Salivary Gland Polytene Chromosomes
Rapid Embryogenesis- 10 minute cycles
Variations in the cell cycle during Drosophila development Canonical S G1 G2 No gaps No mitosis M Imaginal tissues (adult eyes, wings, etc.) S S Posters 350B 340A M G Early embryos Adult nurse cells Larval tissues
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 P PLU P P P P P P PNG GNU P P PNG- ser/thr protein kinase PLU- 19kDa ankyrin repeat protein, activating subunit GNU-27kDa protein; promotes dimerization and activation
PNG, PLU, GNU Translation [Cyclin B] / CDK1 DNA Replication Chromosome condensation Mitosis
Is PNG required to inhibit PUM repression of cycli lin B translation throughout the embryo? png mutant PLU PLU PNG GNU PNG GNU PUM ? No Translation Translation ? PUM 3 ’ UTR 3 ’ UTR cyclin B cyclin B Does removal of PUMILIO in a png mutant restore translation?
Mutations in pum umil ilio io suppress the png 331 331 8 mutant phenotype png 3318 pum/+ png 3318 pum png 3318 pum tubulin DNA
Transition from Meiosis to Embryonic Mitosis PNG Kinase PUM SMG Cyclin B Deadenylation mRNA deg. M Maternal- Zygotic Transition S Tadros et al. Dev. Cell 12, 143 (2007)
Meiosis in Female Drosophila Completion of meiosis as Primary Arrest Secondary Arrest egg passes Prophase I through Metaphase I oviduct Egg Oocyte Activation Maturation Adapted from Xiang et al. PLoS Biol 5(12): e323
Analysis of Protein Changes at Egg Activation • 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 Adapted from: Xiang Y et al., PLos Biol, 2007
Proteomics Results 3500 proteins scored in replicates 132 proteins upregulated at egg activation 230 proteins downregulated at egg activation (Zach Whitfield- Talk #10 today)
Polysome Profiling Results- 5300 mRNAs analyzed Upregulated Proteins- mRNAs move from monosome to polysomes Activated Egg Mature Oocyte (stage 14) 5’ 3’ 3’ 5’ Downregulated Proteins- mRNAs move off polysomes Activated Egg Mature Oocyte (stage 14) 3’ 3’ 5’ 5’
Role of PNG in Translation at Egg Activation Upregulated Proteins PNG Dependent Activated Egg Mature Oocyte (stage 14) 3’ 5’ 3’ 5’ Downregulated Proteins Largely PNG Independent Activated Egg Mature Oocyte (stage 14) 5’ 3’ 3’ 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.
PNG and PLU 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)
PhD Students: Postdoctoral Fellows: Deborah Burney-Sigman Myles Axton Masatoshi Hara Janet Carminati Sharon Bickel Iva Kronja (Poster 174C) Julie Claycomb Giovanni Bosco Jared Nordman Doug Fenger Astrid Clarke Belinda Pinto (Poster 325A) Jane Kim Hannah Cohen Jessica Von Stetina Janice Lee Kimberley Dej (Poster 350B) Wes Miyazaki Lisa Elfring Yingdee Unhavaithaya Daniel Moore Aki Hayashi Andrea Page-McCaw Cintia Hongay Eugenia Park Irena Ivanovska Technical Support Jillian Pesin Anne Kerrebrock Tamar Resnick Raissa Fomina Elaine Lahue Irena Royzman Heidi LeBlanc Helena Kashevsky Fay Shamanski Jacqueline Lopez Albert Smith Laura Lee Tracy Tang Cristina Nogueira Julie Wallace Bruce Reed Allyson Whittaker Noa Sher Fang Xie Leah Vardy Jessica Alexander Laura Frawley (Poster 340A) Brian Hua Zach Whitfield (Talk #10)
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