Calmodulin-driven Nuclear Trafficking linked to diabetes and insulin signaling

受调控的核运输对于维持转录因子、核蛋白激酶和复制因子的水平和活性至关重要。我们发现Ca+2/钙调蛋白是核输入的激活剂，并建议在细胞激活过程中，Ca+2在核输入的调节中发挥作用。这种进化上古老的钙依赖的输入途径被认为是在细胞激活过程中促进核蛋白的不同亚群的摄取。随着发现依赖钙调素的核输入缺陷是某些形式的人类性逆转的基础，钙调素调节的运输途径的生理学意义变得清晰起来。HMG-box结构转录因子SRY和SOX9必须进入Sertoli细胞核，并与靶DNA紧密结合，才能保证男性性腺的正常发育。因此，SRY是男性化的主要触发因子；SRY的核运输需要释放这一触发因子。在一组常染色体性反转(斯威尔斯综合征，驼核发育不良)的人类患者中，SRY和SOX9的核运输被阻断。与这些缺陷相关的突变位于SRY和SOX9的氨基末端附近的一个保守的钙调蛋白结合基序中。因此，SOX转录因子家族似乎使用Ca+2/钙调蛋白作为输入受体和分子开关，允许核输入和DNA结合。当三个胰岛素相关受体在小鼠中被消融时，类似的常染色体性反转表型发生，这表明细胞内的信号级联可能影响SRY和SOX9的正常功能。最近，编码TCF7L2的基因被证明是冰岛人群中的糖尿病易感基因。该转录因子是含有钙调蛋白结合基序的HMG-box家族的成员。钙调素对核蛋白的输入，虽然在功能上是冗余的，与经典的RAN依赖途径无关，但受到细胞内钙动员的独立调节。钙调蛋白依赖的导入异常是常染色体性反转患者的主要缺陷。这些性反转综合征是导致人类疾病的核进口途径缺陷的第一个直接例子。其他涉及SOX家族转录因子的人类疾病在依赖钙调素的核输入中也可能存在类似的缺陷。除了分析HMG-box转录因子在哺乳动物中的输入外，我们最近还发现依赖钙调素的核输入途径在酵母中发挥作用。我们目前正在采用基因方法来定义这一新的输入途径的其他成分，并试图确定这一过程的小分子抑制剂。由于SOX2蛋白对干细胞的多能性特别重要，我们目前的重点是了解核进口在其调控中的作用。在线虫中，POP-1是与BAR-1相关的关键结构转录因子，BAR-1是Beta-catenin的蠕虫同源物。这些研究是在酵母和线虫系统中进行的。目前正在从遗传学和生物化学的角度研究与已知的核运输途径的关系。这些新的方法正在转化为临床和基础科学研究的潜在成像策略。

Regulated nuclear transport is critical for the maintenance of the levels and activities of transcription factors, nuclear kinases, and replication factors. We identified Ca+2/calmodulin as an activator of nuclear import and suggested a role for Ca+2 in the regulation of nuclear import during cell activation. This evolutionarily ancient, calcium-dependent import pathway was proposed to facilitate uptake of a distinct subset of nuclear proteins during cell activation. The physiological significance of the Ca+2-calmodulin-regulated transport pathway became clear with the finding that defects in calmodulin-dependent nuclear import underlie certain forms of human sex reversal. The HMG-box architectural transcription factors SRY and SOX9 must enter the nucleus of Sertoli cells and bind tightly to target DNA for proper male gonad development. Thus, SRY acts as the primary trigger for maleness; nuclear transport of SRY is required to release this trigger. In a subset of human patients with autosomal sex reversal (Swyers syndrome, Campomelic dysplasia) nuclear transport of SRY and SOX9 is blocked. The mutations associated with these defects reside in a conserved calmodulin-binding motif near the amino terminus of SRY and SOX9. Thus, the SOX family of transcription factors appears to use Ca+2/calmodulin both as import receptor and molecular switch allowing for nuclear import and DNA binding. A similar autosomal sex reversal phenotype occurs when three insulin-related receptors are ablated in mice suggesting that intracellular signaling cascades may impact the normal functions of SRY and SOX9. More recently, the gene encoding TCF7L2 was shown to be a diabetes susceptibility locus in the Icelandic population. This transcription factor is a member of the HMG-box family containing a calmodulin-binding motif. The import of nuclear proteins by calmodulin, while functionally redundant with the canonic Ran-dependent pathway, is subject to independent regulation by intracellular Ca+2 mobilization. Abnormalities in calmodulin-dependent import emerge as the primary defect in patients with autosomal sex-reversal. These sex-reversal syndromes represent the first direct examples of a defect in a nuclear import pathway leading to human disease. Other human disorders involving SOX-family transcription factors may have similar defects in calmodulin dependent nuclear import. In addition to analysis of the import of the HMG-box transcription factors in mammals, we have recently shown that the calmodulin-dependent nuclear import pathway functions in yeast. We are currently taking genetic approaches to define additional components of this novel import pathway and seek to identify small molecule inhibitors of the process. Because the SOX2 protein is particularly important for Stem Cell pluripotency, our focus is currently on understanding the role of nuclear import in its regulation. In the nematode, pop-1 is a key architectural transcription factor associated with bar-1, the worm homolog of Beta-catenin. These studies are being carried out in the yeast and C. elegans systems. The relationship with known nuclear transport pathways is being investigated genetically and biochemically. These novel approaches are being translated into potential imaging strategies for both clinical and basic science studies.