- How is the patterning established during early Drosophila development? (Long answer: talk about different genes involved in pattern formation, starting with the maternal polarity genes. Include the consequence of gene mutations.)
a. Patterning is not determined by site of sperm entry but rather by interactions that occur in the ovary between the egg and nursing and follicular cells; those interactions will influence location of maternal determinants.
b. Maternal polarity genes
i. bicoid and hunchback regulate anterior structures.
ii. nanos and caudal regulate posterior structures.
iii. Maternal egg polarity genes regulate zygotic Gap genes; expressed in an overlapping pattern.
iv. The expression of gap genes regulates the transcription of the Pair-rule genes further dividing the embryo.
v. Finally, the pair-rule genes regulate the segment polarity genes
c. Gap genes
i. Map out basic subdivisions along the embryo’s anterior-posterior axis
ii. Mutations cause “gaps” in the animal’s segmentation
d. Pair-rule genes
i. Define pattern in terms of pairs of segments
ii. Mutations result in embryos having half the normal number of segments
e. Segment polarity genes
i. Set the anterior-posterior axis of each segment
f. Mutations produce segments where part of each segment is missing
i. Bicoid missing = no head, acron, thorax,
- When are A-P and D-V axes specified during Drosophila development? How is this different (or not) from C. elegans (or Sea urchin) early development?
a. Occurs during oogenesis, which is before fertilization
i. By patterning, interactions between egg and follicular cells
b. Different from sea urchin and C. elegans as it occurs when in cleavage/blastulation
- What are the differences between two different types of blastoderm formed sequentially during early Drosophila development?
a. Syncytial blastoderm
i. Most nuclei are at the periphery but still with a common cytoplasm
b. Cellular blastoderm
i. Membranes grow inwards to separate each nuclei into single cells
- Define germ band (in respect to germ layers and function).
a. Germ band (ventral blastoderm) starts to elongate→ posterior end of the embryo with the pole cells comes to the dorsal side of the egg
b. parasegments; appear at the extended germ band
c. Parasegments are transient; segments form from the posterior 2/3 of one parasegment and anterior 1/3 of the following parasegment
d. Germ band retracts; epidermal grooves rearrange themselves to form definitive segments
- What are the types of genes involved in determination of anterior-posterior axis (anterior-posterior pattern formation) in Drosophila?
a. Special follicle cells/ border cells determine the axis
b. Two sets of maternal egg polarity genes in the egg, expressed in different regions into transcription and/or translation factors regulate the expression of genes within these regions and in between.
c. bicoid and hunchback regulate anterior structures.
d. nanos and caudal regulate posterior structures.
- The graph and picture below show the distribution of the bicoid protein during the early Drosophila development. Is bicoid protein acting as a transcription or translation factor at this point? Explain your reasoning. What does becoid protein control (expression of which proteins)?
a. It’s a translation factor in the early stage as the later it enters the cells for activation of hunchback
b. Bicoid represses translation of maternal mRNA for protein Caudal in the anterior - it works as translation factor in this stage
c. Generates caudal protein gradient (P-A) in syncytial blastoderm
- Where are bicoid and nanos mRNAs produced during Drosophila oogenesis and where are they localized in the early embryo?
a. They are in nurse cells
b. Bicoid Localized in anterioir end of oocyte and nanos in the posterior
- You isolate embryos laid by a female Drosophila that lacks both copies of the bicoid gene (homozygous mutant). Describe (in 1-3 sentences or with a clearly labeled diagram) what would be the spatial distribution of caudal protein that is made from the corresponding maternal caudal mRNA in these mutant embryos at the syncytial blastoderm stage. Explain in additional 1-3 sentences how and why this distribution is different from that seen in normal embryos
b. The caudal would then be in the posterior
- Thinking question (possible bonus): You inject synthetic bicoid mRNA into the middle of the egg laid by female homozygous mutant for a bicoid. Describe (in 1-3 sentences or with a clearly labeled diagram), what do you expect to be the spatial distribution of caudal protein at the syncytial blastoderm stage that is made from the corresponding maternal caudal mRNA in these embryos. Explain how this distribution is achieved in additional 1-3 sentences.
a. Caudal is surrounding the bicoid
b. Creates two gradients as its in the back ground
- What are the major posterior determinants in an early Drosophila development?
a. The organizing center acts through a gradient of Nanos protein. Nanos blocks the translation of maternal hunchback mRNA in the posterior. Caudal is expressed in posterior.
- What is the terminal embryonic determinant in an early Drosophila development? What is the consequence in the homozygous mutant for this protein?
a. Torsal protein, activates gap genes
b. Formation of the acron and tel wont occur
- Compare syncytial with autonomous and conditional cell specifications in respect to interactions between cells (be careful here, it’s somewhat a tricky question).
Autonomous: specification by differential aquisition of certain cytoplasmic molecules present, btwn itself by migration
Conditional: interaction btwn cells, allow for different functions
Syncytial: interaction between cytoplamic regions prior to cellularization of blastoderm
- What are homeotic genes and what is their role?
a. ultimately activated by the positioning signals provided by different morphogen combinations in different segments – define what is going to become of each segment
- What are the two gradients that establish dorsal-ventral polarity in the Drosophila embryo? Briefly describe how the % (level) of ventralization is controlled.
a. dorsally positioned ligand called Gurken and ventrally positioned transcription factor Dorsal (both synthesized from maternal mRNA).
b. Dorsal induces the transcription of genes responsible for ventralization and inhibits others that (if transcribed) specify a dorsal fate
c. The amount of Dorsal determines the % of “dorsalization” (ventralization)
d. Large amount of Dorsal→ cells will become mesoderm
e. Smaller amount of Dorsal→ glial or ectodermal tissue
- How is the dorsal-ventral polarity determined in the Xenopus embryo?
a. Ventral side is determined by point of sperm entry as it triggers the microtubules separating the cortical and internal cytoplasm
b. Dorsal side – where clear cortical cytoplasm replaces pigmented cortical cytoplasm (grey crescent)
- What is a grey crescent? What is the importance of a grey crescent?
a. Where the first cleavage furrow bisects
b. Dorsal part of embryo
c. Right on top of blastopore lip
- What are the two most important purposes of a blastocoel (in general)?
a. Allows for cell migration during gastrulation
b. Prevents early cell-cell interactions
- What is a regulative development? What is autonomous development? How is the change from regulative to autonomous development demonstrated (or rather proven) during Xenopus gastrulation?
a. regulative development (in other words, cells’ fates are conditional):
i. 1) An individual cell (early in development) can develop into cell lineages beyond its normal fate – nuclear equivalence.
ii. 2) A cell’s fate is determined through interactions with neighboring cells (induction/competence) – inductive interactions
b. Early gastrula – regulative development:
Conditional cell fate (=dependent); cell fate WILL change if exposed to different environment
c. Late gastrula – mosaic development:
Autonomous cell fate (=determined or independent); cell fate is set – change in environment cannot influence it anymore
d. Proved through transplantation where the epidermis vs neural plate is formed
- How would you define dorsal blastopore lip, Spemann’s organizer and Nieuwkoop’s center? What is the relationship between those “organizers”? Use a diagram (don’t forget to label d-v orientation, dorsal and ventral parts of the embryo, grey crescent, point of the sperm entry etc.).
a. dorsal blastopore lip as the organizer. They transplanted cells from the dorsal lip of one species onto a second embryo, on the opposite the site of the recipients dorsal lip.
b. Cells from the donor’s dorsal lip:
c. 1) Induced the host’s ventral cells/tissues (opposite side from the dorsal lip) to change their fates and to get involved in forming the second neural tube and dorsal mesoderm tissues instead of epidermis
d. 2) Organized host and donor cells/tissues into forming a secondary embryo with clear dorsal-ventral and anterior-posterior axis
f. The dorsal blastopore lip OR Spemann’s organizer itself is INDUCED by the dorsal most vegetal cells of the blastula (=the vegetal cells positioned at the very dorsal end); those cells have been called the Nieuwkoop Center; in other words, Nieuwkoop Center induces (establishment of) the organizer.
g. Spemann’s organizer is frequently called (just) organizer
- What is beta-catenin (…. factor)? What does that imply? What is the role of -cateninn during early Xenopus development? When did we mention -catenin before? Was the role of -catenin in that organism similar or not?
a. Before fertilization β-catenin is expressed from maternal mRNA throughout the embryo
b. After fertilization - β-catenin is in the dorsal part of the embryo; protein Vg1 is in the vegetal part (also coded by maternal mRNA); both important in defining the Nieuwkoop center.
c. Accumulation covers both the Nieuwkoop center and the organizer regions.
d. Before fertilization GSK-3 inhibitors Dsh (Disheveled), GBP and ligand Wnt are in vesicles at the most vegetal part of the egg;
e. After fertilization those vegetal vesicles are translocated towards the dorsal part of the embryo
f. Dsh, GBP and ligand Wnt are released and distributed throughout the future dorsal 1/3 of the 1-cell embryo
g. Dsh and GBP bind to GSK-3 and block its action so that the b-catenin is not degraded in those areas (but it is degraded in the future ventral part as Dsh and GBP do not reach that far);
h. Nuclei of blastomers in the dorsal region receive high b-catenin (gradient is formed)
i. Mentioned in both chicken and xenopus
i. Telolecithal (delamination, intercalation, convergent extension) vs mesolecital eggs (invagination and involution)
