Identification of a novel retrovirus expressed in rat Sertoli cells and granulosa cells.
Differential display reverse transcriptase-polymerase chain reaction (DDRT-PCR) was used to identify a novel retrovirus, designated SC1, that is expressed at high levels in rat granulosa cells and prepubertal Sertoli cells. The initial DDRT-PCR screen was performed using RNA from cultured prepubertal rat Sertoli cell, liver, and brain samples. SC1 was detected in the prepubertal rat Sertoli cell samples but not in those from liver and brain. SC1 cDNA was 6 kilobases in length and contained regions encoding for the gag, pol, and env retroviral proteins. Northern blot analysis failed to detect expression of the SC1 gene in total RNA isolated from adult brain, heart, spleen, lung, liver, skeletal muscle, kidney, prostate, and epididymis. Similarly, Northern blot analysis of testes from rats at various ages of development showed that high-level expression of the SC1 gene was limited to prepubertal testis samples. In situ hybridization analysis localized the SC1 mRNA to the seminiferous tubules of prepubertal testes and at a much lower level in Sertoli cells of adult testes. Northern blot analysis of total RNA isolated from Sertoli cells from 20-, 27-, and 35-day-old rat Sertoli cells and type A spermatogonia, pachytene spermatocytes, and round spermatids showed expression of the SC1 gene to be restricted to 20- and 27-day-old Sertoli cells, with no expression detected in germ cells. Furthermore, Northern blot analysis also showed expression of the SC1 gene in rat ovaries, and the level of expression was affected during eCG/hCG-induced ovulation. Expression of SC1 mRNA was localized by in situ hybridization of eCG-treated ovaries to the granulosa cell layer in developing follicles. Southern blot analysis showed SC1 to be endogenous in the rat and absent in mouse and human cell genomes. Transient transfection assays using the SC1 promoter region showed high promoter activity in MSC-1 and cultured prepubertal rat Sertoli cells, and no activity in 3T3 or MCF-7 cell lines.
Anway
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MD
Center for Reproductive Biology, School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4660, USA.
Johnston
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Crawford
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Griswold
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MD
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002478:Cells, Cultured; D016922:Cellular Senescence; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D005260:Female; D006107:Granulosa Cells / Q000821:virology*; D017403:In Situ Hybridization; D008297:Male; D008969:Molecular Sequence Data; D010053:Ovary / Q000821:virology; D010062:Ovulation Induction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D012367:RNA, Viral / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D012190:Retroviridae / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012708:Sertoli Cells / Q000821:virology*; D012741:Sexual Maturation; D014162:Transfection
D006063:Chorionic Gonadotropin; D012333:RNA, Messenger; D012367:RNA, Viral