the amplification more than 10,000-fold** → photoreceptors (cones) not overwhelmed and can still register changes in light intensity.
* **Adaptation deps on neg feedback**: e.g. turning on light in dark room → (Ca2+ channels *close* in response to light) → IC [Ca2+] plummets → inhibits Ca2+-dep proteins/enzymes resp for signal amplification.
* Adaptation freq occurs in IC signal paths that respond to EC signals → allow cells to respond to fluctuations in signal concen, regardless of magnitude of initial concen.
63
Many EC signals acting via GPCRs affect activity of adenylyl cyclase and thus alter IC [cAMP]. What's more common, activated α subunits switching ON or OFF adenylyl cyclase?
Most commonly, activated G-protein α subunit switches on (excites/stims) adenylyl cyclase (hence, **Gs**) → dramatic and sudden ↑ in synth cAMP fr ATP, wh is always present in cell.
65
All GPCRs are seven-pass xmem structures. Do all ECRs share a similar structure? How does ECR structure influence its function?
Unlike seven-pass xmem GPCRs, ECRs typ have only **one** **xmem** **segment**, wh spans lipid bilayer as **single α helix**.
* Single α helix is poorly suited to xmt a conform change across bilayer
* Instead, EC signal binds ECR's cytoplasmic domain → IC tails of two ECRs to come t/g in pmem → form dimer and **activate kinase domains**, i.e. *ea* *receptor tail p-lates the other* (e.g. two RTKs p-late specific tyr's on ea/o) → triggers assembly of *transient* IC signaling complex on ECR's cytosolic tails.
66
Typ, an EC signal binding the EC domain of RTK causes two receptors to assoc into a dimer. What happens if the EC signal itself is a dimer?
Typ, EC signal binding EC domain of RTK causes two receptors to assoc into a dimer.
* **If EC signal itself is a dimer → can physically cross-link receptors**.
* Other EC signals induce conform change in RTKs → RTKs dimerize (not shown).
* Either case, dimer formation brings kinase domains of ea cytosolic receptor tail into contact → activates kinases to p-late tail of adj RTK (on several tyr's) → ea newly p-lated tyr serves as specific docking site for diff IC signal protein.
67
Typ, an EC signal binding the EC domain of an RTK causes *two* receptors to dimerize (or cross-link if EC signal is a dimer). Either case, dimer formation brings _______ domains of ea cytosolic receptor _____ (head/tail) into contact → _______ (in/activates) kinases to p-late _____ (head/tail) of adj RTK on several ____ (name AA) → ea newly p-lated ___ (AA) serves as specific __________ for diff IC signal protein.
Typ, an EC signal binding the EC domain of an RTK causes *two* receptors to dimerize (or cross-link if EC signal is a dimer). Either case, dimer formation brings **IC/kinase** domains of ea cytosolic receptor **tail** into contact → **activates** kinases to p-late tail of adj RTK on several **tyrosines** (**tyr****/R**) → ea newly p-lated**tyr**serves as specific**docking site** for diff IC signal protein.
68
All IC docking signals contain a specialized ______________ that recogs/binds specific p-lated ____ (AA) on cytosolic ____ (head/tail) of activated RTK. Some IC signals are activated by __________ upon binding docking site on ECR, wh allows for propagation of the signal. Other IC signals function solely as ________ → couple ECR to other signaling proteins → help build active signaling complex.
All IC docking signals contain a specialized **interaction** **domain** that recogs /binds specific p-lated **tyrosines (tyr/R)** on cytosolic **tail** of activated RTK. Some IC signals are activated by **p-lation** upon binding docking site on ECR, wh allows for propagation of the signal. Other IC signals function solely as **scaffolds** → couple ECR to other signaling proteins → help build active signaling complex.
* Interaction domain may also allow IC signal to recog/bind another IC signal protein or p-lated lipids synthd on cytosolic leaf of pmem.
71
Diff RTKs recruit diff collections of IC signals → diff effects. Most RTKs, h/e, activate _________ and \_\_\_\_.
Diff RTKs recruit diff collections of IC signals → diff effects. Most RTKs, h/e, activate a **plipase** **C** → trigger *inositol* *plipid* *signaling path* (same mechanism as GPCRs), as well as activate **Ras**—a small GTP-binding protein.
* Virtually all RTKs activate Ras, incl **platelet-derived growth factor** (**PDGF**) receptors, wh mediate cell proliferation in wound healing, and **nerve growth factor** (**NGF**) receptors, wh are imp in dev of certain vertebrate neurons.
72
Ras is a small \_\_\_\_-binding protein bound by ______ (sugar/lipid/protein) _____ (head/tail) to ________ (non/cytoplasmic) face of pmem.
Ras is a small **GTP**-binding protein (*monomeric* GTPase) bound by **lipid** **tail** to **cytoplasmic** face of pmem.
* Key member of IC signal complex (formed on activated RTK's cytosolic tail).
Recall (GTPases, GEFs, and GAPs): two classes of molecular switch proteins: protein p-lation (larger class) and GTP-binding. GTP-binding proteins are activated by GTP binding → stims intrinsic GTP-hydrolyzing (GTPase) activity → shut themselves off by hydrolyzing their bound GTP to GDP (+ Pi). Two main types of GTP-binding proteins in IC signal path:
* Trimeric GTP-binding proteins (Trimeric GTPases; "G proteins") - relay messages fr GPCRs.
* **Monomeric GTP-binding proteins (Monomeric GTPases)** - aided by two sets of regulatory proteins:
* Guanine ntide exchange factors (GEFs) - activate switch by promoting exchange of GDP for GTP.
* GTPase-activating proteins (GAPs) - inactivate switch by promoting GTP hydrolysis.
* Caution: GAPs are *GTPase*-activating, i.e. stim GTPase activity, wh *hydrolyzes GTP to GDP* and *inactivates* to assoc switch protein.
73
Ras resembles α subunit of a G protein → functions as molecular switch in v similar way. Explain.
**Ras has *two distinct conform states*: active when GTP bound, inactive when GDP bound**.
* **Ras guanine nucleotide exchange factor** (**Ras-GEF**) - *activating* protein → stims Ras to exchange GDP for GTP → Ras switches to activated state → can now stim several downstream signaling paths.
* **Ras GTPase-activating protein** (**Ras-GAP**) - *deactivating* protein → (after some post-activation delay) promotes hydrolysis of GTP to GDP → Ras switches to inactivated state.
* Caution: *GTPase*-activating, i.e. stims GTPase activity wh inactivates switch (Ras)
**Summary**: adaptor protein docks on partic phosphotyrosine on activated receptor → adaptor recruits Ras-GEF that stims Ras to exchange bound GDP for GTP → Ras activated → Ras stims several downstream signaling paths → Ras-GAP hydrolyzes bound GTP for GDP → Ras inactivated.
74
In active state (\_\_\_ bound via Ras-GEF), Ras initiates phos cascade → _______ phos/activate one/an in seq → carries signal fr pmem to \_\_\_\_\_\_, incl three-protein-kinase module: __________ signaling module, so named bc of final kinase in chain.
In active state (**GTP** bound via Ras-GEF), Ras initiates phos cascade → **ser/****thr****protein kinases**phosactivate one/an in seq → carries signal fr pmem to**nucleus**, incl three-protein-kinase module:**MAP-kinase signaling module**, so named bc of final kinase in chain—**mitogen-activated protein kinase**(**MAP kinase**).
* **Mitogens are EC signals that** **stim** **cell prolif**.
* In MAP-kinase pathway, Ras activates MAP kinase kinase kinase → activates MAP kinase kinase → activates MAP kinase → phos's various effector proteins, incl certain xcr regulators.
* Dep on wh other genes are active and other signals, changes to gene expr may stim cell prolif, promote cell survival, or induce cell differentiation.
75
In active state (GTP bound via Ras-GEF), Ras initiates p-lation cascade → ser/thr protein kinases p-late/activate one/an in seq → carries signal fr pmem to nucleus, incl the MAP-kinase signaling module, in wh Ras activates ____________ → p-lates/activates ____________ → p-lates/activates ____________ → p-lates various _______ proteins, incl certain xcr regulators.
In MAP-kinase pathway, Ras activates **MAP kinase kinase kinase** → p-lates/activates **MAP kinase kinase** → p-lates/activates **MAP kinase** → p-lates various **effector** proteins, incl certain xcr regulators.
* Mitogen-activated protein kinase (MAP kinase).
* Mitogens are EC signals that stim cell proliferation.
* Dep on wh other genes are active and other signals, changes to gene expr may stim cell proliferation, promote cell survival, or induce cell differentiation.
76
~30% of human cancers contain a Ras mutation. Based on the typ characteristics of cancer, would such a mutation more likely involve a continuously active or inactive GTPase mechanism?
Mutated Ras typ has GTPase *inactivated* → can't shut itself off → promotes uncontrolled cell proliferation and dev of cancer.
77
T/F: Most signaling pathways dep on proteins that physically interact w one/an.
True
Most signaling pathways dep on proteins that physically interact w one/an.
Several ways to detect such direct contact, e.g. using a protein as “bait” to isolate a receptor that binds partic hormone:
* E.g. insulin - one could attach insulin to a chromatography column. Cells that respond to insulin are broken open and their contents poured over the column. Proteins that bind to insulin will stick to this column and can later be eluted and identified.
**Co-immunoprecipitation** - identify protein-protein interactions.
* Cells exposed to target EC signal can be broken open, and antibodies used to grab receptor known to recog signal.
* If receptor is *strongly* assoc w other proteins, these will be captured as well. Thus, researchers can identify wh proteins interact when an EC signal stims cells.
Once two proteins are known to bind ea/o, recombinant DNA tech used to pinpoint wh parts of proteins are reqd for interaction.
* E.g. to det wh p-lated tyr on RTK a certain IC signal binds, a series of mutant receptors is constructed, ea missing a diff tyr fr its cytoplasmic domain.
Ultimately, one wants to assess protein's role in signaling path. A first test may involve using recombinant DNA tech to introduce a gene encoding a constantly active form of the protein into cell to see if this mimics effect of EC signals, e.g. intro mutant/constantly active form of Ras. Alt, a specific signal protein can be *inactivated:*
* E.g. “knock down” Ras gene in cells by RNA interference. If cell doesn't proliferate in response to EC mitogens, indicates importance of *normal* Ras signaling in proliferative response.
78
Which signaling pathway do RTKs activate to produce lipid docking sites in the pmem?
RTKs activate the **PI-3-kinase-Akt signal path** to produce lipid docking sites in the pmem.
* **Phosphoinositide 3-kinase** (**PI 3-kinase**) - phos inositol plipids in pmem → plipids serve as docking sites for specific IC signals (e.g. **Akt**), wh relocate fr cytosol to pmem → activate one/an.
* Crucial path used by RTKs to promote cell growth/survival.
* **Akt** (also '**protein kinase B**', **PKB**) - ser/thr protein kinase; indirectly promotes growth/survival of many cell types, typ by *inactivating* signal proteins that promote cell death (e.g. **Bad**).
79
Phosphoinositide 3-kinase (PI 3-kinase) activate ____________ in pmem, wh then serve as docking sites for specific IC signals, such as \_\_\_\_\_\_, wh relocate fr cytosol to pmem → activate one/an.
Phosphoinositide 3-kinase (PI 3-kinase) activate **inositol plipids** in pmem, wh then serve as docking sites for specific IC signals, such as **Akt** (**PKB**), wh relocate fr cytosol to pmem → activate one/an.
* Crucial path used by RTKs to promote cell growth/survival.
* phos'd plipid also recruits PK1, wh, in addition to PK2, are reqd to activate Akt (PKB).
80
Akt, also called \_\_\_\_\_\_\_\_\_\_\_\_\_, is a ser/ thr protein kinase that is recruited to docking sites on pmem (inositol plipids activated by \_\_\_\_\_\_) and indirectly promotes ____________ of many cell types, typ by inactivating other proteins, such as \_\_\_\_.
Akt (Akt kinase), also called **protein kinase B (PKB)**, is a ser/thr protein kinase that is recruited to docking sites on pmem (inositol plipids activated by **PI 3-kinase**) and indirectly promotes **growth/survival** of many cell types, typ by inactivating other proteins, such as **Bad**, wh promote cell death.
* Akt is recruited to docking sites on pmem and activated by phos via PK1 and PK2; PK1 is also recruited lipid docking sites on pmem.
* Bad - cytosolic protein *inactivated* by Akt kinase; in *active* state, promotes apoptosis by binding/inhibiting Bcl2 (protein), wh otherwise suppresses apoptosis. Akt inactivates Bad → Bad releases Bcl2 → Bcl2 suppresses apoptosis, therefore Akt kinase indirectly promotes cell survival/growth.
81
Bad is a cytosolic protein inactivated by Akt. In its active state, Bad promotes apoptosis by binding/inhibiting Bcl2 (protein), wh otherwise suppresses apoptosis.
Therefore, what effect does Akt have on cell survival/growth?
Akt inactivates Bad → Bad releases Bcl2 → Bcl2 suppresses apoptosis, therefore **Akt indirectly promotes cell survival/growth**.
82
Tor (target of rapamycin) is a large ser/thr kinase that stims cell growth. Briefly describe the mechanism and its assoc w cancer.
**Tor** (**target of rapamycin**) is a large ser/thr kinase; stims cell growth by **enhancing protein synth and inhibiting protein degradation**; *indirectly* activated by PI 3-kinase-Akt path.
* Rapamycin - anticancer drug that inactivates Tor. I.e. blocks cell growth.
83
A cell's survival mechanism involves an EC survival signal, such as \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_, activates ____ → recruits/activates ___________ → p-lates an inositol plipid embedded in cytosolic leaf of pmem → p-lated inositol plipid attracts and activates IC signal proteins, such as \_\_\_\_, wh bear a special recog domain → activated IC signal ________ (releases fr/embeds in) pmem and p-lates various downstream proteins on specific ser's/thr's.
A cell's survival mechanism involves an EC survival signal, such as **insulin-like growth factor (IGF)**, activates **RTK** → recruits/activates **PI 3-kinase** → p-lates an inositol plipid embedded in cytosolic leaf of pmem → p-lated inositol plipid attracts and activates IC signal proteins, such as **Akt kinase**, wh bear a special recog domain → activated Akt kinase **releases fr** pmem and p-lates various downstream proteins on specific ser's/thr's.
* Protein kinase 1 is also recruited to p-lated inositol plipid on pmem.
* Both PK1 and cytosolic PK2 p-late/activate Akt, wh then releases fr pmem...
84
In a cell's *growth* pathway, an EC signal (e.g. growth factor) binds to RTK → activates ____________ signaling path → ___ indirectly activates Tor by p-lating/inhibiting a protein that helps suppress Tor → activated Tor stims protein _____ and inhibits protein _______ by p-lating key proteins in these processes.
In a cell's *growth* pathway, an EC signal (e.g. GF) binds to RTK → activates **PI 3-kinase-Akt** signal path → **Akt kinase** indirectly activates Tor (also a kinase) by inhibiting a protein that helps suppress Tor → activated Tor stims protein **synth** *and* inhibits protein **degradation**.
85
Would you expect to activate RTKs by exposing exterior of cells to antibodies that bind to assoc proteins? Would your answer be diff for GPCRs?
* Bc ea antibody has two antigen-binding sites, it can cross-link receptors and cause them to cluster on cell surface → likely to activate RTKs, wh are typ activated by dimerization.
* For RTKs, clustering allows individual kinase domains of receptors to p-late adj receptors in the cluster.
* Activation of GPCRs is more complicated, bc the ligand has to induce a partic conform change; only v special antibodies mimic receptor ligands sufficiently well to induce conform change that activates a GPCR.
86
\_\_\_\_\_\_ is a receptor protein that directly acts as xcr regulator; activated by binding of ______ → cleaves cytosolic tail → receptor is free to move to nucleus, whr it helps activate approp set of receptor-responsive genes.
**Notch** is a receptor protein that directly acts as xcr regulator; activated by binding of **Delta** → Delta cleaves Notch's cytosolic tail → Notch's tail (receptor) migrates to nucleus, thru pore, and helps activate approp set of Notch-responsive genes.
* **Delta** - xmem signal protein on surface of adj cell; binds Notch (receptor on adj cell) and cleaves it fr pmem.
* Crucially imp in all animals, both during dev and in adulthood.
87
Describe the interaction b/w Delta and Notch.
Delta, a xmem signal protein, binds/activates Notch, a receptor on adj cell. Delta also cleaves Notch's cytosolic tail, wh releases the tail fr pmem and into cytosol → free Notch tail migrates to nucleus, thru pore, and directly activates Notch-responsive genes.
88
Compare the major GPCR and ECR signaling paths.
See figure.
90
T/F: cAMP-dep protein kinase (PKA) is typ held inactive in a complex w a regulatory protein.
True
cAMP-dep protein kinase (PKA) is typ held inactive in a complex w a regulatory protein.
* **cAMP binds PKA's** ***regulatory protein*** → PKA changes conform/activates → PKA catalyzes phosphorylation of partic serines (ser/S) or threonines (thr/T) on specific IC proteins → affect activity of these target proteins.
93
Explain why cAMP must be broken down rapidly in a cell to allow rapid signaling.
Rapid breakdown keeps IC [cAMP] low: the lower cAMP levels are, the larger/faster the ↑ achieved upon activation of adenylyl cyclase, wh synths new cAMP (fr ATP).
* Recall: cAMP PDE is continuously active in cell → eliminates cAMP v quickly (cAMP → AMP, *not ATP*) → IC [cAMP] can change rapidly in response to EC signals, rising/falling tenfold in matter of seconds.
95
Inositol phospholipids are plipids w inositol—a ______ (sugar/lipid/protein)—attached to its ______ (non/cytosolic) ______ (head/tail); present in small qty in ________ (non/cytosolic) leaf of pmem.
Inositol phospholipids are plipids w inositol (IP3)—a **sugar**—attached to its **cytosolic head**; present in small qty in **cytosolic** leaf of pmem.
101
Why do you suppose cells have evolved IC Ca2+ stores for signaling even though there is abundant EC Ca2+?
Recall (Ch.15) that the pmem constitutes a rather small area compared w total mem surfaces in a cell. The ER is especially abundant and spans entire volume of cell as a vast network of mem tubes/sheets. The Ca2+ stored in the ER can therefore be released thru/o cytosol, wh is imp bc rapid clearing of Ca2+ ions fr cytosol by Ca2+ pumps prevents Ca2+ fr diffusing any signif distance in cytosol.
105
Enzyme-coupled receptors don't assoc w a G protein, like GPCRs. Instead, their cytoplasmic domain either acts as a(n) _______ itself or forms complex w another protein that acts as a(n) \_\_\_\_\_\_\_.
Enzyme-coupled receptors don't assoc w a G protein, like GPCRs. Instead, their cytoplasmic domain either acts as an **enzyme** itself or forms complex w another protein that acts as an **enzyme**.
106
Most growth factors act as _______ (local/widespread) mediators and can act at v low concens. ECR response to such signals is typ _____ (slow/fast).
Most growth factors act as **local** mediators and can act at v low concens (~10–9 to 10–11 M). ECR response to such signals is typ **slow** (hours); effects may req many IC xdctn steps that typ lead to change in gene expr.
107
Receptor tyrosine (tyr/R) kinases (RTKs) are the largest class of ECRs. Its ________ (non/cytoplasmic) domain functions as ______________ → ____________ (de/phosphorylates) partic tyr's on specific IC signaling proteins.
**Receptor tyrosine (****tyr****/R) kinases** (**RTKs**) are the largest class of ECRs. Its **cytoplasmic** domain functions as **an enzyme (tyrosine protein kinase)** → **phosphorylates** partic tyr's on specific IC signaling proteins.
* Abnormalities in signaling via RTKs (and other ECRs) have major role in dev of most cancers.
112
Docked IC signals can xmt signal along several routes simult to many destinations in cell → activate/coord many biochem changes reqd to trigger complex response, e.g. cell proliferation or differentiation. To terminate response, p-lated tyr's are *dephosphorylated* by \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_.
Docked IC signals can xmt signal along several routes simult to many destinations in cell → activate/coord many biochem changes reqd to trigger complex response, e.g. cell proliferation or differentiation. To terminate response, p-lated tyr's are dephosphorylated by **protein** **tyr** **phosphatases**.
113
T/F: In some cases, activated RTKs are dragged into interior of cell by endocytosis → destroyed by digestion in lysosomes.
True
In some cases, activated RTKs (as well as some GPCRs) are dragged into interior of cell by endocytosis → destroyed by digestion in lysosomes.
114
Diff IC signaling pathways interact, enabling ea cell type to produce the appropriate response to a combination of EC signals. In the absence of such signals, most animal cells have been programmed to respond how?
Diff IC signaling pathways interact, enabling ea cell type to produce the appropriate response to a combination of EC signals. **In the absence of such signals, most animal cells have been programmed to kill themselves by undergoing apoptosis**.
115
If some cell-surface receptors, including Notch, can rapidly signal to the nucleus by activating latent transcription regulators at the plasma membrane, why do most EC receptors use long, indirect signaling cascades to influence gene transcription in the nucleus?
The more steps there are in an IC signaling pathway, the more places the cell has to regulate the pathway, amplify the signal, integrate signals fr diff pathways, and spread the signal along divergent paths
116
T/F: The EC signal molecule acetylcholine has diff effects on diff cell types in an animal and often binds to diff cell-surface receptor molecules on diff cell types.
True.
ACh, for example, slows the beating of heart muscle cells by binding to a GPCR and stimulates the contraction of skeletal muscle cells by binding to a different acetylcholine receptor, which is an ion-channel-coupled receptor.
117
T/F: After ACh is secreted from cells, it is long-lived, bc it has to reach target cells all over the body.
False
ACh is *short-lived* and exerts its effects *locally*. Indeed, the conseqs of prolonging its lifetime can be disastrous. Compounds that inhibit the enzyme acetylcholinesterase, wh normally breaks down ACh at a nerve–muscle synapse, are extremely toxic: for example, the nerve gas sarin, used in chemical warfare, is an acetylcholinesterase inhibitor
118
T/F: Both the GTP-bound α subunits and nucleotide-free βγ complexes—but not GDP-bound, fully assembled G proteins—can activate other molecules downstream of GPCRs.
True
Nucleotide-free βγ complexes can activate ion channels, and GTP-bound α subunits can activate enzymes. The GDP-bound form of trimeric G proteins is the inactive state.
119
T/F: IP3 is produced directly by cleavage of an inositol plipid w/o incorporation of an additional phosphate group,
True
The inositol plipid that is cleaved to produce IP3 contains three phosphate groups, one of which links the sugar to the diacylglycerol lipid. IP3 is generated by a simple hydrolysis rxn.
120
T/F: Calmodulin regulates IC [Ca2+].
False
Calmodulin recogs/binds/deps on Ca2+ for activation, but does not regulate IC [Ca2+] thereafter.
* Ca2+ binds/activates calmodulin → conform change (α helix 'jackknifes' to surround target) → enables interaction w wide range of target proteins, e.g. CaM-kinases.
* Ca2+/calmodulin-dep protein kinases (**CaM-kinases**) - partic imp class of calmodulin targets; activated by binding to calmodulin complexed w Ca2+ → p-late selected proteins.
* Imp in learning/memory and heart cells.
121
T/F: Tyrosine phosphorylation serves to build binding sites for other proteins to bind to RTKs
True
Tyrosine phosphorylation serves to build binding sites for other proteins to bind to RTKs
122
The Ras protein functions as a molecular switch that is set to its “on” state by other proteins that cause it to expel its bound GDP and bind GTP. A GTPase-activating protein helps reset the switch to the “off” state by inducing Ras to hydrolyze its bound GTP to GDP much more rapidly than it would without this encouragement. Thus, Ras works like a light switch that one person turns on and another turns off.
You are given a mutant cell that lacks the GTPase-activating protein. What abnormalities would you expect to find in the way in which Ras activity responds to EC signals?
You would expect a high background level of Ras activity, bc Ras cannot be turned off efficiently.
* Ras is already GTP-bound, so Ras activity in response to EC signal would be greater than normal, but this activity would be liable to saturate when all Ras are converted to the GTP-bound form.
* The response to a signal would be much less rapid, bc signal-dep increase in GTP-bound Ras would occur over an elevated background of preexisting GTP-bound Ras.
* The increase in Ras activity in response to a signal would also be prolonged compared to the response in normal cells.
123
Compare and contrast signaling by neurons, wh secrete nxmtrs at synapses, w signaling carried out by endocrine cells, wh secrete hormones into the blood.
Both types of signaling can occur over a long range: neurons can send action potentials along very long axons (think of the axons in the neck of a giraffe, for example), and hormones are carried via the bloodstream thru/o the organism.
* Bc neurons secrete large amounts of nxmtrs at a synapse, the concens of these signals are high; nxmtr receptors, therefore, need to bind to nxmtrs w only low affinity. Hormones, in contrast, are vastly diluted in the bloodstream, where they circulate at often minuscule concens; hormone receptors therefore generally bind their hormone w extremely high affinity.
* Whereas neuronal signaling is a private affair, w one neuron talking to a select group of target cells thru specific synaptic conns, endocrine signaling is a public announcement, w any target cell w approp receptors able to respond to the hormone in the blood. Neuronal signaling is very fast, limited only by the speed of propagation of the AP and the workings of the synapse, whereas endocrine signaling is slower, limited by blood flow and diffusion over larger distances.
124
In a series of experiments, genes that code for mutant forms of an RTK are introduced into cells. The cells also express their own normal form of the receptor from their normal gene, although the mutant genes are constructed so that the mutant RTK is expressed at considerably higher concentration than the normal RTK.
What would be the conseqs of introducing a mutant gene that codes for an RTK (A) lacking its EC domain, or (B) lacking its IC domain?
The mutant RTK lacking its EC ligand-binding domain is inactive. It cannot bind EC signals, and its presence has no conseqs for the function of the normal RTK.
The mutant RTK lacking its IC domain is also inactive, but its presence will block signaling by the normal receptors. When a signal molecule binds to either receptor, it will induce their dimerization. Two normal receptors have to come t/g to activate ea/o by phosphorylation. In the presence of an excess of mutant receptors, h/e, normal receptors will typ form mixed dimers, in wh their IC domain cannot be activated bc their partner is a mutant and lacks a kinase domain.
125
What are the similarities and diffs b/w rxns that lead to the activation of G proteins and the rxns that lead to the activation of Ras?
Activation in both cases dep on proteins that catalyze GDP–GTP exchange on the G protein or Ras protein.
* Activated GPCRs perform this function directly for G proteins.
* By contrast, activated ECRs (tyr's p-lated) assemble multiple signaling proteins into a signaling complex; one of these is an **adaptor protein** that recruits a GEF that fulfills this function for Ras.
126
Why do you suppose cells use Ca2+ (which is kept by Ca2+ pumps at a cytosolic concentration of 10–7 M) for intracellular signaling and not another ion such as Na+ (which is kept by the Na+ pump at a cytosolic concentration of 10–3 M)?
Bc IC [Ca2+] is so low, an influx of relatively few Ca2+ ions leads to large changes in its cytosolic concentration. Thus, a tenfold increase in cytosolic Ca2+ can be achieved by raising its concen into the micromolar range, wh would req far fewer ions than would be required to change significantly the cytosolic concen of a more abundant ion such as Na+.
In muscle, a greater than tenfold change in cytosolic Ca2+ concen can be achieved in microseconds by releasing Ca2+ fr SR, a task that would be difficult to accomplish if changes in the millimolar range were required.
127
It seems counterintuitive that a cell, having a perfectly abundant supply of nutrients available, would commit suicide if not constantly stimulated by signals from other cells. What do you suppose might be the advantages of such regulation?
In a multicellular organism such as an animal, it is imp that cells survive only when and where they are needed. Having cells dep on signals fr other cells may be a simple way of ensuring this. A misplaced cell, for example, would probably fail to get the survival signals it needs (as its neighbors would be inappropriate) and would therefore kill itself.
This strategy can also help regulate cell numbers: if cell type A depends on a survival signal from cell type B, the number of B cells could control the number of A cells by making a limited amount of the survival signal, so that only a certain number of A cells could survive. There is indeed evidence that such a mechanism does operate to help regulate cell numbers—in both developing and adult tissues
128
The contraction of the myosin–actin system in muscle cells is triggered by a rise in IC Ca2+. Muscle cells have specialized Ca2+ channels—called ryanodine receptors bc of their sensitivity to the drug ryanodine—embedded in SR mem. In contrast to the IP3-gated Ca2+ channels in ER mem, the signaling molecule that opens ryanodine receptors is Ca2+ itself. Discuss the conseqs of ryanodine channels for muscle cell contraction.
Ca2+-activated Ca2+ channels create a positive feedback loop: the more Ca2+ that is released, the more Ca2+ channels open. The Ca2+ signal in the cytosol is therefore propagated explosively thru/o the entire muscle cell, thereby ensuring that all myosin–actin filaments contract almost synchronously.
129
Two protein kinases, K1 and K2, function sequentially in an IC signal path. If either kinase contains a mutation that permanently inactivates its function, no response is seen in cells when an EC signal is received. A diff mutation in K1 makes it permanently active, so that in cells containing that mutation a response is observed even in the absence of an EC signal. You characterize a double-mutant cell that contains K2 with the inactivating mutation and K1 with the activating mutation. You observe that the response is seen even in the absence of an EC signal.
In the normal signaling pathway, does K1 activate K2 or does K2 activate K1?
K2 activates K1. If K1 is permanently activated, a response is observed regardless of the status of K2. If the order were reversed, K1 would need to activate K2, which cannot occur because in our example K2 contains an inactivating mutation.
130
A. Trace the steps of a long and indirect signaling pathway from a cell-surface receptor to a change in gene expression in the nucleus.
B. Compare this pathway with two short and direct pathways from the cell surface to the nucleus.
A. Three examples of extended signaling pathways to the nucleus are (1) extracellular signal → RTK → adaptor protein → Ras-activating protein → MAP kinase kinase kinase → MAP kinase kinase → MAP kinase → transcription regulator; (2) extracellular signal → GPCR → G protein → phospholipase C → IP3 → Ca2+ → calmodulin → CaM-kinase → transcription regulator; (3) extracellular signal → GPCR → G protein → adenylyl cyclase → cyclic AMP → PKA → transcription regulator.
B. An example of a direct signaling pathway to the nucleus is Delta → Notch → cleaved Notch tail → transcription.
131
How does PI 3-kinase activate the Akt kinase after activation of RTK?
When PI 3-kinase is activated by an activated RTK, it phosphorylates a specific inositol phospholipid in the plasma membrane. The resulting phosphorylated inositol plipid then recruits to the pmem both Akt and another protein kinase (PK1) that helps phosphorylate and activate Akt. A third kinase (PK2) that is permanently assoc w the pmem also helps activate Akt.
Cell Biology (BIO 301)
flashcards
Decks in class (15)
# Cards
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Ch.4 - Protein Struc/Func73
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Ch.7, Part 1 - DNA to RNA42
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Ch.1/2 - Intro & Chem Components of Cells24
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Ch.11 - Membrane Structure50
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Ch.15, Part 1 - Cell Compartments54
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Endomem Sys & Protein Sorting (BIO201 Review)42
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Ch.7, Part 2 - RNA to Protein38
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Ch.12, Part 1 - Membrane Transport58
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Ch.12, Part 2 - Channels & Electrical Properties of Membranes73
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Ch.15, Part 2 - Cell Compartments78
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Ch.16 - Cell Signaling106
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Ch.17 - Cytoskeleton98
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Ch.18 - Cell Division Cycle97
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Ch.19 - Sex Reprod & Genetics17
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Ch.20 - Tissues, Stem Cells, Cancer106
