Lecture 32 - Developmental genetics II Flashcards
Why mRNAs don’t diffuse freely like proteins, in the syncytium ?
Are bound to specific proteins that target them to a specific place
What’s the point of having enhancers and silencers sequences near to each other (like in eve gene for example)
When repressor concentration is high, it will block the enhancer effect
Segment polarity genes: 2 functions
- Encode components of two cell-cell signaling pathways (Hedgehog and Wingless)
- Define A and P within each segment
Segment polarity genes: What kind of protein
TFs, secreted proteins, membrane receptors, kinases, …
Segment polarity genes: What regulates their transcription
Pair-rule genes
What happens in segment polarity genes mutants
Segments have mirroring of one half of the segment
Cascade of TFs to regulate early drosophila embryogenesis. 5 types of genes and function
Egg-polarity genes (maternal-effect): A-P axis
Gap genes: Broad subdomains
Pair-rule genes: Individual segments identified
Segment polarity genes: Define polarity of individual segments
Segment identity genes (homeotic genes): Define identity of individual segments
(notes) what genes are responsible for forming 14 equivalent segments + what we mean by serially reiterated segments
Segment polarity genes. Serially reiterated = have equal capacity of becoming different parts of the body
Homeotic mutations meaning and name of gene category identified
Mutations in segment identity genes. Hox genes
What happens in homeotic mutants (name the phenomenon)
A particular structure is replaced by another one normally found somewhere else (homeotic transformation)
2 examples of homeotic mutations
Ultrabithorax (Ubx, a TF) : second thorax and set of wings replace halteres
Antennapedia (Antp): leg in place of antenna
Number of homeotic genes (hox genes) identified in drosophila + type of protein and common characteristic
- all encode homeodomain family transcription factors
What’s a homeodomain
A DNA binding domain with a helix-helix structure
Where the hox genes are found in the drosophila genome and particular characteristic
In 2 gene complexes (antennapedia complex and bithorax complex) on third chromosome. Colinearity: Aligned in chromosome in same order as body parts when their function is needed
What genes activate/repress hox (or selector) genes
Gap genes and pair-rule genes
Pattern of expression of hox genes
Each is expressed in specific segments and some overlap
How is hox genes transcription regulated once they are commited
Through epigenetic regulation (their faith is maintained during development) methylation or acetylation depending on segment
What will ultimately determine the structure that a segment develops
The specific hox genes that are expressed in this segment
Developing wing and developing haltere: where is Ubx expressed
Not expressed in wing. Expressed in haltere
What happens at the level of the Ubx gene in the ultrabithorax mutant
Ubx is mutant so halteres develop into a second pair of wings and thorax
How was the Ubx gene linked to the ultrabithorax mutant phenotype (how was its function uncovered ?)
Forced expression of Ubx in wings and it converted them to halteres
Name of portions of body structure that stick out of the body and hox gene that controls that
appendages (antennae, legs, wings, halteres). Gene distal-less (Dll)
When is dll expressed in appendages and in abdominal segments
Expressed in earliest stage in appendage development (about 20 cells)
Not expressed in abdominal segments
What repressed Dll expression in the abdomen and type of protein
Ubx and Abd-A (abdominal A) Hox genes. ARE TRANSCRIPTION FACTORS
In double mutants (Ubx - and Abd-A -) what happens to Dll expression in the abdominal segments
Dll is derepressed and expressed in abdominal segments
In Ubx mutants, what happens to Dll expression
Dll is derepressed in abdominal segment A1 and is expressed there
Regulatory sequences in the Dll gene
Has binding sites for several TFs, including 2 for Hox genes
What happens when the Dll gene’s regulatory sequences that are binding sites for Hox genes are mutated
Dll is not repressed and is expressed in abdominal segments
What are the Dll gene regulatory sequences other than the binding sites for Ubx and Abd-A
Binding sites for segment polarity genes: sloppy-paired (slp) - anterior (halves of) segments. engrailed (en) posterior (halves of) segments
What is minimally required to repress dll expression in abdominal segments
- Ubx OR Abd-A
- Slp OR En
How do Ubx, Abd-A, Slp and En TFs work to repress Dll
Cooperative regulation
What happens when Slp is mutated
Dll is derepressed in segments aA1 to aA7 (a meaning the anterior half of these segments)
What happens when En is mutated
Dll is derepressed in segments pA1 to pA7 (p meaning the posterior half of these segments)
What happens when Slp and En are mutated
Dll is derepressed in segments A1 to A7
Which genes function to define A and P within each segment (2 examples of these) and effect of mutations in these.
Segment polarity genes. Mutation = mirror effect in segments. Examples = gooseberry and patched genes
Summary of effects of Ubx mutant (2)
Halteres develop into second thorax and wing pair
Dll is derepressed in segment A1 (Abd-A still present to repress Dll in segments A2 to A7. Ubx represses in 1 to 7)
Hox genes conservation on other species
Conserved in mammals. All duplicated twice. 4 Hox gene clusters in mammals.
Hox genes characteristic in mammals that is similar to to flies’
Colinearity. The order of hox genes on the chromosome parallels the order of body parts in which they are expressed
Example of serially repeated structure in vertebrates
Vertebraes
Normal number of lumbar and sacral vertebrae in vertebrates
6 lumbar and 4 sacral
2 types of mutants causing changes in vertebraes identity in vertebrates
Mutant in Hoxd11 (hox gene) has an additional (7) lumbar vertebra
Double mutant in Hoxd11 and Hoxa11 (hox genes) has 2 additional (8) lumbar vertebrae
Summary of 5 types of genes (involved in EG) effect
Maternal-effect genes (specify A and P) Gap genes (divide A-P axis into broad subdomains) Pair-rule genes (divide A-P axis into 7 stripes - expression/non-expression) Segment polarity gene (define + organize the 14 segments, and define A and P halves of each segment) Homeotic genes (define structures produced by each segment)
Classify Bicoid, Nanos, Even-skipped, Engrailed and Ultrabithorax genes (in the 5 categories) and find the odd one out of the group
Bicoid: maternal-effect. Nanos: maternal-effect. Even-skipped: Pair-rule. Engrailed: Segment polarity. Ubx: Homeotic. Odd one = Nanos (bc not a TF)
One technique of developmental timing in C.elegans
Use of miRNAs
C.elegans total number of cells and pattern of cell divisions
959 (somatic) cells total. Cell divisions follow a stereotyped pattern (lineages). Number of muscle cells, gonad cells, neuron cells is well known.
What the screen for heterochronic mutants in C.elegans led to
Found that mutants in gene let-7 repeat the L4 division at the adult stage
Consequence of mutations that affect V1-4 vulval cell division pattern in C.elegans (2)
Gives rise to hypodermal cells and let-7 mutants die of vulval rupture
let-7 function
encodes short (22 nt) miRNAs derived from double stranded RNA precursors. Are incorporated in the RISC complex and bind complementary mRNA’s 3’ UTR
General consequence of mutations in genes like let-7
The target protein(s) is not repressed
Proteins targeted by let-7 and give one example and where it is targeted
let-7 targets several protein-coding mRNAs. (ex. lin-41)
Recognize two sequences in lin-41’s 3’ UTR
Mutation in lin-41 consequence
Too little Lin-41 protein -> L4 is not performed in the V1-4 vulval cell division pattern
Mutation in let-7 vs mutation in lin-41
let 7 mutations -> too much lin-41 -> L4 repeated
lin-41 mutation -> too little lin-41 -> L4 not performed
2 constructs used to show that let-7 targets lin-41 3’ UTR
- Insert GFP in lin-41 protein to make a fusion protein (lin-41 promoter, GFP, lin-41 sequence, lin-41 3’UTR)
- col-10 (an exogenous promoter), lacZ, lin-41 3’UTR
How they proved that let-7 targets lin-41 3’ UTR using the 2 constructs
Followed percentage of animals that expressed GFP and LacZ in hypodermial cells during L1 to L4 and adult stage and percentages of animals for both constructs at each phase were similar
Segments over which Abd-A, Abd-B and Ubx repress Dll
Abd-A : From A2 to A7
Abd-B: From A4 to A7
Ubx: From A1 to A7
