The long term goal of this proposal is to characterize the genetic control of neural tissue development in Drosophila embryos. Two genes expressed early in all neural precursors (pan-neural genes) that encode likely transcription factors were identified by the enhancer-trap method.
Specific aims of the proposed study are focused on one of these genes, referred to as scratch (scrt), and address two parallel issues. The first is how the scrt gene influences the neural development as a single gene, and the second is how scrt interacts with other genes to define neurons as a tissue-type. The predicted scrt protein possesses five zinc fingers and shares significant sequence similarity with the product of the snail (sna) gene. The scrt and sna genes are also expressed in similar although not identical neuroectodermal patterns. To understand how scrt regulates embryonic neurogenesis we will first generate deletion and point mutant alleles of scrt and then analyze the embryonic phenotypes of these mutants. We will also mis-express scrt during neurogenesis in wild-type and mutant embryos using inducible promoters and P-transformation. To understand how scrt interacts with other genes, we will dissect scrt and sna pan-neural promoter elements using P-element lacZ-promoter fusion gene constructs and derived deletions to characterize the nature of upstream regulatory inputs. We will determine whether pan-neural promoter elements can be subdivided into independent subelements and whether scrt and sna share such elements. We will assay expression of endogenous genes and lacZ-promoter constructs in mutants to link particular promoter subelements with specific upstream regulatory genes. In parallel with these studies scrt mutations will be combined with sna point mutations and deletions and with other previously identified mutations affecting neurogenesis to address the possibility that scrt acts in concert with other genes. We will examine embryos and adult flies carrying an scrt mutation along with a test mutation for enhanced or suppressed phenotypes. We will extend these studies by conducting EMS screens for genes interacting with scrt (e.g modifiers of the scratched eye phenotype of putative scrt alleles). Finally, we will use low stringency hybridization and PCR amplification with scrt zinc finger probes and primers to search for structurally related pan-neural genes.
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