An ovarian follicular epithelium protein of the silkworm (Bombyx mori) that associates with the vitelline membrane and contributes to the structural integrity of the follicle

Abstract

We have cloned and functionally characterized a novel protein, BmVMP30, which is synthesized by the cells of the follicular epithelium of the ovarian follicles of the domesticated silkworm Bombyx mori, secreted from them and associated with the vitelline membrane. BmVMP30 is a 30 kDa protein that bears limited structural features reminiscent of other insect vitelline membrane proteins. Although BmVMP30 does not share pronounced similarities or signature motifs with other reported proteins, its temporal and spatial expression and its behavior throughout oogenesis suggest that it is a novel member of the insect vitelline membrane protein family. The protein is expressed exclusively in the cells of the follicular epithelium during stages 3 315 to 3 31 of vitellogenesis, secreted from them and, ultimately, localized at the junction between the oocyte and the eggshell, where the vitelline membrane is located. Treatment of follicles with an antisense oligonucleotide that encompasses the translation initiation codon results in the production of an N-terminally truncated protein and disruption of the integrity of the follicular epithelium. Antisense oligonucleotide treatment, however, has no e¡ect on the implementation of the developmental program that directs the autonomous progression of ovarian follicles through the last stages of vitellogenesis and choriogenesis. ß

Vitellogenesis and Post-Vitellogenic Maturation of the Insect Ovarian Follicle

“…However, after fertilization no BmVMP30 protein could be detected by Western blot analysis in the eggshell fraction of the developing embryos. Thus, the temporal and spatial expression pattern of BmVMP30 during and after oogenesis is consistent with the behavior expected for a VMP (Kendirgi et al, 2002). Although BmVMP30 does not have significant sequence identity with any Drosophila or mosquito VMPs, including the VM domain, it does share with them a distinct overall proline distribution, a similar general hydropathy profile and the lack of introns in the gene structure (Kendirgi et al, 2002).…”

“…During the vitellogenic stages that precede the onset of choriogenesis, other important events occur in follicles, including the synthesis of vitelline membrane proteins and ovarian (maternal) ecdysteroids by the cells of the follicular epithelium (Kadono-Okuda et al, 1994; Kendirgi et al, 2002), the reinitiation of meiosis in the oocytes (Lanot et al, 1990), and morphogenetic movements of the follicular epithelium prior to their commitment to chorion protein gene expression (Spradling, 1993;Montell, 2001). …”

“…Bombyx mori A cDNA clone, BmVMP30, encoding a putative VMP has been isolated from a screen for genes that are differentially expressed with respect to stages À35/À1 (Kendirgi et al, 2002). BmVMP30 is a 30 kDa protein that is synthesized and secreted by the cells of the follicular epithelium during late vitellogenesis (stages À18 to À1).…”

“…Strong knockdowns of BmVMP30 gene expression, achieved by antisense oligonucleotide technology, resulted in the appearance of a severe phenotype suggesting disruption of the integrity of the follicular epithelium, while no interference was observed with the initial steps of the program that controls the activation of chorion protein genes ( Figure 16; Kendirgi et al, 2002). Because the knockdown experiments were carried out in in vitro culture, the long-term effects on follicle development and eggs (e.g., completion of choriogenesis) could not be assessed.…”

Architectural factor HMGA induces promoter bending and recruits C/EBP and GATA during silkmoth chorion gene regulation

“…Antisense DNA assays were performed essentially as described in [26]. Briefly, a 4-day-old pupa was dissected and follicles corresponding to vitellogenic (-10 to -6, Series A) and to early choriogenic stages (+1 to +5, Series B) were collected and allowed to develop ex vivo in appropriate medium (Grace's Insect Medium, 0.5% fetal calf serum, 50 mgr/ml gentamycin) and environment (28 o C, 100% oxygen atmosphere).…”