Block 2 Presentation 2 Transduction part 2

Question 1

Signaling through enzyme-coupled cell-surface receptors

Answer 1

Signaling through enzyme-coupled cell-surface receptors

Receptors that are kinases or that bind kinases. The kinase domains are shown in red, and the phosphorylation sites are indicated with red arrows.
A: Tyrosine kinase receptors.
B: JAK-STAT receptors.
C: Serine–threonine kinase receptors.

(pic is located on slide 5 of part 2)

Question 2

Epidermal growth factor (EGF)

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Answer 2

Epidermal growth factor (EGF) is one of the Receptor Tyrosine Kinases

Signal protein: Epidermal growth factor (EGF)

receptor: EGF receptors

Responses: stimulates cell survival, growth, proliferation, or differentiation of various cell types; acts as inductive signal in development

Question 3

Receptor Tyrosine Kinases

Answer 3

Receptor Tyrosine Kinases

• The structure of receptor tyrosine kinases
  
  • Single transmembrane polypeptides
  • Extracellular ligand binding domain
  • Tyrosines on cytosolic tail of receptor

receptor tyrosine kinases phosphorylate tyrosines. they phosphorylate themselves (autophosphorylate) on the cytoplasmic tail.

Additional information (not required)

Figure 24-4 RECEPTOR TYROSINE KINASES. Domain architecture of nine of the 20 families of receptor (R) tyrosine kinases, with ribbon models of several domains. The globular domain of the EphB2 receptor is a β sandwich with a ligand-binding site that includes the exposed loop on the front of this model (PDB file: 1IGY). The extracellular part of the insulin-like growth factor consists of two similar β-helical domains connected by cysteine-rich domains (PDB file: 1IGR). The cytoplasmic kinase domain from the insulin receptor is similar to most known kinases (PDB file: 1IRK). Kinase inserts and C-terminal extensions contain tyrosine phosphorylation sites. Receptor names: EphR, receptor for ephrin, membrane-bound ligands in the nervous system, the largest class of receptor tyrosine kinases; PDGFR, platelet-derived growth factor receptor; FGFR, fibroblast growth factor receptor; VEGFR, vascular endothelial growth factor; Met, receptor for hepatocyte growth factor; TrkA, receptor for nerve growth factor; RET, a cadherin adhesion receptor; Axl, receptor for the growth factor Gas6; EGFR, epidermal growth factor receptor. Domain names: Ig, immunoglobulin; F3, fibronectin-III; CAD, cadherin..)

Question 4

Achondroplasia

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Answer 4

Example of a dominant negative disorder: Achondroplasia

results from mutation of a receptor tyrosine kinase

it is autosomal dominate disorder.

• Achondroplasia also known as short-limbed dwarfism; charaterized by small stature with short limb, large head, low nasal bridge, prominent forehead, lumber lordosis.
  
  • Incidence is ~ 1 in 10,000
• Pathology
  
  • Failure of cartilage cell proliferation at the epiphyseal plates of the long bones, resulting in failure of longitudinal bone growth, causing short limb.
• Mutation in Fibroblast Growth Factor Receptor 3 (FGFR3) gene;
• ~ 80% of patients are due to new mutation;
• increased risk with late paternal age
• Achondroplasia is a well known, incompletely dominant skeletal disorder of short-limbed dwarfism and large head
• Most achondroplastic individuals have normal intelligence and lead normal lives within their physical capabilities.
• Marriages between two achondroplastic individuals are not uncommon.
• Achondroplasia, an autosomal dominant disorder that often occurs as a new mutation. Note small stature with short limbs, large head, low nasal bridge, prominent forehead, and lumbar lordosis in this typical presentation.

Question 5

Receptor Tyrosine KinaseThree Major Signaling Branches

Answer 5

Receptor Tyrosine KinaseThree Major Signaling Branches

1. Ras/MAPK signaling pathway
2. Phospholipase C-gamma signaling pathway
3. PI 3-Kinase/AKT signaling pathway

to get one of the above branches going, what needs to occur at the beginning of each branch is that you need a protein in that pathway to interact with the receptor complex. so a member of that pathway will need the ability to dock to a phosphotyrosine.

Regulatory proteins with SH2 domains (Src homology 2 domains) recognize and bind to phosphorylated tyrosines on receptor. so proteins with SH2 domains have the ability to bind to phosphorylated tyrosines.

• This binding stimulates regulatory proteins

Question 6

Three types of intracellular signaling complexes associated with receptors

Answer 6

Three types of intracellular signaling complexes associated with receptors

• Assemblies help simplify and localize signaling through receptors
• The assembly of signaling complexes depends on various conserved interaction domains, which are found in many intracellular signaling proteins.

(slide 10 for larger pics)

Question 7

Activation of RTKs

Answer 7

The activation of RTKs

Ligand binds, receptors aggregate, autophophorylation occurs between receptors -Activated receptor tyrosine kinases phosphorylate themselves

Question 8

Phosphorylated tyrosines on receptor tyrosine kinases serve as?

Answer 8

Phosphorylated tyrosines on receptor tyrosine kinases serve as docking sites for intracellular signaling proteins

Proteins with either SH2 (for SRC homology region) domains, or less commonly, PTB domains (for phosphotyrosine-binding) can bind phosphorylated tyrosines

Question 9

SH2 domains vary for specificity with neighboring amino acids

Answer 9

SH2 domains vary for specificity with neighboring amino acids

• The SH2 domain is a compact module.
• Each SH2 domain has distinct sites for recognizing phosphotyrosine and for recognizing a particular amino acid side chain, different SH2 domains recognize phosphotyrosine in the context of different flanking amino acid sequences
• This variation allows receptor tyrosine kinases to bind with specificity, multiple proteins containing SH2 domains

Question 10

SUBUNIT DIMERIZATION MECHANISM FOR ACTIVATING THE EGF RECEPTOR TYROSINE KINASE

Answer 10

SUBUNIT DIMERIZATION MECHANISM FOR ACTIVATING THE EGF RECEPTOR TYROSINE KINASE

• In the absence of EGF, intramolecular interactions preclude dimerization.
• EGF binding changes the conformation of the extracellular domains allowing dimerization of two receptors, bringing together two cytoplasmic kinase domains.
• Transphosphorylation activates both kinases and creates phosphotyrosine binding sites for SH2 and PTB domains of downstream signal transduction proteins.

Question 11

Ras/MAPK Signaling Pathway

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Answer 11

Ras/MAPK Signaling Pathway

1. GRB2 (contains SH2 domain) interacts with tyrosine phosphorylated receptor, localizing it to the membrane
2. Sos is associated with GRB2 and becomes active (- Sos is a guanine-nucleotide exchange factor)
3. Sos stimulates Ras (a monomeric G-protein) to release GDP and bind GTP.
4. Ras is now active
5. Ras activates RAF (a MAPKKK)
6. RAF activates MEK (a MAPKK)
7. MEK activates ERK (a MAPK)
8. MAPKs phosphorylates transcription factors (such as AP-1)
9. Transcription factors induce cells to grow and divide

Ras in inactivated by GTPase activating protein (GAP)

Question 12

Signal transduction by tyrosine kinase receptors.

Answer 12

Signal transduction by tyrosine kinase receptors.

Signal transduction by tyrosine kinase receptors.

(1) Binding and dimerization.
(2) Autophosphorylation.
(3) Binding of Grb2 and SOS.
(4) SOS is a GEF (guanine nucleotide exchange protein) that binds Ras, a monomeric G-protein anchored to the plasma membrane.
(5) GEF activates the exchange of GTP for bound GDP on Ras.
(6) Activated Ras containing GTP binds the target enzyme Raf, thereby activating it, and a series of kinases known as the MAP kinase pathway.

Question 13

Ras is a?

Answer 13

Ras is a monomeric GTPase

• Ras is active when bound to GTP
• It is inactive when bound to GDP
• SOS is a GEF which “turns on” Ras
• GAPs “turn off” RAS
• Active RAS binds to RAF

Question 14

RAS-MAPK Signaling

Answer 14

RAS-MAPK Signaling

• The MAP kinase module activated by Ras
• The three-component module begins with MAP kinase kinase kinase called Raf.
• Ras recruits Raf to the plasma membrane and helps activate it.
• Raf then activates the MAP kinase kinase Mek, which then activates the MAP kinase Erk.
• Erk in turn phosphorylates a variety of downstream proteins, including other protein kinases, as well as gene regulatory proteins in the nucleus.

Question 15

Oncogenes and signal transduction

Answer 15

Oncogenes and signal transduction

Oncogene proteins act as growth factors (e.g., EGF), growth factor receptors (e.g., ErbB), and intracellular signaling molecules (Ras and Raf). Ras and Raf activate the ERK MAP kinase pathway leading to the induction of additional genes (e.g., fos) that encode potentially oncogenic transcriptional regulatory proteins. Proteins with known oncogenic potential are highlighted with a yellow glow.

Question 16

Noonan Syndrome

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Answer 16

Noonan Syndrome results from mutations in Ras/MAPK signaling

• Noonan Syndrome is an autosomal dominant condition which is characterized by short stature, distinctive craniofacial features, congenital cardiovascular disease, and other more variable clinical findings such as developmental delay and intellectual disability, bleeding tendencies, lymphatic abnormalities and genitourinary abnormalities.
• Occurs in ~1 in 1,000 individuals
• Often genes encoding Ras/MAPK signaling proteins are mutated
  
  • PTPN11 (Protein Tyrosine phosphatase Shp2)
  • KRAS
  • SOS
  • RAF1

Also note (in the picture): Excessive nuchal skin/webbed neck is sometimes seen in individuals with Noonan syndrome

A 12-year-old female with Noonan Syndrome. Typical webbed neck. Double structural curve with rib deformity

Question 17

Activation of phospholipase C-g by protein-tyrosine kinases

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Answer 17

Activation of phospholipase C-g by protein-tyrosine kinases

• PLC-γ has SH2 domains that associate with receptor protein-tyrosine kinases.
• Tyrosine phosphorylation increases PLC- γ activity, stimulating hydrolysis of PIP2 to produce InsP3 and DAG.
  
  • InsP3 stimulates calcium release.
  • DAG stimulates PKC pathway

Question 18

EGF RECEPTOR TYROSINE KINASE SIGNALING PATHWAY

Answer 18

EGF RECEPTOR TYROSINE KINASE SIGNALING PATHWAY

1. A, Ligand binding changes the conformation of the extracellular domains of the receptor.
2. B, Extracellular domains dimerize, bringing together the tyrosine kinase domains of two receptor subunits in the cytoplasm. Direct interactions and transphosphorylation activate the kinases and create specific docking sites for effector proteins with SH2 domains.
3. C, Phospholipase C-gamma binds one phosphotyrosine and is activated by phosphorylation to break down PIP2 into diacylglycerol and IP3
4. D, A complex of the adapter protein Grb2 and the nucleotide exchange factor SOS binds another phosphotyrosine. SOS catalyzes the exchange of GDP for GTP on the membrane-associated small GTPase Ras. Ras-GTP attracts the cytoplasmic serine/threonine kinase Raf to the plasma membrane.
5. E, Raf phosphorylates and activates the dual-function kinase MEK.
6. F, MEK phosphorylates and activates MAP kinase.
7. G, MAP kinase enters the nucleus and activates latent transcription factors.

Question 19

NFAT and Cyclosporin A

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Answer 19

NFAT and Cyclosporin A

• Stimulated T cells trigger PLCg resulting in calcium being released
• Calcium binds and activates calmodulin
• Calmodulin binds and activates a phosphatase called Calcineurin
• Calcineurin dephosphorylates NFAT allowing it to go to the nucleus and stimulate IL-2 gene expression.
• Cyclosporin A : important immunosuppressant used to prevent rejection of transplanted organs
• Works by inhibiting Calcineurin

Question 20

Receptor tyrosine kinases can also activates?

Answer 20

Receptor tyrosine kinases can also activates the PI 3 Kinase/AKT Pathway

• Phosphatidylinositol-3-kinase (PI 3-Kinase) binds phosphorylated tyrosines on receptor and becomes stimulated
• Phosphatidylinositol-3-kinase then phosphorylates phospholipid PIP2 in membrane.
• Regulates cell growth, survival and movement

Question 21

PI 3 Kinase

Answer 21

• PI 3-Kinase produces lipid docking sites in the plasma membrane
• PI 3-kinase phosphorylates PI (4,5)P2 (PIP2) into PI(3,4,5) P3 (PIP3)
• Intracellular signaling proteins can interact with PIP3 via a specific interaction domain, such as pleckstrin homology domain (PH).

Question 22

AKT activation

Answer 22

AKT activation

• AKT and the phosphoinositol-dependent kinase (PDK1) bind PIP3
• Another kinase (usually mTOR) phosphorylates AKT, this causes a conformational change that allows PDK1 to phosphorylate AKT
• The activated AKT now dissociates from the plasma membrane and phosphorylates various targets, including the Bad protein.
• Bad is involved with apoptosis.

Question 23

The PI 3-kinase/Akt pathway

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Answer 23

The PI 3-kinase/Akt pathway

1. Ligand binds receptor, receptors aggregate and autophosphorylate each other
2. PI 3 Kinase binds phosphorylated tyrosine on receptor and becomes active
3. PI 3 Kinase phosphorylates PIP2 to become PIP3
4. PDK and AKT (PKB) bind PIP3 molecules
5. PDK and mTORC2 then phosporylate and activate AKT (PKB)
6. AKT then phosphorylates downstream targets

Question 24

The PI 3-kinase pathway and cell survival

Answer 24

The PI 3-kinase pathway and cell survival

Survival factors such as NGF activate receptor protein-tyrosine kinases, leading to activation of PI 3-kinase and formation of PIP3. PIP3 recruits the protein kinase Akt to the plasma membrane where it is activated as a result of phosphorylation by PDK. Akt then appears to phosphorylate a number of proteins that contribute to cell survival. The targets of Akt that have been implicated in suppression of apoptosis include the Bcl-2 family member Bad, caspase-9, several transcription factors, and the protein kinase GSK-3, which affects cell metabolism and protein synthesis.

Question 25

AKT & Forkhead (FOXO)

Answer 25

AKT & Forkhead (FOXO) * Akt phosphorylates several target proteins, transcription factors, and other protein kinases. * Transcription factors include members of the Forkhead or FOXO family. * If growth factors are not present, Akt is not active, and FOXO travels to the nucleus where it stimulates transcription of genes that inhibit cell proliferation, or induce cell death. * Akt phosphorylation of FOXO sequesters it in inactive form.

Question 26

The PTEN tumor suppressor gene acts as?

Answer 26

The PTEN tumor suppressor gene acts as a phospholipid phosphatase * (A) Under normal growth conditions, stimulatory signals from the insulin receptor activate the enzyme phosphoinositide kinase (PI3-kinase), which phosphorylates phosphatidylinositol 4,5-bisphosphate (PIP2) to generate phosphatidylinositol 3,4,5-trisphosphate (PIP3), a lipid signaling molecule. Downstream, PIP3 activates several effectors, including the proto-oncogene product PKB/Akt. * The role of PTEN is to dephosphorylate PIP3, acting as a negative control on PKB/Akt activation. * (B) If a mutation in PTEN renders it unable to carry out its phosphatase function, PIP3 can no longer be deactivated, so continues to propagate its signal downstream. * This may result in the continued activation of PKB/Akt , which, in combination with other factors, could lead to increased cell growth and possible tumor development.

Question 27

Insulin affects several signaling pathways to regulate resting glucose levels ## Footnote

Answer 27

Insulin affects several signaling pathways to regulate resting glucose levels * Insulin stimulates the uptake of glucose into muscle and adipose cells and by stimulating glycogen metabolism. It thereby lowers blood glucose levels * The inability to produce insulin can cause type I diabetes * Type II diabetes appears to result from resistance to insulin, rather than an inability to produce it. * The insulin signaling pathway influences glucose homeostasis by regulating multiple pathways. * The insulin receptor is a multi-subunit receptor tyrosine kinase. * When it binds insulin, recruitment and activation of the IRS1 protein initiates signal transduction, leading to glucose import, stimulation of glycogen synthesis, and regulation of gene expression

Question 28

Insulin-Dependent Diabetes Mellitus (IDDM)(Type I Diabetes)

Answer 28

Insulin-Dependent Diabetes Mellitus (IDDM)(Type I Diabetes) * IDDM is usually caused by autoimmune destruction of islet β cells in the pancreas; this autoimmune reaction is triggered by an unknown mechanism. * The destruction of islet β cells causes insulin deficiency and thereby dysregulation of anabolism and catabolism, resulting in metabolic changes similar to those observed in starvation. * Among North American whites, IDDM is the second most common chronic disease of childhood, increasing in prevalence from 1 in 2500 at 5 years of age to 1 in 300 at 18 years of age. * Loss of insulin reserve occurs during a few to many years. * The earliest sign of abnormality is the development of islet autoantibodies when blood glucose concentrations, glucose tolerance (ability to maintain normal blood glucose levels after ingestion of sugar), and insulin responses to glucose are normal. * This period is followed by a phase of decreased glucose tolerance but normal fasting blood glucose concentration. * With continued loss of β cells, fasting hyperglycemia eventually develops but sufficient insulin is still produced to prevent ketosis; during this period, patients have non-insulin-dependent diabetes mellitus. * Eventually, insulin production falls below a critical threshold, and patients become dependent on exogenous insulin supplements and have a propensity to ketoacidosis.

Question 29

Type II Diabetes Mellitus

Answer 29

Type II Diabetes Mellitus Non-Insulin-Dependent Diabetes Mellitus (NIDDM) * NIDDM results from a combination of genetic susceptibility and environmental factors. * NIDDM usually affects obese individuals in middle age or beyond, although an increasing number of children are becoming affected. * The population risk of NIDDM is 6% to 7% in the United States. * NIDDM has an insidious onset and is diagnosed usually by an elevated glucose level on routine examination. * **_One in six patients with NIDDM will develop end-stage renal disease or will require a lower extremity amputation for severe vascular disease_** * **_One in five will become legally blind from retinopathy._** * In contrast to patients with IDDM, patients with NIDDM usually do not develop ketoacidosis. * In general, the development of NIDDM is divided into three clinical phases. * First, the plasma glucose concentration remains normal despite elevated blood levels of insulin, indicating that the target tissues for insulin action appear to be relatively resistant to the effects of the hormone. * Second, postprandial hyperglycemia develops despite elevated insulin concentrations. * Third, declining insulin secretion causes fasting hyperglycemia and overt diabetes. * In addition to hyperglycemia, the metabolic dysregulation resulting from islet β-cell dysfunction and insulin resistance causes atherosclerosis, peripheral neuropathy, renal disease, cataracts, and retinopathy. * Chronic hyperglycemia can be monitored by means of measurements of the percentage of hemoglobin that has become modified by glycosylation, referred to as HbA1c. * Rigorous control of blood glucose levels, as determined by HbA1c levels as close to normal as possible (\<7%), reduces the risk of complications by 35% to 75% and can extend the average life expectancy.

Question 30

Intracellular Movement of Insulin

Answer 30

Intracellular Movement of Insulin 1. Gene coding for insulin are transcribed to mRNA in the nucleus. 2. After moving into the cytoplasm, translation of the mRNA is initiated on cytosolic ribosomes, with formation of an N-terminal hydrophobic signal sequence that aids in the transport of the mRNA and ribosomes to the RER. 3. The N-terminal signal sequence penetrates the membrane of the RER. Further elongation directs the polypeptide chain into the lumen of the RER, resulting in the formation of preproinsulin 4. the signal sequence is cleaved and proinsulin is formed in the lumen (cisternal space). 5. Proinsulin is transported from RER to the Golgi complex, where it is cleaved forming insulin and C-peptide. 6. insulin and C-peptide in secretory granules. 7. secretory granules are secreted by exocytosis, releasing insulin and C-piptide. (Pic on slide 39)

Question 31

Insulin Signaling

Answer 31

Insulin Signaling * Gluconeogenesis results in the generation of glucose from non-carbohydrate carbon substrates such as lactate, glycerol and glucogenic amino acids. * Glycogenolysis is the breakdown of glycogen polymers into glucose monomers. * Lipolysis is the breakdown of lipids and involves the hydrolysis of triglycerides into free fatty acids followed by further degradation into acetyl units by beta-oxidation. slide 40 has the mechanism

Question 32

what is the mechanism for Insulin signaling recruiting GLUT transporters to the cell membrane? (empty, have to type out the pic on slide 41)

Answer 32

Insulin signaling recruits GLUT transporters to the cell membrane

Question 33

Insulin Receptor Signaling

Answer 33

Insulin Receptor Signaling

Question 34

Insulin receptor signaling Example of Receptor Tyrosine Kinase Growth Factor Signaling

Answer 34

Insulin receptor signalingExample of Receptor Tyrosine Kinase Growth Factor Signaling * The insulin receptor is a dimer of two membrane-spanning α–β pairs. * The tyrosine kinase domains are shown in red, and arrows indicate auto-phosphorylation. * The activated receptor binds IRS molecules (insulin receptor substrates) and phosphorylates IRS at multiple sites, thereby forming binding sites for proteins with SH2 domains: Grb2, phospholipase Cγ(PLCγ), and PI 3-kinase. Both PLCγ and PI 3-kinase are associated with various phosphatidylinositol phosphates (all designated with PIP) in the plasma membrane. Grb2 associates with a protein known as GAP1 (Grb2-associated protein), which has a pleckstrin homology domain that associates with phosphatidylinositol phosphates in the membrane.

Question 35

The insulin receptor–AKT signaling pathway

Answer 35

The insulin receptor–AKT signaling pathway * Activation of AKT (PKB) has four important consequences * First, it leads to movement of GLUT4 from vesicles in the cytosol to the plasma membrane, allowing glucose uptake. * Second, AKT can phosphorylate glycogen synthase kinase-3 (GSK-3), reducing its activity. This leads to an increase in glycogen synthesis * Third, it alters TSC/mTOR signaling pathway and increase protein translation * Fourth, it alters FOXO pathway leading to deceased glucose synthesis

Question 36

The “Fountain of Youth” pathway

Answer 36

The “Fountain of Youth” pathway * mTOR stands for mammalian target of rapamycin. * Rapamycin is an immunosuppressive drug which was recently discovered to also prolong life in mice (anti-aging effect). * The mTOR containing complexes mTORC1 and mTORC2 are central components of nutrient- and hormone-sensitive pathways that regulate cell growth and proliferation. * Abnormal regulation of these pathways have been linked to numerous human diseases and disorders including diabetes, Cowden syndrome, Peutz Jeghers syndrome, Von Hippel Lindau syndrome, sporadic cancers and tuberous sclerosis. * Drugs, such as mTOR inhibitor temsirolimus (Torisel), target this pathway to help treat cancer.

Question 37

TSC/mTOR Signaling Pathways (need to be written up cause pic might be too small) **Favorite it!**

Answer 37

TSC/mTOR Signaling Pathways --------------------------------------------------- notes **eLF4E binding protein** is an inhibitor that inhibits **eIF4E** The cell knows if there is **low** energy in the cell to initiate the **AMPK** pathway (on the left) by measuring **nucleotide levels** (how much AMP, ATP, if there is a high level of AMP, not much energy is available.) DO NOT CONFUSZE mTORC1 with mTORC2! * *mTORC2** _phosperylates Akt_ * *mTORC1** is down stream to mTORC2. **mTORC1** goes after 4eBP binding proteins (like eLF4E binding protein that inhibits eLF4E) shutting it off. It also goes after P70s6k kinases (like S6) activating it. Once Akt is active, it will shut off the Gap TSC1 & 2. this means Rheb and mTORC1 is on, which means the inhibitor is turned off (eLF4E binding protein), which means eLF4E is on and can bind to the methyl cap and initial translation. Gap TSC1 & 2 controls Rheb by the damn way

Question 38

Rapamycin (Sirolimus)

Answer 38

Rapamycin (Sirolimus) * A potent immunosupressant used to prevent transplant rejection (by interfering with the cell's ability to respond to iL-2) * Mode of action: * Binds to FK-binding protein 12 (FKBP12) * The rapamycin-FKBP12 complex binds and inhibits mTOR Complex 1 (mTORC1) stopping it's activity blocking cell growth ---------------------------------------------- Reminder! remember cyclosporin A job was to block the production of iL-2? good... don't get it mixed up with rapamycin.

Question 39

Central role of mTOR and TSC1/TSC2 complex in regulating protein synthesis

Answer 39

Central role of mTOR and TSC1/TSC2 complex in regulating protein synthesis ## Footnote (slide 50 for larger pic)

Question 40

what are the 3 major branches of receptor tyrosin kinase? **Favorite it!**

Answer 40

The 3 major branches of receptor tyrosin kinase (an example of enzyme coupled receptor) are 1. Ras MAPK kinase 2. Phospholipase c gama route 3. Pi3-kinase/ Akt route. --------------------------------------- additional info another example of enzyme couple coupled receptors is serine threonin kinase

Question 41

Role of TSC1 & 2 in the mTOR signaling

Answer 41

Role of TSC1 & 2 in the mTOR signaling * Tuberous sclerosis complex (TSC) tumor suppressors (TSC1 and TSC2) is regulated by multi-site phosphorylation and acts as a point of integration for cellular signaling * When active, the TSC1-TSC2 complex acts as a GTPase activating protein (GAP) for the Ras-related small G protein Rheb. * Thus turning Rheb off by stimulating its intrinsic GTPase activity. * In the presence of growth factors and nutrients, this TSC1-TSC2 complex is turned off, allowing the GTP-bound active version of Rheb to accumulate and turn on downstream pathways such as mTORC1 * The mTORC1 complex is a critical regulator of cell growth and proliferation.

Question 42

Tuberous sclerosis

Answer 42

Tuberous sclerosis * Tuberous sclerosis is an inherited disorder whose key features include multiple facial angiofibromas, hypopigmented macules, periungual fibromas, seizures, Shagreen patch, cardiac rhabdomyoma, and renal lesions. * Many patients also have ocular and neurologic manifestations, including mental retardation. * The disorder results from a mutation in either TSC1 or TSC2 * ~Two-thirds of cases may be due to spontaneous genetic mutations. * TSC2 mutations account for the majority of sporadic cases * these individuals usually show a more severe phenotype that includes renal lesions and neurologic deficits ---------------------------------------------- In case you haven't noticed.... TSC1 and 2 got it's name from it's associating desease Tuberous sclerosis with complex 1 and 2 added to it!

Question 43

AMP-activated protein kinase (AMPK)

Answer 43

AMP-activated protein kinase (AMPK) * A pivotal regulatory molecule in the metabolism of carbohydrates and fats * Example of hepatic targets of AMPK: * mTORC1 – decreased protein synthesis * TSC2 - decreased protein synthesis * Transducer of regulated CREB activity 2 (TORC2) –protein is sequestered in the cytoplasm when phosphorylated by AMPK, leading to decreased expression – at the transcription level of gluconeogenic enzymes * (Gluconeogenic = generation of glucose from non-carbohydrate carbon substrates) -------------------------------------- DO NOT CONFUSE TORC2 ( stands for transducer of regulated CREB activity) is a TRANSCRIPTIONAL REGULATOR which is a target of AMPK mTORC2 PHOSPHORYLATES AKT

Question 44

AMPK Activation

Answer 44

AMPK Activation * AMPK can be activated in several ways, all of which depend on increased AMP levels within the cell * Activated by phosphorylation by LKB1 or calmodulin kinase kinase * The drug Metformin activates AMPK via LKB1

Question 45

Metformin **Favorite it**

Answer 45

Metformin Has MULTIPLE modes of action. * Metformin – a drug used for treatment of type 2 diabetes (The drug Metformin activates AMPK by influencing LKB1) * Metformin reduces blood glucose levels by inhibiting hepatic gluconeogenesis, which is active in these patients because of the liver’s resistance to the effects of insulin. * Metformin also reduces lipid synthesis in the liver * Metformin stimulates glucose uptake **_by the muscle_**, which is significant for reducing circulating blood glucose levels ------------------------------------------------ info regarding the picture (also on slide 57) one propose meechanism for how Metformin works is by preventing or decreasing gene expression of the enzymes involved in gluconeogenesis

Question 46

Activation of CREB **Favorite it!**

Answer 46

Activation of CREB * Under normal conditions, increases of hepatic cAMP levels (e.g. in response to glucagon) activates CREB, which, in combination with TORC2, leads to increased transcription of genes required for gluconeogenesis. * Under conditions of insulin resistance, this pathway remains stimulated, even in the presence of insulin. ------------------------------------------- proposed mechanism for metformin drug Metformin works by preventing or decreasing gene expression of the enzymes involved in gluconeogenesis

Question 47

Action of metformin on gluconeogenesis **Favorite it!!!!**

Answer 47

Action of metformin on gluconeogenesis * Metformin stimulates the activation of AMPK, * AMPK phosphorylates TORC2 and sequesters it in the cytoplasm, thereby decreasing synthesis of gluconeogenic enzymes and reducing hepatic output of glucose -------------------------------------------------- top half of pic is the action of metformin drug for diabetes type ll (slide 59) DO NOT CONFUSE TORC2 with mTORC2

Question 48

serine/threonine kinase pathway **favorite it!**

Answer 48

Some growth factors transduce signals via receptor serine/threonine kinase (an enzyme coupled receptor) * Transforming Growth Factor Beta receptor is an example of serine/threonine kinase TGF beta signaling pathway (normally see this pathway compromised in colon cancers) 1. TGF ligand binds a complex of type I and II receptors 2. Type II receptor phosphorylates type I receptors 3. Type I receptors phosphorylates an R-Smad (Smad2 or Smad3) 4. R-Smad complexes with Smad4 (a co Smad). Smad 4 forms a heteroduplex with the regulatory Smad, disassociate with the receptor complex and migrates into the nucleous. 5. In the nucleus, the Smad complex associates with transcription factors to repressing gene expression/ cell proliferation ------------------------------------------------ TGF beta usually tell cells not to proliferate. so if this pathway is shutdown... you get proliferation that can lead to cancer. in colon cancer, often it's either the receptor gene or Smad 4 is not working.

Question 49

Signaling Pathway of TGF-beta | (Picture form)

Answer 49

Signaling Pathway of TGF-beta

Question 50

Other Types of Enzyme-Linked Receptors

Answer 50

Other Types of Enzyme-Linked Receptors Protein-tyrosine phosphatases remove phosphate groups from phosphotyrosine, counterbalancing the effects of protein-tyrosine kinases. * Example: CD45 * CD45 is a receptor which is expressed on T and B lymphocytes. * Following antigen stimulation, CD45 dephosphorylates a phosphotyrosine that inhibits the enzymatic activity of Src family Receptor Guanylyl Cyclases * Ligand binding to receptors stimulates the cyclase activity of the receptor to synthesize cGMP

Question 51

JAK-STAT Signaling Pathway favorite it

Answer 51

JAK-STAT Signaling Pathway * Cytokine receptors activate the JAK-STAT signaling pathway, providing a fast track to the nucleus * Tyrosine kinase-associated receptors called JAK-STAT are often used by cytokines to regulate the proliferation of certain cells involved in the immune response. * The receptor itself has no intrinsic kinase activity, but it binds the tyrosine kinase JAK (Janus kinase) * STAT proteins are located in the cytosol and are referred to as latent gene regulatory proteins because they only migrate into the nucleus and regulate transcription after they are activated * There are many different STAT proteins. * Receptors for different cytokines bind different STATs

Question 52

The receptor protein Notch **Favorite it**

Answer 52

The receptor protein Notch is a latent gene regulatory protein * Notch signaling is very important during development * For example, when individual epithelial cells begin to develop as neural cells, they signal to their neighbors not to do the same through Delta binding receptor Notch * Upon binding Delta, Notch is cleaved on either side of the plasma membrane **by gama secretases** * The freed cytoplasmic tail of Notch then migrates to the nucleus, associates with other regulatory factors bound to promoter elements and modulates gene transcription

Question 53

Fibroblast growth factors (FGF1 to FGF24) **Favorite it!**

Answer 53

Fibroblast growth factors (FGF1 to FGF24) is one of the receptor tryosine kinases Signal protein: Fibroblast growth factors (FGF1 to FGF24) Receptor: FGF recepts (FGFR1-FGFR4, plus multiple isoforms of each) Responses: stimulate proliferation of various cell types, inhibit differentiation of some precursor cells; act as inductive signals in development.

Question 54

Insulin **Favorite it!**

Answer 54

Insulin is one of the receptor tyrosine kinases ## Footnote Signal protein: insulin Receptor: insulin receptor Responses: stimulates carbohydrate utilization and protein synthesis.

Question 55

platelet-derived growth factors (PDGF,AA,BB,AB) **Favorite it!**

Answer 55

Platelet-derived growth factors is one of the receptor tyrosine kinases ## Footnote signal protein: platelet-derived growth factors (PDGF,AA,BB,AB) receptor: PDGF receptors (alpha and beta) responses: stimulate survival, growth, proliferation, and migration of various cell types

Question 56

Vascular endothelial growth factor **Favorite it!**

Answer 56

Vascular endothelial growth factor is one of the receptor kinases ## Footnote signal protein: Vascular endothelial growth factor receptor: VEGF receptors Reponses: stimulates angiogenesis

Question 57

The Wnt/b-catenin signaling pathway **Favorite it!**

Answer 57

The Wnt/b-catenin signaling pathway (commonly compromised in colon cancer) * Wnt proteins are secreted signal molecules that act as local mediators and morphogens to control many aspects of developments. * Wnt protein bind to Frizzled receptors and inhibit the degradation of b-catenin * APC is a key protein in the regulation of b-catenin * APC is often mutated in some forms of colon cancer * b-catenin accumulates and translocates to the nucleus * Once in the nucleus, b-catenin migrates to the nucleus, displaces Groucho and associates with coactivator ----------------------------------- additional info common genes mutated in this pathway is APC and not as common, beta cateninin

Question 58

The Wnt/b-catenin signaling pathway (when Wnt is absent) **Favorite it!**

Answer 58

The Wnt/b-catenin signaling pathway When Wnt is absent * The “Destruction Complex” composed of **_APC_**, axin, GSK3 and CK1 phosphorylates b-catenin * b-catenin is ubiquitylated and degraded * b-catenin levels kept low in the cell what they do is phosphorylate targets. when this happens, E3 ubiquitin ligases will now recognize the target as something that needs to be marked with ubiquitin. once marked, proteasomes destroys the target protein. the one most know target for the wnt/b-catenin signaling pathway is the 5 function protein beta cateninin. ------------------------------------------- common genes mutated in this pathway is APC and not as common, beta cateninin

Question 59

The Wnt/b-catenin signaling pathway (when Wnt is present) **Favorite it!**

Answer 59

The Wnt/b-catenin signaling pathway * Wnt binds and activates receptors Frizzled and LRP * Dishevelled becomes activated * The “Destruction Complex” is turned off * b-catenin is no longer marked for destruction and its levels increase within the nucleus * b-catenin modulates gene expression it will associate with other transcriptional regulator and need to changes in gene expression. ------------------------------------------------ common genes mutated in this pathway is APC and not as common, beta cateninin | (when Wnt is present)

Question 60

Hedgehog signaling

Answer 60

Hedgehog signaling * The receptor for Hedgehog is a transmembrane protein (Patched) that inhibits a second transmembrane protein (Smoothened) by an unknown mechanism. * Binding of Hedgehog inhibits Patched, leading to activation of Smoothened, which initiates a signaling pathway leading to activation of a transcription factor Ci (Drosophila) or Gli (mammals). BASICALLY: In the absence of hedgehog, transcriptional regulator get destroyed. When hedgehog is present, the activation subunit will be allowed to stay around and go into the nucleus and modify transcription. At the cell’s surface, you’re going to have hedgehog receptors called patch. Whenn patch binds to hedgehog, destruction wont occur.

Question 61

NF-kB-Dependent Signaling Pathway information

Answer 61

NF-kB-Dependent Signaling Pathway * The NF-kB proteins are latent gene regulatory proteins that are present in most animal cells and are central to many stressful, inflammatory, and innate immune responses. * These responses occur as a reaction to infection or injury and help protect stressed multicellular organisms and their cells. * Upon cell signaling, the inhibitory protein (IkB) dissociates from NF-kB in the cytosol. * NF-kB migrates to the nucleus , where it interacts with coactivators to alter gene transcription --------------------------------------- additional info The activation of NF-kB pathway by TNF. Both TNF and its receptors are trimers. The binding of TNF causes a rearrangement of the clustered cytosolic tails of the receptors, which now recruit various signaling proteins, resulting in the phosphorylation and activation of IKK complex. IKK then phosphorylates IkB, which marks the protein for ubiquitylation and degradation in proteosomes. The releasedNF-kB translocates into the nucleus, where in collaboration with coactivator proteins, it stimulates the transcription of its target genes.

Question 62

NF-kB-Dependent Signaling Pathway

Answer 62

NF-kB-Dependent Signaling Pathway 1. TNF binds the receptor 2. IkB Kinase becomes active and phosphorylates IkB 3. IkB is ubiquitinated and degraded 4. NF-kB is no longer bound to IkB and it’s nuclear localization signal is now exposed 5. NF-kB is transported into the nucleus through the nuclear pores 6. NF-kB binds to promoter elements and modulates gene expression

Question 63

Signal Transduction and the Cytoskeleton

Answer 63

Signal Transduction and the Cytoskeleton * The functions of most cells are directly affected by cell adhesion and the organization of the cytoskeleton. * Receptors responsible for cell adhesion initiate intracellular signaling pathways that regulate other aspects of cell behavior, including gene expression. * Growth factors often induce cytoskeletal alterations resulting in cell movement or changes in shape. * Components of the cytoskeleton thus act as both receptors and targets in cell signaling pathways. * Integrins are the main receptors for the attachment of cells to the extracellular matrix and also interact with the cytoskeleton. * Integrins also serve as receptors that activate intracellular signaling pathways. *

Question 64

Integrin signaling

Answer 64

* Binding of integrins to the extracellular matrix leads to activation of FAK ( focal adhesion kinase), a nonreceptor protein-tyrosine kinase. * Phosphorylation of FAK provides binding sites for several signaling molecules, including the Grb2-Sos complex, leading to activation of Ras, PI 3-kinase, and phospholipase C-γ. (pic on slide 74)

Question 65

Regulation of actin remodeling by Rho family proteins

Answer 65

Regulation of actin remodeling by Rho family proteins * Signaling from integrins and growth factor receptors also regulates the dynamic behavior of the actin cytoskeleton. * Members of the Rho subfamily of small GTP-binding proteins (including Rho, Rac, and Cdc42) play central roles in regulating the organization of the actin cytoskeleton Rho, Rac, and Cdc42 are monomeric G proteins. They are involved in controlling cytoskeleton formation. Cell signaling can effect their behaviors.

Question 66

Stimulation of actin polymerization by Rho family proteins

Answer 66

Stimulation of actin polymerization by Rho family proteins * Rho family members are activated by integrin signaling and growth factor receptors. * Multiple target proteins mediate the cytoskeletal changes. All of the Rho family proteins promote actin polymerization.

Question 67

Regulation of myosin light-chain phosphorylation by Rho

Answer 67

Regulation of myosin light-chain phosphorylation by Rho * Rho also targets a protein-serine/threonine kinase called ROCK, which promotes the formation of stress fibers.

Question 68

Signaling Networks

Answer 68

Signaling Networks * Signaling pathways do not operate in isolation, and intracellular signal transduction is really an integrated network of connected pathways. * Computational modeling of signaling networks is currently a major challenge in cell biology. * The activity of signaling pathways is controlled by feedback loops. * An example of a negative feedback loop is the NF-κB pathway. * NF-κB is activated as a result of phosphorylation and degradation of IκB, but one of the genes activated by NF-κB encodes IκB, generating a feedback loop that inhibits NF-κB activity.

Question 69

Feedback inhibition of NF-κB

Answer 69

Feedback inhibition of NF-κB * This regulation is critical because extent and duration of NF-κB activity can determine the transcriptional response of the cell. * Some target genes are induced by transient NF-κB activity, (30–60 minutes), but induction of other genes requires several hours of sustained NF-κB signaling.

Question 70

Signaling Networks-Crosstalk

Answer 70

Signaling Networks-Crosstalk ## Footnote Crosstalk is the interaction between signaling pathways, such as junctions between Ca2+ and cAMP signaling, between the cAMP and ERK pathways, and between integrin signaling and receptor protein-tyrosine kinases. The downstream signaling pathways activated by receptor tyrosine kinases or by G-protein linked receptors overlap and are interconnected

Question 71

Elements of signaling networks

Answer 71

Elements of signaling networks * Multiple signal transduction pathways interact with one another to form signaling networks within the cell. * Junctions between pathways can be either positive (one pathway stimulates the other) or negative (one pathway inhibits the other). * Signaling networks can contain positive feedback loops and feedforward relays in which the activity of one component stimulates a distant downstream component.

Question 72

Rho, Rac, and Cdc42 are?

Answer 72

Rho, Rac, and Cdc42 are monomeric G proteins. They are involved in controlling cytoskeleton formation. Cell signaling can effect their behaviors.

Question 73

APC

Answer 73

APC is a key protein in the regulation of b-catenin in the Wnt/b-catenin signaling pathway APC is often mutated in some forms of colon cancer

Question 74

4 major consequences of the Pi3-kinase/ Akt route

Answer 74

4 major consequences of the Pi3-kinase/ Akt route 1. get the GLUT permeases to the cell's surface. so that you can bring in more glucose 2. increase glycogen synthesis. if you bring in glucose, you gotta do something with it right? how do you store glycose? exactly. 3. if you're bringing in glucose, you dont have to make glucose right? FOXO is phosphorylated so it wont get to the nucleus and wont ulter gene expression for genes that deal with glucose synthesis. 4. TSC1&2 mTOR signaling inhibition leading to global rates of translation. tuberous sclerosis

Question 75

Receptor gets triggered by peptide antigen. What branch of the receptor tyrosine kinase are we talking about at that point?

Answer 75

receptor gets triggered by peptide antigen. What branch of the receptor tyrosine kinase are we talking about at that point? **Phospholipase C gamma ** The pathway: when activated, phospholipase C gama cleaves pip 2 into DAG and IP-3. IP-3 stimulates release of calcium from the smooth ER. calcium binds to calmadulin calmadulin bind to calcinerin calcinerin is a phosphatase. it dephosphorylates NFAT (an transcription factor) NFAT (like PH04) bind to importin and is transported into the nucleus. Now NFAT can modulate gene expression. NFAT can stimulate IL-2 while inside the nucleus. A MUST know drug that will suppress this pathway is cyclosphorin A. It will block the calcinerin step so that calcinerin cannot dephosphorylate NFAT...so it's nuclear signaling sequence remains blocked. If it remains blocked, it cannot bind to importin in order to get into the nucleus where it would stimulate IL-2 and thereby produce an immune response.