Does Rho GAP 71E affect cell polarity in Drosophila neuroblasts? Megan Gladwin Colorado State University of Pueblo Prehoda Lab Mike Drummond SPUR 2010 University of Oregon

Neuroblasts are Stem Cells that Give Rise to Neurons Begin to form early in development Neuroblasts delaminate from neuroepithelium Inherit polarity – Apical – Basal Prehoda (2009)

Neuroblasts Must Go through Asymmetric Cell Division Prehoda (2009)

Proteins Determine Neuroblast Polarity Apical – Atypical protein kinase C Basal – Miranda Adapter protein for basal determinants Required for differentiation aPKC Mir

aPKC is a Key Regulator in Neuroblast Polarity Sets stage for other determinants aPKC mutants show delocalized basal factors Prehoda (2009) Miranda aPKC wild type Rolls et al. (2004)

Cdc42 is Necessary for Neuroblast Polarity Wild Type Cdc42 Mutant Atwood et al. (2007)

Cdc42 Cycles Between Inactive and Active Forms Cdc42-GDPCdc42-GTP GAP GEF

What GAP(s) are Necessary for Establishing Proper Neuroblast Polarity? Prehoda (2009)

Question: Does Rho GAP 71E affect cell polarity during asymmetric cell division? Atwood et al. (2009)

Experiment Third instar larvae with 71E RNAi Mutant 71E embyros Stained with antibodies – aPKC, Mir, PH3 and Dpn Confocal microscopy – Look at neuroblasts during metaphase

Wild Type vs. Rho GAP 71E aPKC/ PH3MirMerge WT Larval Rho GAP 71E RNAi WT Embryonic Rho GAP 71E Mutant

aPKC/PH3MirMerge Quantifying Images 71E RNAi neuroblast

aPKC/PH3MirMerge Quantifying Images 71E RNAi neuroblast

Basal over Apical Peaks will give a Ratio to Compare If more aPKC is present on basal domain then ratio will be greater A higher ratio would suggest that aPKC is no longer restricted to the apical domain ApicalBasal

Results Larval Brain NeuroblastsEmbryonic Neuroblasts

Conclusions & Future Directions Rho GAP 71E has an affect on cell polarity Continue to look at neuroblasts with Mir Count neuroblasts within Rho GAP 71E RNAi

Acknowledgements Ken Prehoda Mike Drummond Prehoda Lab – Marisa – Doug – Ryan – Chiharu – Nick – Jon – Michelle – Chris – Ruth Siboni – Amanda Baker – Evan Dewey Peter O’Day SPUR interns NSF REU

Pathway

GAL4