Molecular and Functional Characterization of Mucolipin-1

这项指导职业发展奖的主要目标是为申请者的职业生涯做好准备 作为神经退行性变和钙(2)信号研究的独立研究员。申请人 对生物医学研究事业表现出坚定的承诺，并获得了广泛的 在溶酶体的细胞和分子生物学、生物化学和基因靶向方面的培训和专业知识 然而，她认为，对储存疾病进行一段时间的额外培训后，就会勾勒出 在这项拟议研究的背景下，电生理技术将极大地改善她 作为一名独立科学家的竞争力。申请人所处的环境非常适合支持 她的事业继续发展，拥有最先进的核心设施和现场科学 专家。她的导师穆阿利姆博士是钙信号和离子通道调节方面的专家 在指导年轻调查人员方面有丰富的经验。 候选人的近期目标是阐明Trp-ML1(ML1)的细胞功能，并 确定溶酶体膜离子通道的缺陷如何导致IV型粘脂病(MLIV)， 一种脂肪储存神经退行性疾病。MLIV是由溶酶体在细胞水平上定义的 从晚期开始脂类的积累和成熟溶酶体的形成受阻 内体/杂合细胞器。ML1的确切功能尚不清楚。在初步研究中，我们有 结果表明，ML1是一种H选择性阳离子通道，在溶酶体中被特异性地切割。这个 裂解形式主要见于天然细胞和组织中，可能具有调节脂肪酶的功能 活动。我们现在假设ML1是一种调节溶酶体脂类的双重功能蛋白 通过调节溶酶体pH的变化来代谢。在缺乏功能性ML1的情况下，脂质 新陈代谢异常。以下具体目标将检验这一假设：1)描述 溶酶体靶向性和蛋白水解性切割的分子决定因素，2)确定哪些脂肪酶是 被ML1激活，3)表征ML1与MLS以及与其他可能 在综合体中发挥作用。这些研究的结果将为这种疾病提供细胞和治疗基础。 疾病。

The primary goal of this Mentored Career Development Award is to prepare the applicant for a career as an independent investigator in neurodegenerative and Ca(2+) signaling research. The applicant has shown a firm commitment to a biomedical research career and has acquired extensive prior training and expertise in cell and molecular biology, biochemistry, and gene targeting of lysosomal storage diseases, however, she believes that a period of additional training in the outlined imaging and electrophysiological techniques in the context of this proposed study will dramatically improve her competitiveness as an independent scientist. The applicant's environment is ideally suited to support her continued career development, with access to state-of-the-artcore facilities and on-site scientific experts. Her mentor, Dr. Muallem, is an expert in Ca(2+) signaling and in the regulation of ion channel functions and has had extensive experience in mentoring young investigators. The candidate's immediate goal is to elucidate the cellular functions of TRP-ML1 (ML1), and to determine how defects in this lysosomal membrane ion channel lead to mucolipidosis type IV (MLIV), a lipid storage neurodegenerative disorder. MLIV is defined at the cellular level by a lysosomal accumulation of lipids and by a block in the formation of mature lysosomes from late endosomes/hybrid organelles. The exact function of ML1 is unknown. In preliminary studies, we have shown that ML1 is a H+ selective cation channel that is specifically cleaved in the lysosomes. The cleaved form is predominantly seen in native cells and tissues and may function to regulate lipase activity. We now hypothesize that ML1 is a dual function protein that regulates lysosomal lipid metabolism by mediating alterations in lysosomal pH. In the absence of a functional ML1, lipid metabolism is aberrant. The following specific aims will test this hypothesis: 1) Characterize the molecular determinants of lysosomal targeting and proteolytic cleavage, 2) Identify which lipases are activated by ML1, 3) Characterize the interactions of ML1 with MLS and with other proteins that may function in a complex. Results from these studies will provide the cellular and therapeutic basis of this disease.