Development L1/2 Flashcards
What are major qs in development?
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
What is an eg. of differentiation?
Cells of frog somite express muscle myosin, neighbouring cells in skin and neural tube don’t
(stain for myosin w antibody)
What are egs of pattern formation?
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
What is xenopus?
An African frog commonly studied as model organism
Why: big ol embryo
Xenopus egg cell divides to produce…
Ball of cells called blastula
How can we follow fate of single cells in blastula?
Inject single cell with fluorescent dye
C3 cell line progeny… (In Xenopus)
Contribute to muscle and skin
What is a fate map?
Early embryo mapped with what each cell will become in final embryo
Draw xenopus basic fate map
(see notes for diagram)
What was the first experiment in the late 19th century and why was it flawed?
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)
What was the next important development experiment?
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
Describe Josh Gurdon nuclear transplantation experiments
(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
What are intrinsic and extrinsic signals?
(see notes for diagram)
Describe chick erythrocyte cancer blood cell experiment
Chick erythrocytes are quiescent
If fused with human cancer cell blood cell nucleus enlarges and chick-specific proteins get synthesised in new cell
What happens w human amniocyte and rat muscle cell if fused?
Expression of human liver-specific genes turned off and human muscle-specific genes turned on (see notes for diagram)
How can we find out what the muscle cytoplasm factors are?
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
What does MyoD encode?
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
What means a differentiated state is very stable?
More TFs than MyoD, complex gene regulatory network with multiple feedback loops
Where is Myo D conserved?
Frog, Mouse human… (often in development genes are conserved)
What is extra evidence that MyoD is a master regulator also needed by adults?
MyoD also expressed in adult skeletal muscle
What is in situ hybridisation?
Technique like probes but anti-sense RNA bind sense RNA, RNA probe fluoresently labelled
What happens to egg cell at fertilisation?
Before egg is developmentally paused (transcriptional quiescence), on fertilisation this is released and there is rapid cell division called cleavage
How does xenopus and drosophila development differ?
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)
At what cell cycle in xenopus does transcription of zygotic genes begin ? What is this called?
(see notes for diagram), cycle 12, called the mid-blastula transition
How can you show that proteins and RNA are unevenly distributed in the egg cell?
in situ hybridisation and antibodies (often shows they get partitioned by cleavage)
What happens to Vg1 transcripts?
Fluorescence shows they are localised to vegetal pole of frog egg
What happens in C. Elegans egg?
Asymmetric distribution of protein granules before it divides, protein granules go asymmetrically into the germ cells (future gametes)
Describe drosophila cytoplasmic transfer experiment
(see notes for diagram)
What is the only totipoetent cell type?
Egg cell
What do you put in development intro
Branching diagram of differentiation from totipotent cell, branching=decision made, responding to signals
How do you activate an egg cell?
Promote calcium release or put in calcium buffer, wave of calcium triggers first division
(OR electric shocks)
How are scientists trying to shunt cells back up their differentiation pathway?
Overexpress 4x TFs, in theory fibroblasts could be undifferentiated then made to differentiate to replace damaged liver cells
What is the big problem with this?
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
The more differentiated a cell is…
The harder it is to reprogramme eg. neurons v low % reverse differentiation
Ideally reprogramme stem cell population eg. CSF with haemopoetic stem cells in
