Valcárcel J, Singh R, Zamore P D, Green M R
Program in Molecular Medicine, University of Massachusetts Medical Center, Worcester 01605.
Nature. 1993 Mar 11;362(6416):171-5. doi: 10.1038/362171a0.
Somatic sexual differentiation in Drosophila melanogaster involves a cascade of regulated splicing events and provides an attractive model system for the analysis of alternative splicing mechanisms. The protein Sex-lethal (Sxl) activates a female-specific 3' splice site in the first intron of transformer (tra) pre-mRNA while repressing an alternative non-sex-specific site. We have developed an in vitro system that recapitulates this regulation in a manner consistent with genetic, transfection and fly transformation studies. Using this system, we have determined the molecular basis of the splice site switch. Here we show that Sxl inhibits splicing to the non-sex-specific (default) site by specifically binding to its polypyrimidine tract, blocking the binding of the essential splicing factor U2AF. This enables U2AF to activate the lower-affinity female-specific site. A splicing 'effector' domain present in U2AF but absent from Sxl accounts for the different activities of these two polypyrimidine-tract-binding proteins: addition of the U2AF effector domain to Sxl converts it from a splicing repressor to an activator and renders it unable to mediate splice-site switching.
黑腹果蝇的体细胞性分化涉及一系列受调控的剪接事件，为分析可变剪接机制提供了一个有吸引力的模型系统。蛋白质性致死因子（Sxl）激活transformer（tra）前体mRNA第一个内含子中的一个雌性特异性3'剪接位点，同时抑制一个非性别特异性的可变位点。我们开发了一种体外系统，该系统以与遗传、转染和果蝇转化研究一致的方式重现了这种调控。利用这个系统，我们确定了剪接位点转换的分子基础。在这里我们表明，Sxl通过特异性结合其多嘧啶序列来抑制向非性别特异性（默认）位点的剪接，从而阻止必需剪接因子U2AF的结合。这使得U2AF能够激活亲和力较低的雌性特异性位点。U2AF中存在但Sxl中不存在的一个剪接“效应器”结构域解释了这两种多嘧啶序列结合蛋白的不同活性：将U2AF效应器结构域添加到Sxl中会使其从剪接抑制因子转变为激活因子，并使其无法介导剪接位点转换。
