Coregulation of a Neighboring EST with RHOGAP

• Main Author: Hughes, Tia
• Format: Others
• Published: TopSCHOLAR® 2008
• Subjects: Biology
• Online Access: http://digitalcommons.wku.edu/theses/367; http://digitalcommons.wku.edu/cgi/viewcontent.cgi?article=1370&context=theses

It has been reported that the crossveinless-c locus of Drosophila melanogaster corresponds the RhoGAP88C gene. This determination was based on three criteria: proximity in genetic map position, complementation tests, and similarity of phenotype between RhoGAP88C RNAi constructs and the phenotype of hypomorphic cv-c' mutant homozygotes, both of which eliminate posterior crossveins (Denholm et al. 2005). There are several genes that map to this region of chromosome 3, including both RhoGAP88C, and a poorly annotated PDZ-domain containing gene corresponding to Expressed Sequence Tag (EST) Clot 13975 (approximately 60 kb upstream from the RhoGAP88C transcriptional start). A P-element insertion P[PZ]l(3)06951 is located immediately adjacent to an EST Clot 13975, but the homozygous lethal phenotype of the insertion appears to be unrelated to this EST. The data presented by Denholm et al. (2005) showing that 1(3)06951 fails to complement cv-c1 is relatively weak; only about 35% of 1(3)06951/cv-c' heterozygotes have a broken crossvein phenotype. This is in spite of the fact that 1(3)06951 is homozygous lethal, and cv-c1 homozygotes have a completely penetrant broken/missing crossvein phenotype. I have mobilized 1(3)06951 and recovered lethal insertion mutants 500 bp away from the original insertion that complement 1(3)06951 and fail to complement cv-c1 with a completely penetrant posterior crossveinless phenotype. I hypothesize based, on our data and that of Denholm et al. ( 2005), that P[PZ]l(3)06951 and RhoGAP88C are allelic with one another and distinct from the cv-c locus. I have used Real-Time PCR to show that mutations that fail to complement cv-c and mutations that fail to complement RhoGAP88C result in lower levels of both RhoGAP and Clot 13975 transcripts. This suggests that the mutations in the region may be affecting transcription levels of multiple mRNAs and may be causing some of the difficulty in assigning mutations to specific genetic loci.