MICROVILLAR CYTOSKELETAL-MEMBRANE INTERACTIONS

对细胞骨架如何与膜相互作用的研究将解释 微丝起关键作用的几种细胞现象， 包括趋化、细胞移动、吞噬、胞质分裂和 细胞表面突起的形成。为了执行其中的许多细胞 功能，肌动蛋白细丝必须与血浆密切相关 薄膜。确定肌动蛋白细丝如何与膜相互作用， 例如，有助于阐明癌细胞是如何转移的。这个 肠道微绒毛显示清晰可见的连接核心的连接 肌动蛋白细丝到膜的束，这一尖的标识 组件是完全未知的。螺旋排列的横向链节 沿着微绒毛的长度可以看到延伸到膜的部分 核心；它们由一个110kD的多肽与钙调蛋白复合而成。 对110K-钙调蛋白复合体的研究可以提供所需的洞察 肌动蛋白细丝被锚定。110K-钙调蛋白与F-肌动蛋白的结合 ATP、钙和蛋白质浓度依赖，并表现出 协作性。此外，110K-钙调蛋白复合体类似于肌球蛋白 其修饰肌动蛋白细丝和水解三磷酸腺苷升高的能力 推测它是非肌肉细胞中的一种运动分子。的目标是 本研究旨在研究细胞骨架与细胞外基质的相互作用。 薄膜。特别是，我们将：(A)调查提纯的110K- 钙调蛋白将直接与脂质相互作用，(B)使用交联剂 用完整的微绒毛来探测一种可能有助于 将110K-钙调蛋白连接到膜上，然后对其进行表征 连接蛋白，(C)确定组成微绒毛的成分 TIP区，(D)寻找110K-钙调素蛋白在 其他组织类型，以及(E)继续绘制 100kD多肽包括鉴定ATP结合部位肌动蛋白- 结合部位和分子内钙调素结合部位。

The study of how the cytoskeleton interacts with the membrane will explain several cellular phenomena in which microfilaments play a critical role, including chemotaxis, cell locomotion, phagocytosis, cytokinesis, and the formation of cell surface projections. To perform many of these cellular functions, actin filaments must be intimately associated with the plasma membrane. Determining how actin filaments interact with the membrane will, for example, help to elucidate how cancer cells metastasize. The intestinal microvillus shows clearly visible links which connect a core bundle of actin filaments to the membrane, the identify of this tip component is completely unknown. Helically arranged lateral links extending to the membrane are visible down the length of the microvillar core; they are comprised of a 110 kD polypeptide complexed with calmodulin. Study of the 110K-calmodulin complex can provide needed insight into how actin filaments are anchored. Binding of 110K-calmodulin to F-actin is ATP, calcium, and protein concentration dependent and exhibits cooperativity. In addition, the 110K-calmodulin complex resembles myosin in its ability to decorate actin filaments and to hydrolyze ATP raising speculation that it is a motile molecule in non-muscle cells. The goal of this study is to characterize the interaction of the cytoskeleton with the membrane. In particular, we will: (a) investigate whether purified 110K- calmodulin will interact directly with lipid, (b) use cross-linking experiments on intact microvilli to probe for a protein which may serve to link 110K-calmodulin to the membrane, followed by characterization of this linker protein, (c) determine the components which comprise the microvillar tip region, (d) search for the presence of 110K-calmodulin proteins in other tissue types, and (e) continue to map the functional domains of the 100 kD polypeptide including identification of the ATP-binding site, actin- binding site, and calmodulin-binding site within the molecule.