Functional analysis of the sV23 vitelline membrane protein in Drosophila melanogaster
The extracellular matrix (ECM) is a complex framework found outside of the cell that has been found not only to provide mechanical and structural support to tissues and organs, but also plays a pivotal role in housing molecules that influence cellular processes like migration, differentiation and proliferation. Understanding how the extracellular matrix is assembled provides insight into how this structure contributes to these processes and to the health of the organism. In this study, the eggshell of Drosophila melanogaster is used as an experimental system to study the assembly of a specialized extracellular matrix in vivo . The eggshell is a multilayered structure, consisting of an oocyte proximal vitelline membrane and surrounding chorionic structures, secreted during the late stages of oogenesis. sV23, a major vitelline membrane protein, is required for normal eggshell morphology and function, since sV23 protein null mutants are female sterile and exhibit disorganized chorionic structures. By comparing the behavior of normal and altered versions of the sV23 protein in the null mutant, it was shown that sV23 is secreted as a pro-protein from which C- and N-terminal sequences are sequentially removed. An array of engineered mutations was used to identify regions or amino acids that are important for fertility and the formation of an organized eggshell. Biochemical extraction and fractionation studies revealed sV23 becomes incorporated into a disulfide linked network that increases in size as development progresses. Site directed mutagenesis of targeted cysteine residues indicates that without cysteines, sV23 protein accumulation is negligible. At least one of the cysteines is essential for sV23 accumulation, but that more than one appears necessary for its function. Since sV23 mutants with a single cysteine residue cannot incorporate into large disulfide linked networks, this mutant provides a means to characterize initial disulfide mediated protein interactions.
Manogaran, Anita, "Functional analysis of the sV23 vitelline membrane protein in Drosophila melanogaster" (2003). Dissertations (1962 - 2010) Access via Proquest Digital Dissertations. AAI3093146.