Baker L P, Daggett D F, Peng H B
Department of Cell Biology and Anatomy, University of North Carolina at Chapel Hill 27599.
J Cell Sci. 1994 Jun;107 ( Pt 6):1485-97. doi: 10.1242/jcs.107.6.1485.
Focal adhesion kinase is a recently characterized tyrosine kinase that is concentrated at focal contacts in cultured cells. It is thought to play an important role in the regulation of the integrin-based signal transduction mechanism involved in the assembly of this membrane specialization. In this study, we examined the immunocytochemical distribution of focal adhesion kinase in Xenopus skeletal muscle and its role in the formation of two sarcolemmal specializations, the myotendinous junction and the neuromuscular junction, using a monoclonal antibody (2A7) against this protein. Immunoprecipitation of Xenopus embryonic tissues with this antibody demonstrated a single band at a relative molecular mass of 116 kDa. A distinct concentration of immunolabeling for focal adhesion kinase was observed at the myotendinous junction of muscle fibers in vivo. At this site, the labeling for this protein is correlated with an accumulation of phosphotyrosine immunolabeling. Focal adhesion kinase was not concentrated at the neuromuscular junction in muscle cells either in vivo or in vitro. However, it was localized at spontaneously formed acetylcholine receptor clusters in cultured Xenopus myotomal muscle cells, although its distribution was not exactly congruent with that of the receptors. In these cells, the accumulation focal adhesion kinase was induced by polystyrene microbeads. In addition, beads also induce the formation of acetylcholine receptor clusters and myotendinous junction-like specializations. By following the appearance of the focal adhesion kinase relative to the formation of these sarcolemmal specializations at bead-muscle contacts in cultured muscle cells, we conclude that the accumulation of this protein was in pace with the development of the myotendinous junction, but occurred well after the clustering of acetylcholine receptors. These results suggest that focal adhesion kinase may be involved in the development and/or maintenance of the myotendinous junction through an integrin-based signaling system. Although it can accumulate at acetylcholine receptor clusters formed in culture, it does not appear to be involved in the development of the neuromuscular junction.
粘着斑激酶是一种最近被鉴定的酪氨酸激酶，在培养细胞中集中于粘着斑。它被认为在基于整合素的信号转导机制的调节中起重要作用，该机制参与了这种膜特化结构的组装。在本研究中，我们使用针对该蛋白的单克隆抗体（2A7），研究了粘着斑激酶在非洲爪蟾骨骼肌中的免疫细胞化学分布及其在两种肌膜特化结构，即肌腱连接和神经肌肉接头形成中的作用。用该抗体对非洲爪蟾胚胎组织进行免疫沉淀，显示出一条相对分子质量为116 kDa的条带。在体内肌肉纤维的肌腱连接处观察到粘着斑激酶免疫标记的明显集中。在这个部位，该蛋白的标记与磷酸酪氨酸免疫标记的积累相关。粘着斑激酶在体内或体外的肌肉细胞的神经肌肉接头处均未集中。然而，它定位于培养的非洲爪蟾肌节肌细胞中自发形成的乙酰胆碱受体簇，尽管其分布与受体并不完全一致。在这些细胞中，聚苯乙烯微珠诱导了粘着斑激酶的积累。此外，微珠还诱导了乙酰胆碱受体簇和肌腱连接样特化结构的形成。通过追踪培养的肌肉细胞中粘着斑激酶相对于这些肌膜特化结构在微珠-肌肉接触处形成的出现情况，我们得出结论，该蛋白的积累与肌腱连接的发育同步，但在乙酰胆碱受体聚集之后才出现。这些结果表明，粘着斑激酶可能通过基于整合素的信号系统参与肌腱连接的发育和/或维持。尽管它可以在培养中形成的乙酰胆碱受体簇处积累，但似乎不参与神经肌肉接头的发育。
