Eggshell Assembly inDrosophila:Processing and Localization of Vitelline Membrane and Chorion Proteins

AbstractTheDrosophilaeggshell consists of three major proteinaceous layers: the vitelline membrane, the inner chorionic layer, and the outer endochorion. During the latter stages of oogenesis, the proteins that comprise these layers are synthesized and secreted by epithelial follicle cells which surround the maturing oocyte. While there is considerable knowledge of the structural units which comprise the eggshell layers, there is little knowledge of how individual proteins function or interact with one another to form the structure. Immunoelectron microscopy was used to follow the distribution of four different eggshell proteins in the assembling and mature eggshell. sV23 and sV17, follicle cell proteins synthesized during the early stages of eggshell formation (stages 8–10), were distributed within the vitelline membrane layer at all stages. Despite marked temporal differences in their accumulation profiles, s36 and s18, putative chorion proteins, were similarly distributed throughout the floor, pillars, and roof of the endochorion. Although the vitelline membrane appears to be morphologically complete by stage 11, developmental Western blots and immunolocalization data indicate that molecular dynamism persists within the layer throughout the subsequent choriogenic stages. During early chorion formation the vitelline membrane appears to act as a reservoir for chorion proteins since s36 was found predominantly in the vitelline membrane layer of stage 12 egg chambers. During the late choriogenic stages (13–14), both sV17 and sV23 are processed to smaller derivatives. Interactions between the eggshell layers were suggested by ultrastructural analysis of a sV23 protein null mutant which showed that the structural integrity of the outer chorion is dependent upon the presence of a vitelline membrane component

Procedure for the permeabilization and cryobiological preservation of Drosophila embryos

The authors describe the detailed protocol developed in their laboratory at Oak Ridge for the permeabilization and cryobiological preservation of embryos of Drosophila melanogaster, Oregon R strain. The protocol is supplemented by notes containing two sorts of information. One category includes references to the appropriate portions of their published papers giving the scientific rationale and experimental basis for important steps. The other category is concerned with the criticality of certain steps and the precision with which they need to be performed. As an aid to investigators, the authors list even ordinary pieces of equipment. Brand names and model numbers are given where it is either important or convenient for readers to know precisely what is used