van den Ham R, van Dissel-Emiliani F M F, van Pelt A M M
Department of Biochemistry, Cell Biology and Histology, Faculty of Veterinay Medicine, University of Utrecht, Utrecht, The Netherlands.
Biol Reprod. 2003 Apr;68(4):1369-75. doi: 10.1095/biolreprod.102.004853. Epub 2002 Oct 31.
Previously, we found that the poly(A)+ RNA of the scaffolding subunit A (alpha isoform) of protein phosphatase 2A (PP2A-Aalpha) was clearly expressed by fetal gonocytes but weakly expressed by adult single (As), paired (Apr), and aligned (Aal) A spermatogonia. The scaffolding subunit A of PP2A (PP2A-A) is the major subunit in the formation of a functional PP2A holoenzyme. In this study, we investigated the expression of PP2A-A during testicular development in more detail using in situ hybridization, immunohistochemistry, and Western blot with testes of rats of various ages from 16 days postcoitum (pc) to adulthood. The expression of PP2A-A was detected in fetal proliferative gonocytes at 16 days pc, declining thereafter during the quiescent period of the gonocytes. From the day of birth to the start of spermatogenesis (Day 4 postpartum [pp]), the number of PP2A-A-immunopositive gonocytes increased. At Day 4 pp, the first A1 spermatogonia appeared along the basement membrane; all were PP2A-A positive. In the adult, PP2A-A was upregulated during the differentiation of the As, Apr, and Aal spermatogonia to the A1 spermatogonia and expressed thereafter by all other spermatogonia. Spermatocytes from the pachytene stage onward and all spermatids in the adult testis also showed clear expression of PP2A-A. In Sertoli cells, PP2A-A was detected during their proliferative period at 19 days pc to 15 days pp. The presence of a functional enzyme was confirmed by the additional detection of the catalytic subunit C of PP2A using Western blot analyses at various ages during testicular development. This apparent pattern of expression of PP2A-A during testicular development suggests that PP2A may play an important role in the proliferation of distinct populations of testicular cells and during meiosis and sperm maturation.
此前,我们发现蛋白磷酸酶2A(PP2A)的支架亚基A(α异构体,PP2A - Aα)的聚腺苷酸加尾RNA(poly(A)+ RNA)在胎儿生殖母细胞中明显表达,但在成年单倍体(As)、配对(Apr)和排列(Aal)A型精原细胞中表达较弱。PP2A的支架亚基A(PP2A - A)是功能性PP2A全酶形成中的主要亚基。在本研究中,我们使用原位杂交、免疫组织化学和蛋白质印迹法,对从受孕后16天(pc)到成年的不同年龄大鼠的睾丸进行研究,更详细地调查了PP2A - A在睾丸发育过程中的表达情况。在受孕后16天的胎儿增殖性生殖母细胞中检测到PP2A - A的表达,此后在生殖母细胞的静止期表达下降。从出生日到精子发生开始(产后第4天[pp]),PP2A - A免疫阳性生殖母细胞的数量增加。在产后第4天,首个A1型精原细胞沿基底膜出现;均为PP2A - A阳性。在成年期,PP2A - A在As、Apr和Aal型精原细胞向A1型精原细胞分化过程中上调,此后在所有其他精原细胞中表达。成年睾丸中从粗线期开始的精母细胞和所有精子细胞也显示出PP2A - A的清晰表达。在支持细胞中,在受孕后19天至产后15天的增殖期检测到PP2A - A。通过在睾丸发育的不同年龄使用蛋白质印迹分析额外检测PP2A的催化亚基C,证实了功能性酶的存在。PP2A - A在睾丸发育过程中的这种明显表达模式表明,PP2A可能在睾丸细胞不同群体的增殖以及减数分裂和精子成熟过程中发挥重要作用。