salivary gland polytene chromosomes
TRANSCRIPT
Salivary Gland Polytene Chromosomes
Rapid Embryogenesis- 10 minute cycles
Variations in the cell cycle during Drosophila development
Canonical S
M
G2 G1
Imaginal tissues (adult eyes, wings, etc.)
Adult nurse cells Larval tissues
S
G
No mitosis
S
M
No gaps
Early embryos
Posters 350B 340A
..
Unregulated DNA replication and Absence of mitosis in png, plu, or gnu mutants
wild type png, plu, or gnu mutant
early
late
The PNG kinase is inactive!
PNG, PLU, and GNU form an active protein kinase complex
PNG GNU
P
P
P
P
P
PLU P P P
P
PNG- ser/thr protein kinase PLU- 19kDa ankyrin repeat protein, activating subunit GNU-27kDa protein; promotes dimerization and activation
PNG, PLU, GNU
[Cyclin B] / CDK1
DNA Replication Chromosome condensation Mitosis
Translation
Is PNG required to inhibit PUM repression of cyclin B translation throughout the embryo?
PNG GNU PLU
png mutant
No Translation
cyclin B 3’UTR
? PUM
? PUM
PNG GNU PLU
cyclin B 3’UTR
Translation
Does removal of PUMILIO in a png mutant restore translation?
Mutations in pumilio suppress the png331 8 mutant phenotype
png3318 pum/+
png3318 pum
png3318 pum
tubulin DNA
M Maternal- Zygotic Transition
PNG Kinase
S
PUM
Cyclin B
SMG
Deadenylation mRNA deg.
Tadros et al. Dev. Cell 12, 143 (2007)
Transition from Meiosis to Embryonic Mitosis
Primary Arrest Prophase I
Secondary Arrest Metaphase I
Completion of meiosis as egg passes through oviduct
Adapted from Xiang et al. PLoS Biol 5(12): e323
Oocyte Maturation
Egg Activation
Meiosis in Female Drosophila
Analysis of Protein Changes at Egg Activation
Adapted from: Xiang Y et al., PLos Biol, 2007
• Quantitative proteomic analysis of proteins that increase or decrease at activation
•Polysome profiling-identify all mRNAs recruited or released from polysomes
Iva Kronja Poster 174C
3500 proteins scored in replicates 132 proteins upregulated at egg activation 230 proteins downregulated at egg activation (Zach Whitfield- Talk #10 today)
Proteomics Results
Mature Oocyte (stage 14) Activated Egg
Downregulated Proteins-mRNAs move off polysomes
Polysome Profiling Results- 5300 mRNAs analyzed
Upregulated Proteins-mRNAs move from monosome to polysomes
Mature Oocyte (stage 14) Activated Egg
5’ 5’
5’ 5’
3’ 3’
3’ 3’
Mature Oocyte (stage 14) Activated Egg
Downregulated Proteins
Role of PNG in Translation at Egg Activation
Upregulated Proteins
Mature Oocyte (stage 14) Activated Egg
PNG Dependent
Largely PNG Independent
3’ 3’
3’ 3’ 5’ 5’
5’ 5’
PNG serves to activate translation generally at the developmental transition from oocyte to embryo
• Pursue biological problems in unbiased way.
• Generosity of the community in providing resources. • Application of varied approaches including high throughput. • Drosophila is an amazing model organism with many more exciting problems to solve.
Fay Shamanski Myles Axton Debbie Burney-Sigman Janet Carminati Lisa Elfring Doug Fenger Helena Kashevsky Laurie Lee Leah Vardy Masatoshi Hara Iva Kronja (Poster 174C)
PNG and PLU
Masatoshi Hara Iva Kronja (Poster 174C) Jared Nordman Belinda Pinto (Poster 325A) Jessica Von Stetina (Poster 350B) Yingdee Unhavaithaya
PhD Students: Deborah Burney-Sigman Janet Carminati Julie Claycomb Doug Fenger Jane Kim Janice Lee Wes Miyazaki Daniel Moore Andrea Page-McCaw Eugenia Park Jillian Pesin Tamar Resnick Irena Royzman Fay Shamanski Albert Smith Tracy Tang Julie Wallace Allyson Whittaker Fang Xie Jessica Alexander Laura Frawley (Poster 340A) Brian Hua Zach Whitfield (Talk #10)
Postdoctoral Fellows: Myles Axton Sharon Bickel Giovanni Bosco Astrid Clarke Hannah Cohen Kimberley Dej Lisa Elfring Aki Hayashi Cintia Hongay Irena Ivanovska Anne Kerrebrock Elaine Lahue Heidi LeBlanc Jacqueline Lopez Laura Lee Cristina Nogueira Bruce Reed Noa Sher Leah Vardy
Technical Support Raissa Fomina Helena Kashevsky