Final Flashcards
1
Q
Is bicoid haplosufficient or haploinsufficient? What does this mean?
A
It is haplosufficient and therefore only one functional copy of the gene is required for normal function
2
Q
What is the Dorsal vertebrate homologue?
A
NF kB TF
nuclear factor kappa beta transcription factor
3
Q
What is the Toll vertebrate homologue?
A
Toll-like receptors
4
Q
What is the Gurken vertebrate homologue?
A
Epidermal Growth Factor (RTK ligand)
5
Q
What is the Torpedo vertebrate homologue?
A
Epidermal Growth Factor receptor (RTK)
- Important in immunology
6
Q
What is the Dpp vertebrate homologue?
A
Dpp = decapentaplegic
Homologue is BMP (paracrine factor)
7
Q
What is the Sog vertebrate homologue?
A
Sog = short gastrulation Homologue = chordin
A BMP inhibitor
8
Q
What is Twist?
A
A gene in flies that codes for a TF. The TF is necessary for mesoderm development
Dorsal turns on Twisted gene in the ventral part of the embryo (normally)
9
Q
What cells express the Torpedo (Epidermal Growth Factor receptor) in fly development?
A
The dorsal follicle cells
10
Q
What ligand binds to Torpedo receptors?
A
Gurken binds Torpedo on follicle cell membranes
Torpedo is a receptor tyrosine kinase
11
Q
Why isn’t Gurken in the ventral side of the embryo?
A
It’s mRNA was associated with the oocyte nucleus that was transported to dorsal, anterior portion of the nucleus - Once translated it diffuses, but the distance is too far for it to reach the ventral side
12
Q
Pipe
A
An enzyme made by the follicle cells
- Sulfates ventral vitelline proteins
Synthesis of Pipe is inhibited by RTK signalling (Gurken binding Torpedo on dorsal follicle cells)
13
Q
What are the three proteases in the big fly developmental pathway?
A
GD (gastrulation defective), Snake, Easter
Snake and Easter are zymogens
14
Q
What happens when cactus is phosphorylated?
A
It will be degraded and dorsal is then free to move into the nucleus
15
Q
What does the tube protein do?
A
It is an adaptor protein that helps Pelle bind the Toll-receptor
16
Q
What is the vertebrate homologue protein to Pelle?
A
IRAK
Interleukin Receptor-Associated Kinase
17
Q
Examples of BMP inhibitors?
A
Sog (flies), Chordin, Follistatin, Noggin, etc
18
Q
What does the blastocoel ensure during development?
A
It provide space between groups of cells to prevent mesoderm inducing signals from reaching the ectoderm.
19
Q
capacitation
A
The physiological changes the sperm must undergo within the female reproductive tract to become competent to fertilize an egg
- sperm must last 6 hours in the female before reaching the egg
- binds to cells in the oviduct and becomes hyper activated
- (bicarbonate, calcium - activate pKA)
20
Q
What is the zona pellucida?
A
Layer that surrounds the plasma membrane of the egg and early embryo. It consists of glycoproteins and proteins (ZP2 in humans where sperm binds)
Sperm binds here to begin fusion
21
Q
What is the acrosome reaction?
A
When the contents of the acrosomal vesicle are expelled through exocytosis - allowing sperm penetration and digestion through the zona pellucida
Exposes the proteins on the acrosomal membrane to the outside (of the plasma membrane. This allows interaction of Izumo with Juno and CD9
22
Q
What is the receptor for Izumo and where does this binding occur in the body?
A
Juno/CD9 is the receptor for Izumo and these proteins are present on the plasma membrane of the egg.
Izumo is present on the acrosomal membrane, and then the PM after the acrosomal reaction, in sperm.
Juxtacrine interaction
23
Q
What protein does the sperm bind to on human zona pellucidas?
A
ZP2 protein
ZP3 in mice
24
Q
How is polyspermy blocked in humans?
A
Enzymes released through the cortical reaction.
Here, ZP2 protein on the zona pellucida is cleaved so that it can no longer bind sperm.
25
What stage of meiosis in the egg paused in right before fertilization?
metaphase 2
26
Where does fertilization most often occur?
In the ampulla, but it can occur in the infundibulum
27
infundibulum
Part of the oviduct where fertilization can occur. However, it more often occurs in the ampulla
28
Amount of sperm in ejaculate versus the amount that get to the egg?
300 million versus 200
29
Where do sperm develop?
In the seminiferous tubules. Generated from puberty to death in males.
30
Acrosomal vesicle
Developed ventrally on the spermatid. It is a modified Golgi apparatus that contains proteolytic enzymes necessary to digest the zona pellucida surrounding the egg
31
Where are the mitochondria on the spermatid?
On the base of the head. They generate ATP that the sperm can use for movement
32
What is the germinal vesicle?
The large diploid nucleus of the primary oocyte (before meiosis)
33
Where are the cumulus/granulosa cells?
surrounding the zona pellucida
34
Chemical processes in sperm capacitation?
Bicarbonate influx causes activation of SACY - SACY then generates cAMP that activates PKA. PKA phosphorylates tyrosine kinases which phosphorylate sperm proteins -- sperm is capacitated!
35
How many types of glycoproteins are present on the surface of the zona pellucida and which one is active in binding sperm?
There are four: ZP1-4.
| ZP2 is the only one active in binding sperm.
36
What must occur for Izumo to be able to bind to Juno/CD9?
Exocytosis of the Izumo ligand present on the acrosomal membrane of sperm to the plasma membrane of sperm
37
What proteins are necessary for fertilization to occur?
Izumo ligand, Juno and CD9 (complex of receptors for Izumo)
38
Polyspermy Block in Humans
Membrane fusion between the sperm and the egg triggers exocytosis of cortical granules by the egg (the cortical reaction)
- digestive enzymes are exocytosed in the cortical granules and they cleave the ZP2 proteins on the surface of the zona pellucida
- This blocks anymore sperm from binding.
39
Does a change in membrane potential occur in the human block to polyspermy?
NO
This is part of the mechanism for the fast block to polyspermy seen in sea urchins. Fast block is not necessary as their is less sperm competition in mammals
40
Cortical Reaction
Exocytosis of cortical granules from the egg that occurs when the sperm fuses with the egg.
The vesicles which are exocytosed contain digestive enzymes that cleave the ZP2 proteins present on the zona pellucida.
Because ZP2 proteins are necessary for sperm binding, cleavage of these proteins blocks any more sperm from binding.
41
Does the zygote have a membrane-bound nucleus?
No. The membranes of the pronuclei dissolve - cells go directly into metaphase. So there is no membrane-bound nucleus at the zygotic stage
42
What is the mid-blastula transition (MBT)?
When the embryo begins existing on its own in terms of gene expression. Prior to the MBT, maternal genes are expressed.
- MZT occurs - Transcription of the zygotic genes begins to occur and maternal mRNAs are degraded
- cell cycle lengthens (gap stages are longer) and synchronous cell divisions cease.
43
MZT
maternal to zygotic transition
| when the zygotic genes begin to be expressed while the maternal mRNAs are degraded
44
Timing of MZT in different species
Drosophila - after the 13th cell division - at the 14th cell division (get MBT) - this is when the cells of the fly become cellularized
Frogs - ~14th cell division
Humans - ~3 days post fertilization. The embryo only has 4-8 cells. Mice are similar in that they have a longer time before MBT than other animals
Mic - ~2 cells stage
45
maternal effect mutations
mutations that affect early development. This is because the mom's genotype/genes are the ones being expressed at this time. Errors in her genes are what causes the developmental error
46
Are cell cycles shorter or longer before the MBT?
They are shorter. Early divisions are relatively rapid because there are no gap stages.
47
Cellularization in Flies
After the 13th cell cycle, each nucleus is surrounded by a cell membrane (when MZT occurs)
Creates cellular blastoderm
Must occur before gastrulation begins!
Driven by microtubules (create the furrow) and microfilaments (actin- ring constriction to pinch off cells)
As soon as cellularization occurs, you get formation of apical adherens junctions to hold the cells together as an epithelium
48
What is the ventral furrow in fly gastrulation?
An invagination (cell layer internalizes) of the ventral surface that internalizes ~1000 cells. These cells are precursors of the mesoderm. Pinches off to form mesoderm once the furrow closes
49
How is the endoderm created in flies?
Anterior and posterior midgut invaginations
50
germ band`
contains the cells that will form the trunk of the fly
the germ band forms from the cells which have gastrulated and undergo convergent extension (cells intercalate between each other to extend the germ band) at the ventral midline.
Forms the thoracic and abdominal segments
51
What is convergent extension?
Where cells intercalate between each other to extend a structure
ie. formation of the germ band
52
What are pole cells?
germ cell precursors (in flies) (oocytes or sperm)
53
What are the maternal effect genes present that get eventually translated to make morphogens?
Anterior - Bicoid and Hunchback
| Posterior - Caudal and Nanos
54
What are the proteins maternal effect genes that result in the anterior-posterior patterning of the embryo - pre-patterning of the embryo?
Bicoid, Nanos, Hunchback, Caudal
Bicoid and nanos protein gradients cause gradients to form in hunchback and caudal.
55
When are bicoid and nanos translated?
At fertilization. Their mRNA is distributed and anchored in the oocyte prior to fertilization.
56
Which genes are activated by MEG in the hierarchy of genes?
Gap genes
57
Gap genes
```
Activated by MEG
Expressed from the zygotic genome
Have broad expression domains
Expressed at the 12th cell division
mutants miss several segments in a row
```
58
Pair-Rule genes
Activated by the products of gap gene expression. Cause formation of 14 segment embryo.
Turned on at 13th cell division - expressed in a series of 7 stripes
mutants are missing portions of every other segment - can reduce the number of segments - ie they have 7 rather than 14 segments because they don't have boundaries.
- each stripe of expression corresponds to every second parasegment
Expressed in parasegments!
ex. fushi tarazu, even-skipped
All code for transcription factors!
58
Segment Polarity genes
Activated by the products of the pair-rule genes
Include the homeotic genes
Most downstream in the hierarchy of genes
Give identity to each of the segments formed
Get expression after the 14th cell division stage - at cellularization (can't signal before cell membranes are formed)
Get 14 stripes of expression
mutants - polarity of each segment being disrupted within each parasegment
ex. engrailed, wingless, hedgehog
59
Which cells express maternal effect genes?
The nurse cells of the egg chamber.
There are nurse cells and 1 oocyte that were all derived from the same cell. The posterior most cell becomes the oocyte.
The nurse cells supply proteins and maternal effect mRNAs that are necessary for the development of the future embryo THROUGH CYTOPLASMIC BRIDGES
60
Which type of cells surrounds the nurse cells and the oocyte?
Follicle cells
61
Oskar mRNA
Is present in the oocyte prior to fertilization.
| Associated with the kinesin motor protein and takes Nanos mRNA toward the + end (posterior) and anchors it here
62
Which motor protein is associated with oskar mRNA
kinesin
63
Which motor protein is bicoid mRNA associated with?
Dynein
64
How is bicoid anchored in the oocyte?
It is transported by dynein motor protein to the ANTERIOR part of the oocyte and anchored here
65
What occurs with respect to the proteins formed at fertilization in flies?
Bicoid and Nanos mRNAs are translated and diffuse to form gradients from anterior and posterior ends, respectively.
Get opposing morphogen gradients - syncytial specification
Depending on the concentration - alter the fate of the cell differentially
66
Are both bicoid and nanos TFs?
No, only bicoid protein is a TF
67
What happens if an embryo is homozygous mutant for bicoid?
They will develop normally, unless the mother of the embryo is a homozygous mutant for bicoid too.
68
What happens if the mother of an embryo is heterozygous for bicoid? homozygous null for bicoid?
Heterozygous - embryo should be fine because bicoid is haplosufficent
Homozygous null - embryo will develop without a head
69
What happens when bicoid is injected into the middle or posterior end of the embryo?
Middle - Get two tails, head forming in the middle
| Posterior - Get a head at the anterior and posterior ends
70
What does the distribution of hunchback and caudal mRNA appear like in the oocyte?
MEG
Their distributions are equal throughout the oocyte. No gradients!
When bicoid and nanos are translated at fertilization however - you then get a gradient of the hunchback and caudal proteins.
71
Protein gradient of hunchback?
MEG - mRNA is distributed equally throughout the oocyte
bicoid mRNA is translated at fertilization - because it is a TF - it can activate the expression of hunchback
So hunchback will be higher in the anterior, similarly to bicoid
72
What does bicoid protein do to caudal mRNA?
Binds it in the anterior (where it is most abundant) and represses its translation by preventing the ribosome from binding.
73
What does Nanos protein do in the posterior?
Nanos mRNA is tethered to Oskar which is an mRNA tethered to kinesin motor protein in the posterior.
At fertilization, Nanos is translated. Nanos protein associates with Pumilio to bind Hunchback mRNA in the posterior and prevent its translation by chopping off its PolyA tail
74
What does the Pumilio protein do?
It associates with Nanos protein in the posterior of the embryo to prevent the translation of Hunchback mRNA.
75
What family of protein is bicoid?
Homeodomain family of transcription factors
Bicoid also activates genes involved in developing structures
76
What are examples of gap genes in Drosophila?
zygotically-expressed genes
hunchback, krupel
The overlapping domain of these two proteins (TF) - they have mutually repressive effects on eachother's genes/function
Mutations in gap genes causes deletion of broad domains
77
What is the difference between a segment and a parasegment?
Parasegments are shifts 1/2 a segment forward relative to the segments.
There are 14 parasegments
A parasegment is made up from the cells in the posterior compartment of one compartment and the anterior compartment of the next segment.
78
Is the gene expression in the early embryo defined by segments or parasegments?
parasegments
Pair-rule gene expression spans a parasegment and therefore its expression spans the boundary of what will become a segment.
79
Where is even-skipped expressed?
Pair-rule gene
| Expressed in the odd parasegments
80
What is wingless and what does it bind to?
Wingless is a segment polarity gene that binds to Frizzled
| activates Wnt signalling
81
Which factors lead engrailed to be expressed?
When high concentrations of either evenskipped or fushi-tarazu proteins are present
82
What factors lead wingless to be expressed?
When neither fushi tarazu nor even-skipped is present
83
What parts of the parasegments/segments does engrailed and wingless mark?
Engrailed is present in the anterior part of the parasegments and posterior part of each segment
wingless is present in the posterior part of the parasegments and the anterior part of each segment.
84
What signalling occurs at the parasegment border?
Pair-rule protein presence leads to activation of engrailed expression. Engrailed codes for a TF which leads to expression of hedgehog. HH protein is secreted and diffuses to cells in the posterior region of adjacent parasegments which express Patched -- activates HH pathway in these cells and leads to activation of target genes - among these genes includes wingless -- get wingless expressed -- wingless is also a secreted protein -- it is released and diffuses to the cells on the anterior region of the adjacent parasegment (those which express HH and engrailed) and binds to Frizzled and initiates Wnt signalling.
HEDGEHOG AND WNT SIGNALLING
85
Where are the homeotic genes found in flies?
Give identity to the segments. There are two complexes that bear the homeotic genes in flies. Antennapedia complex and Bithorax complex
Their order on the chromosome corresponds to their expression domains in the embryo
86
What do you get with an ultrabithorax mutant?
2 sets of wings!
ubx is normally the most posterior homeotic gene expressed in the 3rd thoracic segment. So when the gene is no longer functional, the next most posterior gene in that region is expressed -- antennapedia - responsible for wing formation! get sets of wings on both the 2nd and 3rd thoracic segments
87
Antennapedia - gain of function mutation
If the enhancers get mixed up - antennapedia could function more anteriorly than it is normally expressed -> and because the posterior homeotic gene exerts its function - dominates - then you get legs forming where the antenna normally form! Normally get legs in 2nd thoracic segment
specifies the legs
88
What is the distribution of the dorsal mRNA in the embryo?
dorsal is a MEG
Both the dorsal mRNA and the dorsal protein are distributed evenly throughout the embryo. However dorsal is only translocated into the nucleus on the ventral side of the embryo.
89
Why is dorsal named the way it is?
If dorsal is mutated, you get a dorsalized embryo... all back tissue.
ie. need functioning dorsal to get a ventral side.
90
r
Gurken is transported to the dorsal/anterior of the oocyte with the nucleus
91
What is the amnioserosa?
Most dorsal tissue in fly embryo - extraembryonic tissue.
92
What do the cells with the highest concentration of Dorsal do in fly concentration?
These are the cells on the most ventral spot of the fly embryo. They will invaginate to form the mesoderm!
The high dorsal concentration in these cells turns on genes necessary for ventral development ie. Twist
93
What do cells expressing Twist do during gastrulation?
They invaginate to form the mesoderm (Highest dorsal concentration in these cells)
Twist expression assists with forming ventral structures.
94
Where is cactus normally degraded?
Cactus is normally degraded in the ventral cells of the drosophila embryo. Because cactus is degraded here, Dorsal is then able to enter into the nucleus of these ventral cells
95
Which region of the oocyte is the nucleus moved to in Drosophila development?
The dorsal/anterior part, along with Gurken mRNA.
96
When is gurken mRNA translated and what does the protein do once it is translated?
Gurken is translated at fertilization. It then is secreted and binds to Torpedo (RTK) on the DORSAL follicle cells surrounding the zygote
**too far of a distance for the gurken protein to diffuse to the ventral follicle cells surrounding the zygote
97
Which cells bear the Torpedo receptor?
Follicle cells surrounding the oocyte/zygote.
The torpedo receptor is a RTK -- Gurken protein binding at fertilization activates the RTK pathway - causes the follicle cells to differentiate to a dorsal morphology.
98
What effect does does Gurken translation have on Pipe?
When gurken is translated - stimulates the RTK pathway in follicle cells by binding the Torpedo RTK receptor -- this causes INHIBITION of Pipe expression in dorsal follicle cells
99
Do dorsal or ventral follicle cells synthesize Pipe protein?
Ventral follicle cells because pipe expression is not inhibited by Gurken binding ventrally.
100
What is the vitellin envelope?
It is an extracellular matrix surrounding the Drosophila oocyte - similar to the zona pellucida in vertebrates
