MITOCHONDRIAL CYTOCHROME-C OXIDASE-II MESSENGER-RIBONUCLEIC-ACID IS EXPRESSED IN PACHYTENE SPERMATOCYTES AT HIGH-LEVELS AND IN A STAGE-DEPENDENT MANNER DURING SPERMATOGENESIS IN THE RAT

Screening of a rat seminiferous tubule library (stages VI-VIII) resulted in isolation of several clones to the same abundant mRNA. Sequencing revealed that these encoded all or part of the second subunit of cytochrome C oxidase (COX II), the terminal enzyme in the electron transport chain located in mitochondria. The pattern of expression of the COX II mRNA in the testis was examined by Northern blot and in situ hybridisation combined with specific depletion of pachytene spermatocytes or Leydig cells by treatment with methoxyacetic acid (MAA) or ethane dimethane sulphonate (EDS), respectively. COX II mRNA was found to be strongly expressed in pachytene spermatocytes in adult and prepubertal rats, with the highest levels of expression at stages VII-VIII of the spermatogenic cycle. Although this is the androgen-dependent stage, Northern analysis of total testicular RNA showed that expression of COX II mRNA was not altered detectably by EDS-induced androgen withdrawal nor was expression altered detectably by 24-h treatment with high doses of FSH. Treatment of rats with MAA, which selectively depletes seminiferous tubules at most stages of pachytene spermatocytes, resulted in a marked reduction in COX II mRNA. In the absence of pachytene spermatocytes, COX II mRNA was visualized in preleptotene spermatocytes and in Sertoli cell cytoplasm. The expression of COX II in Sertoli cells was confirmed by examination in situ of a testis devoid of most germ cells and by Northern analysis of RNA from an adult Sertoli cell-enriched preparation. However, using either approach the amount of mRNA in Sertoli cells was considerably lower than that in pachytene spermatocytes. Cytochrome C oxidase has thirteen subunits, three of which (I-III) are encoded on the mitochondrial genome. The physiological significance of the high levels of expression of COX II mRNA in pachytene spermatocytes and its stage-dependent changes is unknown but they presumably reflect requirements for alterations in energy demand as these cells enter the final stages of meiosis. The stage-dependent differences in expression of COX II mRNA in pachytene spermatocytes may also explain differences in the susceptability of these cells to depletion by MAA.