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Abstract

Rat testicular extracellular superoxide dismutase (SODEX): Its purification, characterization, and regulation (PDF) (HTML) 
Mruk,Dolores D.; Cheng,Chin Ho; Cheng,Yan Ho; Mo,Meng-Yun; Grima,Josephine; Silvestrini,Bruno; Lee,William M.; Cheng,Chuen-yan
Biology of Reproduction 59 298-308
Publication date: 1998

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Using multiple HPLC steps, we have identified and purified a68-kDa polypeptide (as estimated by gel permeation HPLC) toapparent homogeneity, from primary Sertoli cell-enriched culturemedium, that consisted of two monomers of 35 ( chain) and 33kDa (ß chain) on SDS-polyacrylamide gel running underreducing conditions. Partial N-terminal amino acid sequenceanalysis of these two monomers revealed sequences of NH-DXGESGVDLADRL(SOD-) and NH-XXDTGESGVDLADXL (SOD-ß), which areidentical to rat extracellular superoxide dismutase (SOD)with the exceptions that SOD- and SOD-ß are missing,respectively, four (Trp-Thr-Met-Ser) and two (Trp-Thr) aminoacids from their N-termini, compared to rat SOD, suggestingthat the cleavage sites of the SOD gene in the testis aredifferent from that of other organs. Studies by sequential useof reverse transcription and polymerase chain reaction (PCR)using two SOD primers have demonstrated the expression ofSOD in the heart, brain, lung, kidney, epididymis, testis,Sertoli, and germ cells, with low expression in the liver andovary and no expression in the uterus, spleen, or thymus. Nucleotidesequence analysis of this 447-base pair PCR product from Sertolicells revealed that its sequence is equivalent to the sequenceof previously published rat SOD. During testicular maturation,the SOD steady-state mRNA level increased significantly from20 to 60 days of age and then declined at 90 days of age. Suchan increase in the testicular SOD expression during maturationis not likely a result of an up-regulation by germ cells, sincegerm cells isolated from either 20- or 60-day-old rats whencocultured with Sertoli cells failed to elicit an increase inSOD expression in the cocultures. Using primary Sertoli cellcultures in vitro, it was found that Sertoli cell SOD expressionwas stimulated by interleukin-1 but not by either interferon-or basic fibroblast growth factor. These results illustratethat Sertoli cells as well as germ cells synthesize and/or secretea testicular variant of SOD that may provide essential cluesto understanding superoxide radical-mediated damage in the gonad.

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