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CTO 2 > 3. Cell differentiation > Flashcards

3. Cell differentiation Flashcards

1
Q

Define differentiation

A

Differentiation - a process by which unspecialised cells become cell specific types (in embryos / in tissue homeostasis)

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2
Q

Approximately how many types of cell tissues there are in a human?

A

~ 200 main types

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3
Q

What makes cells different from one another?

A
  • different macromolecules
  • different metabolites
  • different morphologies and behaviours
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4
Q

Housekeeping vs specialised proteins?

A
  • Housekeeping proteins - proteins found in most cell types for shared essential cell functions (ex ATP synthase)
  • Specialised proteins - proteins specific to a particular cell type (ex insulin synthesising protein in beta cells in the pancreas)
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5
Q

What are the main embryonic germ layers from which all tissues differentiate?

A
  • ectoderm
  • mesoderm
  • endoderm

Trophectoderm (embryo tissue which forms placenta)

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6
Q

Why is differentiation considered progressive?

A
  • forms intermediate (transient) states (progenitor / precursor cells)
  • has a terminal differentiation point - final cell from
    => progressive specialisation, not immediate
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7
Q

What drives specific cell differentiation?

A

Differentiation is driven by interplay between cell’s lineage and its environment

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8
Q

During which process of embryo development are the germ layers formed?

A

Gastrulation

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9
Q

What is haematopoiesis?

A

Haematopoiesis - formation of blood components - differentiation of blood cells

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10
Q

What genetic mechanism drives cell differentiation?

A

Selective activation / inactivation of particular genes

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11
Q

Can the nucleus of a differentiated cell support development of a new organism?

A

Yes - nuclear transfer experiment - sheep Dolly (first cloned mammal)

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12
Q

What is gene constancy?

A

Gene constancy - all cells in a multicellular organism have a full complement genes (full genome)
=> cells differentiate because of gene activity rather than gene content (differential gene expression)

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13
Q

How is gene expression controlled?

A

By control of gene transcription - transcription factors (TF) - genes can individually be switched on/off - different gene expression patterns in different cells

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14
Q

What is the DNA signal sequence which is a recognition site / binding site?

A

TATA box - binds TATA binding proteins - transcription factors (TFIID)

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15
Q

What is the role of TATA box?

A

TATA box controls where transcription starts - binds TF
(doesn’t control when transcription starts)

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16
Q

What is an enhancer?

A

Enhancers - short regulatory nucleotide sequences recognised by TF that enhance the rate transcription

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17
Q

What is a promoter?

A

Promoters - rather long regulatory nucleotide sequences that initiate transcription - polymerases bind

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18
Q

Explain promoters vs enhancers

A

Promoters - long sequences / initiate transcription
Enhancer - short sequences / increase transcription rate

Enhancers activate promoters

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19
Q

What are transcription factors (TF)?

A

Transcription factors (TF) - proteins which activate/repress polymerase function

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20
Q

What are the methods by which TFs can initiate transcription?

A
  • can act directly (directly recruit RNA polymerase to TATA box)
  • can act via chromatin modification (indirectly recruit RNA polymerase: histone acetyl transferase / chromatin-remodelling complex)
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21
Q

What are the two indirect methods of TFs for recruiting RNA polymerase for transcription initiation?

A
  • Histone acetyl transferase (HAT): acetylations loosens histone interactions with DNA - genes more accessible for transcription
  • Chromatin-remodelling complex: chromatin modifying enzymes promote RNA polymerase binding and function by remodelling chromatin and making it more accessible
22
Q

What controls gene’s transcriptional activity in a cell?

A
  • binding sites in enhancer sequences of DNA for the gene
  • presence of appropriate TFs for the gene
23
Q

Give two examples of TF in differentiation

A
  • MyoD - TF which activates expression of genes for muscle myosin II protein - in muscle cell differentiation - recognition site: E box
  • GATA1 - TF which activates expression of several target genes (for α+β globin, erythropoietin receptor, heam biosynthesis enzymes, spectrin) - in RBC differentiation
24
Q

Can one TF target several genes transcription?

A

Yes - MyoD activates transcription of many genes acting in muscle cell differentiation + represses non muscles genes

25
What's the effect of MyoD knockout?
MyoD knockouts produce undifferentiated muscle cell precursors - **myoblasts**
26
How can MyoD expression pattern be investigated?
Modify embryos with **MyoD mRNA staining** (in situ hybridisation) - observe MyoD **expression patterns** where stained (muscle cell differentiation sites)
27
Can TF force other lineage cells to differentiate into lineage to which the TF is specific?
**Yes** - ex: fibroblasts (skin precursors) transfected with MyoD containing plasmid - differentiated into muscle cells
28
What are the correct steps in genetic analysis of TF function?
1. Analysis of **expression pattern** (where it is expressed in developing body?) 2. Analysis of **loss-of-function** (is the TF required for differentiation?) 3. Analysis of **gain-of-function** (can the TF force other lineage cell differentiation into its own - is it sufficient for full differentiation?)
29
How TFs recognise their binding sites?
TF - high specificity - specific am. a. in recognition site - bind **s****pecific base sequence**
30
How are TFs categorised?
TFs are categorised **into families** - based on **3D structure** of their DNA binding domains where binding sites are located
31
Usually, do TFs work on their own or in combinations?
In combinations - **groups of TFs** at binding sites
32
What are the possible TF combination interactions for transcription enhancing?
When several TFs for the same gene regulation: - either TF may be sufficient / **doesn't matter which** binds for transcription - **all TFs required** together for transcription
33
What are the possible TF combination interactions for transcription repressing?
When several TFs for the same gene regulation: - repressing TF may **prevent binding of activating TF** (picture) - recruit proteins that **tighten chromatin** - gene less accessible
34
What regulates TFs and how?
What: - **envronmental signals** (signals from other cells: hormones, growth factors) - **developmental history** (earlier TFs regulate later TFs) How: - controlling their activity by **PTMs** - controlling **their gene expression**
35
How are TFs regulated by cell signalling?
36
How are TFs regulated by PTMs?
PTMs - **ex: phosphorylation** - phosphate alters protein conformation -> activate / inhibit TF
37
How growth factors can control TF activity?
Growth factors regulate TF activity **by phosphorylation** -> target gene activity changed Growth factor signal activates **protein kinase cascade**: 1) phosphorylation of **MAP-kinase-kinase kinase** (activated) -> 2) phosphorylates **MAP-kinase kinase** (activated) -> 3) phosphorylates **MAP kinase** (activated) -> 4) MAP kinase enters nucleus and phosphorylates TFs - **activates TFs for target gene**
38
Explain how epidermal growth factor activates cyclin genes
Epidermal growth factors - mitogen (promotes cell proliferation) 1) Growth factors signal **initiates kinase cascade** - **p****hosphorylates MYC** (TF) on serine (Ser-62) + threonine -> changes MYC conformation to stable active 2) Activated MYC drives **cyclin gene transcription**
39
Explain how hormone erythropoietin (EPO) regulates GATA1 activity
1) Hormone erythropoietin (EPO) **secreted from kidneys** 2) **EPO** binds to receptors - kinase cascade 3) **GATA1** (low activity) phosphorylated - conformational change -> high activity 3) GATA1 increases **DNA binding** 4) Stimulates **blood progenitor proliferation** and **RBC differentiation** Low O2 conditions (ex: hipoxia) stimulate EPO secretion - more RBCs - higher efficiency
40
Why control of TFs is important for orderly progression of differentiation?
TF regulate **gene expression** which drive differentiation -> cell fate: Different TFs regulate different steps in differentiation
41
Explain how leaf progenitor cells differentiate into epidermis and stomata?
Default pathway - epidermis cells Some **progenitor cells induced for stomata** (guard cells) differentiation by **SPCH, MUTE, FAMA** TFs
42
What is a gene regulatory network (GRN)?
Gene regulatory network (GRN) - a set of **genes which interact** to control a specific cell function
43
How is MyoD initially activated in muscle differentiation gene regulatory network?
By **c****o-regulation of Pax3 and Myf5** TFs (work in combination) - for initial MyoD levels - later MyoD activates its own expression Pax3 + Myf5 - transient process
44
How is MyoD regulated in muscle differentiation?
**Autoregulation** - MyoD activates further MyoD expression - becomes **independent** of Pax3 and Myf5 regulation (cell memory)
45
How do growth factors stop MyoD expression?
1) Growth factors promote cell division -> cdk produced 2) Cdk **phosphorylate MyoD and Myf5** -> proteolytic degradation -> **muscle cell differentiation inhibited** -> myoblast proliferation enhanced (growth factor response - cell division achieved)
46
What is RNA interference?
RNA interference (RNAi) - **post transcriptional gene silencing** - degrade mRNAs
47
How is Pax3 degraded during muscle differentiation?
Via **RNA interference** (RNAi): - Pax3 only for early muscle development - must be down-regulated for later stages to proceed - Pax3 mRNA degraded by microRNA (miR-1) - double stranded RNA - bound to an enzyme
48
Explain Waddington's developmental landscape
Represents **cell specialisation** - **binary fate decisions** (which valley will roll into - increasing specialisation - decreasing potency to develop into any other cell type
49
Explain pancreas cell differentiation progress
Binary cell fate choices: specific cells from the gut can choose 1/3 cell lineages - **pre-pancreatic** - **Ngn3 TF needed** for **all endocrine cells** (if Ngn3 missing - no endocrine cells developed - all exocrine) - Ngn3 needed for **general endocrine precursor development** but not for a specific cell type in endocrine cells
50
What decides pre-pancreatic fate in foregut?
- **Pre-pancreatic region** affected by TFs - loops formed - comes closer to notochord - receives signalling - **Fgf2** TF turned on -> further pancreas development
51
Explain erythroid/myeloid cell fate choice
Common myeloid progenitor - **binary cell choice** -> **G****ATA1** / **PU.1** TFs GATA1 / PU.1 **autoregulate and inhibit each other** -> different blood cells differentiate - **ANTAGONISTIC interactions**
52
What are master regulators of organogenesis, give example
Master regulators - TFs which are **essential for the development of all organ** - without it nothing formed - if i**njected into wrong location - organ still formed** Example: - Pax6 in eye development - same TF in many animals - was present in a common ancestor - MyoD in muscle differentiation - Shh in anterior-posterior axis development

CTO 2 (8 decks)

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