A MOLECULAR ANALYSIS OF VITELLINE MEMBRANE PRODUCTION IN DROSOPHILA MELANOGASTER

As a first step towards understanding vitelline membrane (VM) formation, these studies have been directed towards the identification of the VM proteins and their encoding genes. SDS-PAGE and 2-D electrophoresis were used to identify 6 radiolabeled components ranging in molecular weights from 14K to 130K extracted from purified VM preparations. Analysis of in vivo labeled proteins from egg chambers of different developmental stages and stage 10 follicle cells showed that 5 of these VM components are synthesized by the follicle cells during the period of VM deposition. In addition, immunological analysis utilizing serum containing anti-VM activity has confirmed the identity of 3 of these components. Electrophoretic analysis of newly synthesized poly A(\u27+) RNAs radiolabeled in vivo from egg chambers of different developmental egg chambers and stage 10 follicle cells revealed 2 highly labeled RNA species that were specifically synthesized in stage 10 follicle cells. The size of these two abundant RNA species, designated as T(,1) and T(,2) was in the range expected for mRNAs encoding the smaller VM proteins. In vitro translations of the T(,1) and T(,2) RNAs in a reticulocyte translation system in the presence and absence of pancreas microsomal membranes revealed protein products that are initially synthesized in precursor forms and subsequently processed to species co-migrating with VM proteins. The T(,2) RNA was found to direct the synthesis of two species co-migrating with the 23-24 and 17.5K-dalton VM proteins while the T(,1) RNA resulted in the translation of a protein co-migrating with the 14K-dalton VM protein. The cytogenetic localization of gene sites encoding the T(,1) and T(,2) RNAs was determined by in situ hybridization of radiolabeled T(,1) and T(,2) RNAs with Drosophila polytene chromosomes. The T(,2) RNA hybridized to locus 42A of the right arm of the second chromosome while the T(,1) RNA displayed hybridization at 39D-E of the left arm of the same chromosome