| This thesis examines how P elements silence genes in Drosophila melanogaster using the host's heterochromatin proteins. This has been modeled using the transgene P{lacW}ciDPlac, a white+ (w+) P element construct inserted upstream of the ci gene on chromosome 4. In the absence of other P elements, this transgene produces a uniform red eye phenotype [white+ (w)] in w- flies, but the presence of some P elements such as P[SalI] and KP changes the phenotype to a variegated eye color due to random silencing of the w+ transgene expression in different ommatidial cells in the eye. This phenomenon, called P element dependent silencing, is dependent on host heterochromatin proteins such as SU(VAR)2-5 and SU(VAR)3-7. These heterochromatin proteins are dose dependent in their effects. As part of this study, I showed that by increasing the number of the KP elements in the genome the silencing of the transgene gets stronger. This means that the effects of P elements on PDS are also dose dependent.;I also examined the P-Sal mutants for their effect on other P element constructs that are sensitive to the presence of P-Sal. I found that the P{lacW}ciE1, P{hsp26-pt-T}ci2-m1021.R, and P{hsp26-pt-T}39C-12 constructs, which are inserted upstream of the ci, are similar in their response to my P-Sal mutants, while P{hsp26-pt-T}39C-12, which is located more distally on chromosome 4, is different both in phenotype and sensitivity.;The phenotypic differences in my P-Sal mutations could have been due to differences in polypeptide expression. I tried to induce antibodies against the P-Sal protein. Preliminary characterization of this antibody showed it was not specific enough to identify the protein expression (or lack of expression) in the fly protein extract.;To examine the role of the P element product in gene silencing, I induced and recovered 22 mutations in P[SalI]89D (P-Sal hereafter) that affected the process of PDS. These mutations were found in regions previously reported to be required for transposition. However, there were no mutations in the zinc finger domain, which is critical for DNA binding of P element proteins. Moreover, I found two new mutation hot spots in the P element DNA sequence. These sequences do not encode any known protein domains. |
