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Cell Stuff Flashcards

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1
Q

What is the response regulator in Caulobacteria cell division?
What does it do?

A

CtrA
Phosphorylated on aspartate 51
Once phosphorylated, binds DNA
Activates transcription of about 100 genes - many involved in polar morphogenesis and cell division

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2
Q

What happens if there is a mutation in CtrA? Example

A

Developmental defects

Eg ctrA401 - cels no longer divide or differentiate, they just grow long

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3
Q

What is a transcription factor?

A

A protein that binds to specific DNA sequences, controlling the flow (or transcription) of genetic material from DNA to mRNA

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4
Q

How do DivJ and PleC affect CtrA?

A

They regulate CtrA phosphorylation

  • PleC (in swarmer cell/pole) keeps DivK de phosphorylated, leaving DivL/CckA free. DivL promotes CckA kinase activity and consequently phosphorylation of CtrA. CtrA is ACTIVATED
  • DivJ (in stalk cell/pole) keeps DivK phosphorylated. Phosphorylated DivK inhibits DivL/CckA. This inhibits CckA kinase activity and causes CckA to show phosphatase activity, which drives the dephosphorylation of CtrA. CtrA is INACTIVATED
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5
Q

What second species undergoes asymmetrical cell division during sporulation?

A

Bacillus subtilis

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6
Q

Where does sporulation occur in Bacillus subtilis cells?

A

Only at one end of the cell

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7
Q

Spore formation in B. subtilis

A
  1. Rod shaped cell
  2. Asymmetric cell division, produces spore cell 1/4 of the size of mother cell
  3. Mother cell engulfs daughter
  4. Mother cell dies, releasing spore
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8
Q

What transcription factor is key to sporulation in B. subtilis?

A

oF (sigma F)
oF is inactive in the mother cell, but activated in the prespore
It’s regulation depends on three other proteins: Spo11AB, Spo11AA and Spo11E

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9
Q

What is the process by which oF activates spore formation?

A
  • SpollAB protein binds and inactivated the transcription factor sigma F (oF)
  • Binding of oF by SpoIIAB is regulated by a phosphatase called SpoIIE (SpoIIE activity dissociated SpoIIAB from oF)
  • SpoIIE is only expressed on the newly formed septum between the mother cell and prespore
  • Because the mother cell is four times larger than the prespore the effective concentration of SpoIIE is higher in the prespore. Increased SpoIIE activity dissociates SpoIIAB from oF
  • Feedback loops ensure that this situation arises quickly and is stable
  • oF activates spore formation
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10
Q

What determines asymmetrical cell division in C. elegans?

A

Asymmetry is determined by microtubules associated with the sperm pronucleus, which recruit specific proteins

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11
Q

What determines asymmetry in A. thaliana?

A

Asymmetry involves the expression of mRNA for different WOX homeobox transcription factors in different parts of the zygote

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12
Q

Process of asymmetric cell division during embryogenesis in C. elegans

A
  • The zygote divides along its anterior-posterior axis and gives rise to the AB and P1 cells, which produce different cell lineages
  • The two daughter cells contain different PAR proteins (named after the mutant phenotypes, which are PARtitioning defective)
  • Microtubules associated with the sperm pronucleus recruit PAR-2 protein
  • PAR-1 is recruited and downregulates cytoplasmic MEX-5
  • This permits the expression of PIE-1
  • Cleavage across the centre produces one cell expressing PIE-1 and one not expressing PIE-1
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13
Q

In what animals do PAR proteins affect cell polarisation?

A

PAR proteins affect cell polarisation, in all bilateral animals including C. elegans, Xenopus oocytes, Drosophila and mammals

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14
Q

What do PAR proteins switch between?

A

A rapidly mixing cytoplasmic state and a more slowly diffusing membrane-associated state

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15
Q

How to PARs regulate polarisation of cytoplasmic determinants?

A

Cytoplasmic determinants = MEX-5, PIE-1

Polarise by switching between fast- and slow-diffusing cytoplasmic states. This switching is regulated by PARs

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16
Q

What do WOX genes determine in A. thaliana?

A

Apical-basal axis formation or the establishment of shoot and root meristem precursors

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17
Q

What is caused by the wox2 mutation?

A

Single cotyledon

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18
Q

What is caused by the wox8wox9 mutation?

A

Double mutants - severely defective embryo and suspensor

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19
Q

Wha is the distribution of WOX2 and WOX8 before the first cell division in A. thaliana?

A

WOX2 at tip of cell

WOX8 in lower part of cell

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20
Q

What 4 different domains do WOX expression mark?

A
  1. The upper tier of the embryo proper that will develop into cotyledons and shoot meristem (WOX2)
  2. The lower tier of the embryo proper which will generate the hypercotyl, the primary root and the proximal part of the root meristem (WOX9)
  3. The hypophysis from which the QC and columella stem cells are derived (WOX8+9)
  4. The suspensor (WOX8)
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21
Q

What is the name for a cell where all genes are available for use?

A

Totipotent

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22
Q

What is the name for a cell where a range of genes and cell fates are still available but not all?

A

Pluripotent

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23
Q

Example of cell gate determination: single called eukaryote

A

Anabaena
Forms heterocysts
An example of cell fate induction after cell division
Before induction all cells are identical vegetative cells
Cell gate determined by dynamic interactions between diffusible positive (2-OG) and negative (PatS peptide) signals
Feedback loops involving the HetR protease and NtcA transcription factor amplify the signals
The result is optimal spacing of heterocysts

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24
Q

Why does a heterocyst form?

A

Cyanobacteria grow long filaments of photosynthetic vegetative cells
About every 10-15th cell, a vegetative cell differentiated into an anaerobic, nitrogen-fixing heterocyst
Heterocyst supply fixed nitrogen to neighbouring vegetative cells in return for the products of photosynthesis
This separation of cellular functions is necessary because cyanobacteria produce oxygen during photosynthesis, but nitrogenase is unstable in the presence of oxygen
The differentiation of heterocysts is provoked by the absence of a fixed nitrogen source

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25
Features of heterocysts
- Have a thick envelope to limit oxygen penetration - Photosystem II is absent so no oxygen is evolved - High respiratory rate consumes oxygen - Hetercysts are terminally differentiated; their fate is determined
26
How is heterocyst formation regulated?
- Low nitrogen leads to 2-oxoglutarate (2-OG) accumulation - 2-OG is sensed by NtcA, a transcription factor whose DNA-binding activity is stimulated by 2-OG - NtcA activity leads to the expression of HetR - The activator of heterocyst formation, HetR, is expressed in all cells in response to a lack of fixed nitrogen - HetR is a protease with DNA-binding activity - Positive feedback involving NtcA and HetR amplifies the signal - Several cells start to respond to the 2-OG signal simultaneously - They produce PatS, a short peptide - PatS diffuses from the heterocysts and inhibits the differentiation of neighbouring cells, which revert to the vegetative state - PatA counteracts the action of differentiation inhibitors in the differentiating cell - HetN is expressed in mature hetercysts and suppresses the formation of new heterocysts adjacent to previously formed heterocysts - A portion of PatS and HetN containing the RGSGR pentapeptide diffuses away from source cells to neighbouring cells establishing an inhibitor concentration gradient along the filament
27
Example of cell fate determination in a multicellular prokaryote
Saccharomyces cerevisiae mating type switch Cell fate is determined by DNA rearrangements that replace one version of the MAT locus with another Different combinations of transcription factors encoded by different versions of the MAT locus control cell fate Fate of the mother cell is switched part way through a series of cell divisions
28
What are the two haploid mating types of S. cerevisiae?
a haploid cell or alpha haploid cell Diploid cells are a/alpha but undergo meiosis when put under stress producing two a cells and two alpha cells A single haploid spore germinates has divides. The new mother cell then switches it’s mating type before dividing again to produce two cells of the opposite mating type from the original spore. These two new cells can then mate cells of the opposite mating type to generate diploids
29
What do a cells produce?
“a factor” - a short peptide (12 aas) that signals to alpha cells, which grow as shmoos towards a source of the a factor
30
What do alpha cells produce?
“alpha factor” - a short peptide mating pheromone (13 aas) that signals to a cells, which grow as shmoos towards the source of alpha factor Ste3: a trans-membrane receptor of the opposite mating pheromone
31
Where is the MAT locus found?
On S. cerevisiae chromosome III
32
What does S. cerevisiae chromosome III also contain?
Two other copies of the mating-type cassette at the HML-alpha and HMRa loci
33
How does mating type switching occur in S. cerevisiae?
An endonuclease (HO) cleaves the MAT locus and triggers recombination between the active MAT and silent HML or HMR cassettes
34
What silences the HML and HMR loci?
A histone deacetylase (HDAC) called Sir2 Sir2 deacetylates histones H3 and H4 in the silencer regions, producing short regions of heterochromatin where transcription does not occur
35
What do a and alpha genes encode?
- Transcription factors - a1 and alpha2 are homeodomain transcription factors - a cells express a1. This has no effect in haploid cells, which express a set of a specific genes - alpha cells express alpha2-alpha2 homodimers, which repress a-specific genes. They also express alpha1, which activated alpha-specific genes - Mating produces diploid cells that express alpha2-alpha2 homodimers. These repress a-specific genes. They also express a1-alpha2 heterodimers, which repress a set of haploid-specific genes
36
What is Ste3?
The a factor receptor which is only expressed on alpha cells
37
What is Ste2?
The alpha factor that is only expressed on a cells
38
What are Ste2 and Ste3 coupled to?
Coupled to and activate the same hetertrimeric G protein, consisting of a G-alpha subunit (Gpa1) and a Gbetagamma heterodomer (Ste4/Ste18) Once the G protein is activated this leads to a MAP kinase cascade and the phosphorylation of the STE12 transcription factor
39
What does phosphorylated STE12 cause?
Synthesis of cell surface proteins that mediate cell-cell adhesion Proteins that mediate cell fusion synthesised Cel cycle arrest Cells turn into shmoos with mating projections
40
Two types of cell death in Bacillus subtilis
1. Cannibalism 2. Spore mother cell lysis - covered in asymmetrical cell division Both responses to severe stress (starvation)
41
Cannibalism in B. subtilis
Cell death triggered by skf and sdp toxins secreted by cells starting to sporulate This causes the lysis of some cells, releasing their nutrients
42
Stages of B. subtilis mother cell death
- Sigma F in prespore permits sigma E activity in mother cell - Sigma E induces transcription of genes including nucB, an extracellular nuclease - Spore detaches from mother cell pole - Mother cell membranes rupture - Mother cell walls collapse due to peptidoglycan hydrolase activity - NucB degrades mother cell DNA
43
Two types of cell death in animals
1. Necrosis | 2. Programmed cell death (PCD)
44
Two types of programmed cell death
Type I: Apoptosis | Type II: Autophagic cell death
45
Definition of apoptosis
Disintegration of cells into membrane-bound particles that are then engulfed and digested by another cell
46
Definition of autophagic cell death
Self-digestion by a cell. Contents are engulfed by double-membraned vesicles and degraded by the cell’s own vacuole lysosomes
47
Animal cell necrosis
“Accidental” cell death Caused by cell damage due to trauma: -Physical: mechanical damage, extreme heat/cold -Chemical: poison -Biological: toxin/venom Ordered digestion by the injured cell or other cells prevented/reduced Subsequent release of naked intracellular contents can cause inflammation
48
Stages of apoptosis
1. Cytoplasm begins to shrink. Nuclear condensation observed (breakdown of chromatin and nuclear structural proteins) 2. Cells continue to shrink 3. Cels package themselves in a form that allows easy clearing by macrophages - often undergo plasma membrane changes that trigger the macrophage response 4. Small vesicles called apoptotic bodies sometimes observed
49
Why evolve PCD?
1. To remove unwanted or dangerous cells eg. infected cells, cancer cells, autoimmune cells, surplus cells 2. To facilitate development: to sculpt or organise new tissues and organs and to remove redundant cells, tissues and organs
50
Examples of organisms using PCD
- The resorption of the tadpole tail during metamorphosis into a frog - The formation of fingers and toes of a foetus requires apoptosis of the tissue between them - The formation of synapses between neurones in the brain requires surplus cells to be eliminated
51
How do cells with damaged DNA respond?
They could cause a cell to disrupt proper embryonic development or become cancerous They increase their production of p53, a potent inducer of apoptosis
52
What animal does not need apoptosis to live a normal life span if their two apoptosis genes are mutated?
C. elegans ced-3 and ced-4 131 cells which would normally undergo apoptosis during development do not, but it does not affect the lifespan of the worm
53
What animals cannot survive development without apoptosis?
Drosophila | Mice
54
How was it proved that apoptosis is essential for digits to form?
KO knockout mice have impaired apoptotic signalling - developmental defect in paws
55
What is the cell death rate in vertebrate and insect central nervous / immune systems?
as high as 80% | nerves must form good connections and immune cells must find appropriate targets or they die
56
What are CTLs?
Cytotoxic T lymphocytes | Can be used to kill virus-infected cells by apoptosis in plants
57
When is apoptosis required during plant development?
- Xylogenesis - death of tracheal elements to produce xylem vessels - Self-incompatibility - Aerenchyma - soft plant tissue containing air spaces
58
Examples of PCD in plants to detach organs
Abscission and senescence - Leaves - Petals (especially after fertilisation) - Fruit
59
How is cell death programmed in C. elegans?
Four components required CED-9 found at the surface of the mitochondria where it keeps CED-4 to prevent it from activating CED-3. When EGL-1 binds to CED-9 it releases CED-4 which activates CED-3 at the nucleus, causing cell death The apoptosome contains two molecules of CED-3 surrounded by 4 dimers of CED-4
60
what is the apoptosome in Drosophila?
Cytosolic signalling platform that activates caspase
61
What amplifies the cell death pathway in mammals?
The release of apoptotic molecules from the mitochondrial intermembrane space, including cytochrome C
62
Diagnostic features of apoptosis
- Maintenance of intact cell membranes during the death process so that the cell does not release its contents and trigger an inflammatory reaction - Fragmentation of the cell into membrane-bound apoptotic bodies - Nuclear and cytoplasmic condensation - Endolytic cleavage of DNA into small oligonucleosomal fragments
63
Evolution of cell death
- Some phage lyse bacteria cells using holins - Some bacterial cells use holins to lyse - Some Bcl-2 related genes involved in losing mitochondria in mammals and lyse bacterial cells too - evolutionary link?
64
What are non-coding RNAs?
- Do not encode protein or peptide sequences | - Encoded by RNA-coding genes
65
Examples of non-coding RNAs
``` Ribosomal RNA (rRNA) Transfer RNA (tRNA) Small nuclear RNA (snRNA) Ribonuclease P microRNA (miRNA) CRISPR RNA ```
66
What are non-coding RNAs involved in?
- Regulating RNA splicing - Gene expression levels - tRNA/rRNA maturation - Telomere synthesis - DNA synthesis - Parasite resistance (destroy parasite genome)
67
microRNAs
18-25 nucleotides Post-transcriptional gene silencing RNA interference
68
Small RNAs
``` 20-300 nucleotides Modification of target RNAs Synthesis of telomeric DNA Chromatin structure dynamics Transcription modulation Structural role ```
69
Medium and large RNAs
``` 300-10000 nucleotides DNA imprinting X-inactivation DNA demethylation Gene transcription Generation of other RNA classes ```
70
Potential for developmental regulation of cells by ncRNA
External signals DNA binding transcriptional activators and repressors Splicing Translational control Post-translational modification of proteins
71
How is miRNA produced?
Encoded as pri-miRNA Synthesised in the nucleus by RNA polymerase II then processed by Drosha Drosha produces a pre-miRNA from the pri-miRNA The pre-miRNA is exported to the cytoplasm In the cytoplasm, the pre-miRNA is cleaved by Dicer to produce double stranded miRNA This is then bound by the RISC complex, which removes the complementary strand, leaving just the protein-bound, mature miRNA This protein-bound miRNA recognises target mRNAs and regulates their activity (usually reduces their activity)
72
What is the role of Argonaute (Ago)?
Cleaves RNA complementary to the miRNA | Argonaute can also sequester miRNAs to stop them binding target RNAs
73
What do mutations in the Argonaute10 gene cause?
Damaged ago10 mutant protein cannot sequester miR166/165 so HD-ZiP genes are degraded by interactions with miR166 and AGO1, and the shoot meristem cannot develop
74
What are Yamanaka factors?
Can be used to change somatic cells into stem cells - induced pluripotent stem (iPS) cells
75
Examples of Yamanaka factors
Oct3/4 Sox2 Klf4 c-Myc
76
What miRNA mutation causes hearing loss in animals?
microRNA-96 (Diminuendo)

Cell and Developmental (4 decks)

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