Chakraborty Papia, William Buaas F, Sharma Manju, Smith Benjamin E, Greenlee Anne R, Eacker Stephen M, Braun Robert E
The Jackson Laboratory (P.C., F.W.B., M.S., B.E.S., A.R.G., R.E.B.), Bar Harbor, Maine 04609; and Department of Neurology (S.M.E.), Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Mol Endocrinol. 2014 Jul;28(7):1055-72. doi: 10.1210/me.2013-1134. Epub 2014 May 13.
Sertoli cell tight junctions (SCTJs) of the seminiferous epithelium create a specialized microenvironment in the testis to aid differentiation of spermatocytes and spermatids from spermatogonial stem cells. SCTJs must be chronically broken and rebuilt with high fidelity to allow the transmigration of preleptotene spermatocytes from the basal to adluminal epithelial compartment. Impairment of androgen signaling in Sertoli cells perturbs SCTJ remodeling. Claudin (CLDN) 3, a tight junction component under androgen regulation, localizes to newly forming SCTJs and is absent in Sertoli cell androgen receptor knockout (SCARKO) mice. We show here that Cldn3-null mice do not phenocopy SCARKO mice: Cldn3(-/-) mice are fertile, show uninterrupted spermatogenesis, and exhibit fully functional SCTJs based on imaging and small molecule tracer analyses, suggesting that other androgen-regulated genes must contribute to the SCARKO phenotype. To further investigate the SCTJ phenotype observed in SCARKO mutants, we generated a new SCARKO model and extensively analyzed the expression of other tight junction components. In addition to Cldn3, we identified altered expression of several other SCTJ molecules, including down-regulation of Cldn13 and a noncanonical tight junction protein 2 isoform (Tjp2iso3). Chromatin immunoprecipitation was used to demonstrate direct androgen receptor binding to regions of these target genes. Furthermore, we demonstrated that CLDN13 is a constituent of SCTJs and that TJP2iso3 colocalizes with tricellulin, a constituent of tricellular junctions, underscoring the importance of androgen signaling in the regulation of both bicellular and tricellular Sertoli cell tight junctions.
生精上皮的支持细胞紧密连接（SCTJs）在睾丸中创造了一个特殊的微环境，以帮助精原干细胞分化为精母细胞和精子细胞。SCTJs必须长期被高度精确地破坏和重建，以允许前细线期精母细胞从基底上皮区室迁移到管腔上皮区室。支持细胞中雄激素信号传导的受损会扰乱SCTJ重塑。Claudin（CLDN）3是一种受雄激素调节的紧密连接成分，定位于新形成的SCTJs，在支持细胞雄激素受体敲除（SCARKO）小鼠中不存在。我们在此表明，Cldn3基因敲除小鼠并不表现出与SCARKO小鼠相同的表型：Cldn3(-/-)小鼠可育，精子发生不间断，并且基于成像和小分子示踪分析显示其SCTJs功能完全正常，这表明其他受雄激素调节的基因必定导致了SCARKO表型。为了进一步研究在SCARKO突变体中观察到的SCTJ表型，我们构建了一个新的SCARKO模型，并广泛分析了其他紧密连接成分的表达。除了Cldn3，我们还鉴定出其他几种SCTJ分子的表达发生了改变，包括Cldn13的下调和一种非典型紧密连接蛋白2异构体（Tjp2iso3）。染色质免疫沉淀用于证明雄激素受体直接结合到这些靶基因的区域。此外，我们证明CLDN13是SCTJs的一个组成部分，并且TJP2iso3与三细胞连接的一个组成部分tricellulin共定位，强调了雄激素信号传导在调节双细胞和三细胞支持细胞紧密连接中的重要性。
