Showing papers by "Himangshu S. Bose published in 2014"

STARD6 is expressed in steroidogenic cells of the ovary and can enhance de novo steroidogenesis.

Abstract: STARD6 is a member of the StAR-related lipid transfer (START) domain family of proteins whose function thus far remains obscure. While it recently was shown to facilitate steroidogenesis in a cell-free setting, it has not been localized to steroidogenic cells of normal reproductive tissues. In a recent microarray study, we detected STARD6 mRNA in cultured porcine ovarian granulosa cells which are steroidogenic. In the present study, we examined regulation of STARD6 mRNA in porcine granulosa cultures, and found that it was not regulated by cyclic AMP, but it was reduced by combined knockdown of the transcription factors GATA4 and GATA6. We detected both STARD6 mRNA and protein in fresh granulosa cells and whole antral follicles and different stage corpora lutea of pig. The highest levels were discovered in the mid-luteal phase corpus luteum. Immunolocalization within ovarian tissues indicated robust STARD6 immunoreactivity in steroidogenic cells of the corpus luteum. Relatively lesser amounts of STARD6 signal were found in granulosa cells, theca cells, and oocytes. To test the ability of STARD6 to facilitate de novo steroidogenesis, non-steroidogenic COS-1 cells were co-transfected with components of the P450 cholesterol side-chain cleavage system, enabling them to make pregnenolone, and STARD6. STARD6 increased pregnenolone production by two- to three-fold over the empty vector control. In summary, STARD6 is found in the pig ovary, exhibits the strongest expression in highly steroidogenic luteal cells, and significantly enhances pregnenolone production in transfected COS cells independent of cyclic AMP treatment. Collectively, these findings indicate that STARD6 may contribute to steroidogenesis in ovarian cells, but also suggests other cellular functions that require cholesterol trafficking.

Passenger protein determines translocation versus retention in the endoplasmic reticulum for aromatase expression

Abstract: Aromatase protein is overexpressed in the breasts of women affected with cancer. In the endoplasmic reticulum (ER), signal sequence and signal anchors (SAs) facilitate translocation and topology of proteins. To understand the function of type-I SAs (SA-Is), we evaluated translocation of aromatase, whose signal anchor follows a hydrophilic region. Aromatase SA-I mediates translocation of a short N-terminal hydrophillic domain to ER lumen and integrates the protein in the membrane, with the remainder of the protein residing in the cytosol. We showed that lack of a signal peptidase cleavage site is not responsible for the stop-transfer function of SA-I. However, SA-I could not block the translocation of a full-length microsomal secretory protein and was cleaved as part of the signal sequence. We propose that interaction between the translocon and the region after the signal anchor plays a critical role in directing the topology of the protein by SA-Is. The positive charges in the signal sequence helped it to override the function of signal anchor. Thus, when signal sequence follows SA-I immediately, the interaction with the translocon is perturbed and topology of the protein in ER is altered. If signal sequence is placed far enough from SA-I, then it does not affect membrane integration of SA-I. In summary, we conclude that it is not just the SA-I, but also the region following it, which together affect function of aromatase SA-I in ER.