Title: Announcements
1Announcements
- Exam this Wednesday my half is 40. Gerry
Prodys half is 60. - Exam regrade policy if you have a question about
how I graded an answer, send me an email
outlining exactly how you felt I went wrong, and
submit the exam for regrade. - You MUST (1) make this request in writing, and
you MUST (2) submit the exam, both within 5
business days of its return. - PLEASE NOTE that I will regrade the entire exam
your grade may go up or it may go down (more
likely).
2Biol/Chem 473
Schulze lecture 4 Eukaryotic gene regulation
Early Drosophila development
3Eukaryotic transcription is complex
4Complexity of eukaryotic transcription
5Insulators may function to organize the genome
into transcriptionally autonomous domains
6Linking complex gene regulation to nuclear
organization
7(No Transcript)
8The best model organism in the whole world!!!!
Drosophila melanogaster black bellied dew
lover fruit fly
9The early Drosophila embryo is a syncytium (a bag
of nuclei)
10(No Transcript)
11(Maternal effect genes)
12(No Transcript)
13Patterning genes classified by mutant effect on
segmentation
The hierarchical nature of these interactions can
be shown by genetics mutations in earlier levels
of the hierarchy severely disrupt later
patterning.
14Seminal publication and subsequent Nobel
15Maternal genes
- Required before the embryo starts transcribing
its own genes - Mother expresses these genes during oogenesis
(mum packs a lunch) - Localization of maternal gene products is the
first step in the cascade
http//flymove.uni-muenster.de/Processes/Segmentat
ion/
16Earliest stages of development run on mRNAs and
proteins deposited by mum during oogenesis
- An efficient way to localize and concentrate
proteins is by localization of mRNA in a
translationally repressed state, followed by
activation of translation when the mRNA reaches
its destination. - How are mRNAs localized?
- How are mRNAs repressed?
- How are localized mRNAs then, when required,
de-repressed?
17Maternal effect Mum packs a lunch for the embryo
Anterior end
Posterior end
18Maternal effect Mum packs a lunch for the embryo
- The bicoid gene encodes a homeodomain
transcription factor. - The oskar gene encodes something we are not quite
sure about. - The gurken gene encodes TGFß homolog - EGF-R
ligand, required for dorso / ventral patterning
of the egg and embryo. - The nanos gene encodes a translational repressor
- zinc finger that targets Hunchback- and
Bicoid-mRNAs to achieve posterior identity.
19Maternal effect Mum packs a lunch for the embryo
- Polarity of the oocyte cytoskeleton is essential
for maternal factor localization (gurken plays a
role here). - Bicoid mRNA gets localized to anterior pole
because another protein, called Staufen,
recognizes a structure in the 3UTR of the bicoid
message (and Staufen protein is itself localized
by the polarity of the cytoskeleton.
20Maternal effect Mum packs a lunch for the embryo
- Bicoid mRNA gets localized to anterior pole
because another protein, called Staufen,
recognizes a structure in the 3UTR of the bicoid
message (and Staufen protein is itself localized
by the polarity of the cytoskeleton.
21Maternal effect Mum packs a lunch for the embryo
These dsRNA UTR sequences are required for
anterior localization of bicoid mRNA by Staufen
protein
- Bicoid mRNA gets localized to anterior pole
because another protein, called Staufen,
recognizes a structure in the 3UTR of the bicoid
message (and Staufen protein is itself localized
by the polarity of the cytoskeleton).
22Gap genes
- The gap genes are expressed by the zygote
(activated in the syncytial blastoderm) - Their expression patterns are regulated by the
maternal gene products, and they regulate each
other - They divide the embryo into broad, unique domains
containing different combinations of
transcription factors
http//flymove.uni-muenster.de/Processes/Segmentat
ion/
23Pair-rule genes
- Pair-rule genes are expressed just before
cellularization in patterns of 7 stripes (half
the number of segments in a wild type larva) - Some pair-rule genes respond to patterns already
laid down by the maternal and gap genes, while
others are regulated by other pair-rule genes - Pair rule gene expression is very dynamic!
(pretty pictures)
http//flymove.uni-muenster.de/Processes/Segmentat
ion/
24Segment polarity genes
- Segment polarity genes are expressed at the onset
of gastrulation - They are expressed in 14 segment bands, refining
the patterns of all the previous genes in the
hierarchy. Maternal, gap and pair-rule gene
expression begins to fade away
http//flymove.uni-muenster.de/Processes/Segmentat
ion/
25Stripe boundaries are precisely defined
26Control regions upstream of pair-rule genes are
complex
27ASSIGNED PAPERTranscriptional regulation of a
pair-rule stripe in Drosophila
Small, S. et al., Genes Development (1991) 5
827-839
28Main players
- Even-skipped (eve) is a homeo-box containing
transcription factor. - Hunchback (hb) has multiple Zn fingers.
- Bicoid (bic) is a homeo-box containing
transcription factor. - Kruppel (Kr) contains a Zn finger.
- Giant (gt) contains a Leucine zipper.
29Wild type expression patterns of putative
regulators of eve stripe 2
- even-skipped protein is red
- hunchback protein is green
- overlap of expression is yellow
30Wild type expression patterns of putative
regulators of eve stripe 2
- even-skipped protein is red
- Kruppel protein is green
- overlap of expression is yellow
31Wild type expression patterns of putative
regulators of eve stripe 2
- even-skipped protein is green
- giant protein is red
- Overlap of expression is yellow
32Wild type expression patterns of putative
regulators of eve stripe 2
High levels of gt
High levels of Kr
33Wild type expression patterns of putative
regulators of eve stripe 2
High levels of gt
High levels of Kr
What predictions arise from this picture?
34eve-lacZ gene fusion
This is a transgene gets injected into the
embryo (P transformed embryos), integrates into
the genome and is expressed stably along with all
the other genes. BUT this transgene will express
lacZ in an even-skipped-stripe 2 dependent manner!
35Questions to ask about in vivo expression patterns
- How do we know the order in which patterning
genes act in development? - How were potential trans-acting regulators of
stripe 2 identified? - What would happen to the domain of expression of
eve stripe 2 in giant or Kruppel mutant embryos? - Do these vivo expression patterns indicate direct
interaction between Kr and gt and the eve stripe
2 regulatory region?