Salivary Gland Polytene Chromosomes

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