A Genetic Screen for Modulators of the Notch Pathway in Drosophila Melanogaster Identifies Not1 as a Positive Regulator of Notch Signaling

Thesis advisor: Marc A.T. MuskavitchThe Notch pathway is an evolutionarily conserved mechanism of intercellular signaling that plays a central role in the development of metazoans. Here I summarize two genetic screens that utilize a rough eye phenotype created by Delta overexpression in the Drosophila eye to identify modulators of Notch pathway signaling activity. Among the many "hits" obtained from both screens, I have mapped to the Not1 gene a single complementation group that exhibits strong genetic interactions with Notch pathway mutants. NOT1 is a component of the CCR4-NOT complex, a global regulator of gene expression that exerts its effects through a variety of mechanisms, including mRNA deadenylation and direct transcriptional repression. I have conducted a series of genetic and molecular experiments in an effort to obtain more insight into the relationship between the CCR4-NOT complex and the Notch pathway. Both Not1 EMS mutations and RNAi-mediated knockdown of NOT1 expression produce phenotypes that mimic those of Notch loss-of-function pathway mutants. Knockdown of NOT1 in the developing bristle organ disrupts Notch-mediated inhibition of neuronal specification, resulting in supernumerary neurons and aberrant sheath cell specification. Knockdown of NOT1 within the developing wing margin disrupts expression of the Notch target genes Cut and Wingless, as well as the Notch ligand Delta. Phenotypic rescue experiments imply that Not1 functions downstream of Notch signal activation and acts directly on Notch target gene expression. These results suggest that NOT1 is required for Notch signal transmission in certain developmental contexts and implicate the CCR4-NOT complex as a positive regulator of the Notch pathway.Thesis (PhD) — Boston College, 2009.Submitted to: Boston College. Graduate School of Arts and Sciences.Discipline: Biology