Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, KS 66160, USA.
Biol Reprod. 2013 Feb 1;88(2):51. doi: 10.1095/biolreprod.112.103135. Print 2013 Feb.
DMRT1 is an evolutionarily conserved transcriptional factor expressed only in the postnatal testis, where it is produced in Sertoli cells and germ cells. While deletion of Dmrt1 in mice demonstrated it is required for postnatal testis development and fertility, much is still unknown about its temporal- and cell-specific functions. This study characterized a novel mouse model of DMRT1-deficient germ cells that was generated by breeding Dmrt1-null (Dmrt1(-/-)) mice with Wt1-Dmrt1 transgenic (Dmrt1(+/-;tg)) mice, which express a rat Dmrt1 cDNA in gonadal supporting cells by directing it from the Wilms tumor 1 locus in a yeast artificial chromosome transgene. Like Dmrt1(-/-) mice, male Dmrt1(-/-) transgenic mice (Dmrt1(-/-;tg)) were infertile, while female mice were fertile. Immunohistochemistry and Western blot analysis showed transgenic DMRT1 expressed in supporting cells of the newborn gonads of both sex and in Sertoli cells of the testis afterbirth. Sertoli cells were evaluated by electron microscopy, revealing that maturation of Dmrt1(-/-;tg) Sertoli cells was incomplete. Morphological analysis of testes from 42-day-old mice showed that, compared to Dmrt1(-/-) mice, Dmrt1(-/-;tg) mice have improved seminiferous tubule structure, with lumens present in many. Immunohistochemistry of the polarity markers ESPIN and NECTIN-2 showed that DMRT1 in Sertoli cells is required for NECTIN-2 expression and influences organization of ectoplasmic specializations. Further functional analyses of the transgene on a Dmrt1(-/-) background showed that it did not rescue the decrease in Dmrt1(-/-) testis size, but when expressed on a wild-type background, exogenous DMRT1 prevented the normal age-related decline in testis size and enhanced sperm progressive motility. The studies suggest that DMRT1 in Sertoli cells regulates tubule morphology, spermatogenesis, and sperm function via its effects on Sertoli cell maturation and polarity. Furthermore, expression and function of transgenic DMRT1 in Sertoli cells establishes a novel mouse model of DMRT1-deficient germ cells generated by breeding Dmrt1-null mice with Wt1-Dmrt1 transgenic mice (rescue; Dmrt1(-/-;tg)).
DMRT1 是一种进化上保守的转录因子，仅在出生后的睾丸中表达，在那里它由支持细胞和生殖细胞产生。虽然在小鼠中删除 Dmrt1 表明它是出生后睾丸发育和生育所必需的，但它的时间和细胞特异性功能仍知之甚少。本研究通过将 Dmrt1 缺失（Dmrt1（-/-））小鼠与 Wt1-Dmrt1 转基因（Dmrt1（+/-;tg））小鼠交配，产生了一种新型的 DMRT1 缺失生殖细胞小鼠模型，该模型通过将大鼠 Dmrt1 cDNA 从酵母人工染色体转基因中的 Wilms 肿瘤 1 基因座引导到性腺支持细胞中表达。与 Dmrt1（-/-）小鼠一样，雄性 Dmrt1（-/-）转基因小鼠（Dmrt1（-/-;tg））不育，而雌性小鼠可育。免疫组织化学和 Western blot 分析显示，转基因 DMRT1 在新生性腺的支持细胞中表达，在出生后的睾丸支持细胞中也有表达。电子显微镜评估支持细胞，结果显示 Dmrt1（-/-;tg）支持细胞的成熟不完全。对 42 天大的小鼠睾丸进行形态学分析表明，与 Dmrt1（-/-）小鼠相比，Dmrt1（-/-;tg）小鼠的生精小管结构得到改善，许多小管内存在管腔。极性标记物 ESPIN 和 NECTIN-2 的免疫组织化学显示，Sertoli 细胞中的 DMRT1 对于 NECTIN-2 的表达是必需的，并且影响外质特化的组织。进一步在 Dmrt1（-/-）背景下对转基因的功能分析表明，它不能挽救 Dmrt1（-/-）睾丸大小的减少，但当在野生型背景下表达时，外源性 DMRT1 可防止睾丸大小随年龄的正常下降，并增强精子的渐进运动。这些研究表明，Sertoli 细胞中的 DMRT1 通过影响 Sertoli 细胞成熟和极性来调节小管形态、生精作用和精子功能。此外，Sertoli 细胞中转基因 DMRT1 的表达和功能在 Dmrt1 缺失小鼠与 Wt1-Dmrt1 转基因小鼠交配产生的 DMRT1 缺失生殖细胞新型小鼠模型中建立（挽救；Dmrt1（-/-;tg））。
