The role of forkhead box A2 to restrict androgen-regulated gene expression of lipocalin 5 in the mouse epididymis.
The role of forkhead box A2 to restrict androgen-regulated gene expression of lipocalin 5 in the mouse epididymis.
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DOI：
10.1210/me.2006-0008
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发表时间：
2006-10
影响因子：
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通讯作者：
Xiuping Yu;Kichiya Suzuki;Yongqing Wang;Aparna Gupta;R. Jin;M. Orgebin‐Crist;R. Matusik
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文献类型：
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作者：
Xiuping Yu;Kichiya Suzuki;Yongqing Wang;Aparna Gupta;R. Jin;M. Orgebin‐Crist;R. Matusik
小鼠附睾维A酸结合蛋白[或Lipocalin5(Lcn5)]是由小鼠附睾头中部/远端的主细胞合成和分泌的。Lcn5基因的5kb启动子片段可以在转基因小鼠中决定雄激素依赖性和附睾区特异性基因的表达。在这里，我们报道了1.8kb的Lcn5启动子在转基因小鼠中赋予附睾区特异性基因表达。为了解释转录调控的机制，我们构建了14个嵌合载体，依次去除了1.8kb的Lcn5启动子100bp，并将其导入附睾腺细胞和非附睾腺细胞。瞬时转染分析表明，1.3kb启动子片段对雄激素的反应最强。在1.2kb到1.3kb区域之间，发现了两个雄激素受体(AR)结合位点。凝胶位移分析证实，在AR结合位点附近有一个Foxa2[Fox(Forkhead Box)A亚类]结合位点。在人和大鼠Lcn5的启动子上也发现了相似的FOXA结合位点，表明FOXA结合位点在物种之间是保守的。我们先前报道，在FOXA家族的三个成员中，FOXA1和Foxa3在附睾中缺失，而Foxa2在附睾主细胞中检测到。在这里，我们报告了Foxa2在附睾部的区域特异性表达模式：在起始段没有观察到染色，近端染色浅，中段和远端染色逐渐加深，在体部和尾部表达最强，Lcn5表达很少或没有表达。在瞬时转染实验中，Foxa2的表达抑制了Lcn5启动子的AR诱导，这与Lcn5在体部和尾部缺乏表达的情况是一致的。我们得出结论，Foxa2作为一种阻滞剂，将Lcn5的AR调节限制在附睾节段特异性模式。
Murine epididymal retinoic acid-binding protein [or lipocalin 5 (Lcn5)] is synthesized and secreted by the principal cells of the mouse middle/distal caput epididymidis. A 5-kb promoter fragment of the Lcn5 gene can dictate androgen-dependent and epididymis region-specific gene expression in transgenic mice. Here, we reported that the 1.8-kb Lcn5 promoter confers epididymis region-specific gene expression in transgenic mice. To decipher the mechanism that directs transcription, 14 chimeric constructs that sequentially removed 100 bp of 1.8-kb Lcn5 promoter were generated and transfected into epididymal cells and nonepididymal cells. Transient transfection analysis revealed that 1.3 kb promoter fragment gave the strongest response to androgens. Between the 1.2-kb to 1.3-kb region, two androgen receptor (AR) binding sites were identified. Adjacent to AR binding sites, a Foxa2 [Fox (Forkhead box) subclass A] binding site was confirmed by gel shift assay. Similar Foxa binding sites were also found on the promoters of human and rat Lcn5, indicating the Foxa binding site is conserved among species. We previously reported that among the three members of Foxa family, Foxa1 and Foxa3 were absent in the epididymis whereas Foxa2 was detected in epididymal principal cells. Here, we report that Foxa2 displays a region-specific expression pattern along the epididymis: no staining observed in initial segment, light staining in proximal caput, gradiently heavier staining in middle and distal caput, and strongest staining in corpus and cauda, regions with little or no expression of Lcn5. In transient transfection experiments, Foxa2 expression inhibits AR induction of the Lcn5 promoter, which is consistent with the lack of expression of Lcn5 in the corpus and cauda. We conclude that Foxa2 functions as a repressor that restricts AR regulation of Lcn5 to a segment-specific pattern in the epididymis.