REGULATION OF LYSOSOMAL ENZYME TRAFFICKING

溶酶体在真核细胞中起着重要的作用， 细胞和细胞外大分子的降解位点。 蛋白质向溶酶体的选择性递送涉及添加 蛋白质的共同地址标记（甘露糖6-磷酸标记），和 膜相关受体结合标记物并介导 蛋白质易位到溶酶体区室。 最近的研究 表明在某些条件下，这种靶向过程可以改变， 使得通常预定用于溶酶体的酶从溶酶体分泌， cell. 这种调节机制在生理上可能是重要的，因为 细胞外溶酶体酶可能在肿瘤生长中起重要作用 和转移以及正常细胞生长。 这项提案的长期目标是确定 用于溶酶体酶的正常和受调节的运输。 拟议 研究将涉及MEP的分子和细胞生物学研究， 一种溶酶体半胱氨酸蛋白酶，其运输受生长调节 因子和细胞转化，以及被认为介导 溶酶体运输：阳离子依赖性甘露糖6-磷酸（Man-6-P） 受体和Man-6-P/IGF-II受体。 MEP cDNA的定点突变和表达将用于 确定负责添加的蛋白质上的结构决定因素 Man-6-P标记 蛋白质和碳水化合物部分的分析 MEP，并检查生长因子对合成和 贩运的两个受体，将进行，以进一步 定义MEP分泌调节的基础。 这些研究将提供信息，以确定 负责选择性磷酸化的识别结构 溶酶体蛋白，并将有助于阐明参与的机制， 调节溶酶体蛋白的运输。

The lysosome plays an important role in eukaryotic cells as the primary site of degradation of both cellular and extracellular macromolecules. Selective delivery of proteins to the lysosome involves addition of a common address marker (the mannose 6-phosphate marker) to the proteins, and membrane-associated receptors that bind the marker and mediate translocation of the proteins to the lysosomal compartment. Recent studies suggest that under certain conditions this targeting process can be altered such that enzymes normally destined for lysosomes are secreted from the cell. This regulatory mechanism may be important physiologically since extracellular lysosomal enzymes may play important roles in tumor growth and metastasis as well as in normal cell growth. The long range goal of this proposal is to determine the molecular basis for normal and regulated trafficking of lysosomal enzymes. The proposed studies will involve molecular and cellular biological studies on MEP, a lysosomal cysteine proteinase whose trafficking is regulated by growth factors and cellular transformation, and the receptors thought to mediate lysosomal trafficking: the cation dependent mannose 6-phosphate (Man-6-P) receptor and the Man-6-P/IGF-II receptor. Site-specific mutagenesis and expression of MEP cDNA will be used to identify structural determinants on the protein responsible for addition of the Man-6-P marker. Analysis of the protein and carbohydrate portions of MEP, and examination of the effects of growth factors on the synthesis and trafficking of the two receptors, will be carried out in order to further define the basis for MEP's regulated secretion. These studies will provide information leading to determination of the recognition structure responsible for selective phosphorylation of lysosomal proteins and will help elucidate the mechanisms involved in regulated trafficking of lysosomal proteins.