Loveland K L, Hayes T M, Meinhardt A, Zlatic K S, Parvinen M, de Kretser D M, McFarlane J R
Institute of Reproduction and Development, Monash University, Clayton, Victoria, Australia.
Biol Reprod. 1996 Apr;54(4):896-904. doi: 10.1095/biolreprod54.4.896.
The testis is one of the most abundant sources of microtubule networks. These networks include mitotic and meiotic spindles, the spermatid manchette and axoneme, and the Sertoli cell cytoskeleton. Microtubules are composed of alpha- and beta-tubulin subunits that are polymerized and stabilized by a variety of microtubule-associated proteins (MAPs). One of these, MAP2, has been extensively characterized as a brain-specific protein with the capacity to bind tubulin, cAMP-dependent kinase, and calmodulin. MAP2 mRNA is processed into at least two variants encoding proteins designated MAP2a, MAP2b, and MAP2c. Of the 5.7 kb of coding sequence in the 9-kb mRNA that encodes MAP2a and MAP2b, a deletion of approximately 4 kb produces mRNA encoding MAP2c, which consists of only the N- and C- terminal regions of MAP2b. To determine whether MAP2 was present in the rat testis, microtubule preparations were isolated from adult rat testis and brain by means of taxol-mediated polymerization and analyzed by gel filtration, ELISA, and Western blotting using polyclonal and monoclonal antibodies reactive with MAP2. A 74-kDa protein corresponding to MAP2c was detected in the testis. These results were confirmed by Northern blot analysis of total RNA from adult rat brain and testis with cDNA probes that distinguish between the known MAP2 splice variants. The predominant mRNAs in testis of 6 kb and 2.5-3.5 kb corresponded to MAP2c. A single 6-kb mRNA with the potential to encode MAP2c was detected in enriched preparations of immature Sertoli cells and adult Leydig cells. Round spermatids contained at least two MAP2 mRNAs between approximately 2.5 and 3.5 kb in size that displayed a stage-specific pattern of expression. Immunohistochemistry showed a MAP2-like protein in both somatic and germ cells, with a particularly distinct localization within the cytoplasm of primary and secondary spermatocytes at stage XIV of the seminiferous cycle during meiotic metaphase. In addition to cytoplasmic staining, a novel localization of this protein was observed in the nucleus of many testicular cells.
睾丸是微管网络最丰富的来源之一。这些网络包括有丝分裂和减数分裂纺锤体、精子细胞的袖套和轴丝，以及支持细胞的细胞骨架。微管由α-和β-微管蛋白亚基组成，这些亚基通过多种微管相关蛋白（MAPs）聚合并稳定下来。其中之一，MAP2，已被广泛表征为一种脑特异性蛋白，具有结合微管蛋白、cAMP依赖性激酶和钙调蛋白的能力。MAP2 mRNA被加工成至少两种变体，编码的蛋白质分别称为MAP2a、MAP2b和MAP2c。在编码MAP2a和MAP2b的9-kb mRNA中的5.7 kb编码序列中，大约4 kb的缺失产生了编码MAP2c的mRNA，它仅由MAP2b的N端和C端区域组成。为了确定MAP2是否存在于大鼠睾丸中，通过紫杉醇介导的聚合作用从成年大鼠睾丸和脑中分离微管制剂，并使用与MAP2反应的多克隆和单克隆抗体通过凝胶过滤、ELISA和蛋白质印迹法进行分析。在睾丸中检测到一种与MAP2c相对应的74-kDa蛋白。用区分已知MAP2剪接变体的cDNA探针对成年大鼠脑和睾丸的总RNA进行Northern印迹分析，证实了这些结果。睾丸中主要的6 kb和2.
