Development L1/2

Question 1

What are major qs in development?

Answer 1

does every cell in body encode the same genetic info??
How are genes switched on and off if not?
How differentiation organised in space and time for pattern formation

Question 2

What is an eg. of differentiation?

Answer 2

Cells of frog somite express muscle myosin, neighbouring cells in skin and neural tube don’t
(stain for myosin w antibody)

Question 3

What are egs of pattern formation?

Answer 3

1) At regular intervals cyanobacterium Anabaena forms big hetercyst cells for N2-fixation
2) Hydra has head with tenticles and mouth to injest prey and at the other end a foot to attach to substrate, if cut in half, cut face of foot end generates new head and head cut face end generates new foot

Question 4

What is xenopus?

Answer 4

An African frog commonly studied as model organism

Why: big ol embryo

Question 5

Xenopus egg cell divides to produce…

Answer 5

Ball of cells called blastula

Question 6

How can we follow fate of single cells in blastula?

Answer 6

Inject single cell with fluorescent dye

Question 7

C3 cell line progeny… (In Xenopus)

Answer 7

Contribute to muscle and skin

Question 8

What is a fate map?

Answer 8

Early embryo mapped with what each cell will become in final embryo

Question 9

Draw xenopus basic fate map

Answer 9

(see notes for diagram)

Question 10

What was the first experiment in the late 19th century and why was it flawed?

Answer 10

Hot pin killed 1/2 cells formed by first frog division, only half of embryo develops, conclusion info to make embryo partitioned at each cell
Flawed: dead cell not removed so normal development blocked on that side (spatial constraints)

Question 11

What was the next important development experiment?

Answer 11

Sea urchin (spiky boi) embryo first 2 cells formed by division separated and cultured individually, any of these cells alone is enough to form entire sea urchin embryo but with smaller vol bc half the cytoplasm
Thus system is able to organise and reorganise construction even when cut in half
Old experiment result that info partitioned at each cell proved wrong

Question 12

Describe Josh Gurdon nuclear transplantation experiments

Answer 12

(see notes for diagram)
Showed nucleus of differentaited cell capable of orchestrating whole tadpole development, clearly genetic info not lost as cells differentiate
Cell differentiation depends on selective expression of parts of a full gene set

Question 13

What are intrinsic and extrinsic signals?

Answer 13

(see notes for diagram)

Question 14

Describe chick erythrocyte cancer blood cell experiment

Answer 14

Chick erythrocytes are quiescent

If fused with human cancer cell blood cell nucleus enlarges and chick-specific proteins get synthesised in new cell

Question 15

What happens w human amniocyte and rat muscle cell if fused?

Answer 15

Expression of human liver-specific genes turned off and human muscle-specific genes turned on (see notes for diagram)

Question 16

How can we find out what the muscle cytoplasm factors are?

Answer 16

Transinfect cells with cDNA made from mRNA made from proliferating myoblasts
Single cDNA for gene MyoD (myogenic differentiation gene) makes cells switch fate and become muscle cells eg. fibroblasts (collagen-making) switch fate

Question 17

What does MyoD encode?

Answer 17

Transcription factor (a helical loop) that binds to upstream sequences of mucle-specific sequences and activates them
Also activates its own transcription via positive feedback maintaining expression of muscle-specific cells and stopping cells going backwards in development

Question 18

What means a differentiated state is very stable?

Answer 18

More TFs than MyoD, complex gene regulatory network with multiple feedback loops

Question 19

Where is Myo D conserved?

Answer 19

Frog, Mouse human… (often in development genes are conserved)

Question 20

What is extra evidence that MyoD is a master regulator also needed by adults?

Answer 20

MyoD also expressed in adult skeletal muscle

Question 21

What is in situ hybridisation?

Answer 21

Technique like probes but anti-sense RNA bind sense RNA, RNA probe fluoresently labelled

Question 22

What happens to egg cell at fertilisation?

Answer 22

Before egg is developmentally paused (transcriptional quiescence), on fertilisation this is released and there is rapid cell division called cleavage

Question 23

How does xenopus and drosophila development differ?

Answer 23

Xenopus ball of cells
Drosophila nuclei first replicate without cell division, then migrate to periphery, then each nucleus gets surrounded by cell membrane. The cleavage process is programmed by maternal gene products. (see notes for diagram)

Question 24

At what cell cycle in xenopus does transcription of zygotic genes begin ? What is this called?

Answer 24

(see notes for diagram), cycle 12, called the mid-blastula transition

Question 25

How can you show that proteins and RNA are unevenly distributed in the egg cell?

Answer 25

in situ hybridisation and antibodies (often shows they get partitioned by cleavage)

Question 26

What happens to Vg1 transcripts?

Answer 26

Fluorescence shows they are localised to vegetal pole of frog egg

Question 27

What happens in C. Elegans egg?

Answer 27

Asymmetric distribution of protein granules before it divides, protein granules go asymmetrically into the germ cells (future gametes)

Question 28

Describe drosophila cytoplasmic transfer experiment

Answer 28

(see notes for diagram)

Question 29

What is the only totipoetent cell type?

Answer 29

Egg cell

Question 30

What do you put in development intro

Answer 30

Branching diagram of differentiation from totipotent cell, branching=decision made, responding to signals

Question 31

How do you activate an egg cell?

Answer 31

Promote calcium release or put in calcium buffer, wave of calcium triggers first division
(OR electric shocks)

Question 32

How are scientists trying to shunt cells back up their differentiation pathway?

Answer 32

Overexpress 4x TFs, in theory fibroblasts could be undifferentiated then made to differentiate to replace damaged liver cells

Question 33

What is the big problem with this?

Answer 33

Cells created are aged cells (making them differentiate involves lots of divisions) possibly with shorter telomeres and could harbour mutations
Also there could be mutations in liver genes that wouldn’t be a problem in normal fibroblasts
(need to identify potentially dangerous patterns which requires time and even more divisions)
Process still long way off

Question 34

The more differentiated a cell is…

Answer 34

The harder it is to reprogramme eg. neurons v low % reverse differentiation
Ideally reprogramme stem cell population eg. CSF with haemopoetic stem cells in