Belkin A M, Zhidkova N I, Balzac F, Altruda F, Tomatis D, Maier A, Tarone G, Koteliansky V E, Burridge K
Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill 27599, USA.
J Cell Biol. 1996 Jan;132(1-2):211-26. doi: 10.1083/jcb.132.1.211.
The cytoplasmic domains of integrins provide attachment of these extracellular matrix receptors to the cytoskeleton and play a critical role in integrin-mediated signal transduction. In this report we describe the identification, expression, localization, and initial functional characterization of a novel form of beta 1 integrin, termed beta 1D. This isoform contains a unique alternatively spliced cytoplasmic domain of 50 amino acids, with the last 24 amino acids encoded by an additional exon. Of these 24 amino acids, 11 are conserved when compared to the beta 1A isoform, but 13 are unique (Zhidkova, N. I., A. M. Belkin, and R. Mayne. 1995. Biochem. Biophys. Res. Commun. 214:279-285; van der Flier, A., I. Kuikman, C. Baudoin, R, van der Neuf, and A. Sonnenberg. 1995. FEBS Lett. 369:340-344). Using an anti-peptide antibody against the beta 1D integrin subunit, we demonstrated that the beta 1D isoform is synthesized only in skeletal and cardiac muscles, while very low amounts of beta 1A were detected by immunoblot in striated muscles. Whereas beta 1A could not be detected in adult skeletal muscle fibers and cardiomyocytes by immunofluorescence, beta 1D was localized to the sarcolemma of both cell types. In skeletal muscle, beta 1D was concentrated in costameres, myotendinous, and neuromuscular junctions. In cardiac muscle this beta 1 isoform was found in costamers and intercalated discs. beta 1D was associated with alpha 7A and alpha 7B in adult skeletal muscle. In cardiomyocytes of adult heart, alpha 7B was the major partner for the beta 1D isoform. beta 1D could not be detected in proliferating C2C12 myoblasts, but it appeared immediately after myoblast fusion and its amount continued to rise during myotube growth and maturation. In contrast, expression of the beta 1A isoform was downregulated during myodifferentiation in culture and it was completely displaced by beta 1D in mature differentiated myotubes. We also analyzed some functional properties of the beta 1D integrin subunit. Expression of human beta 1D in CHO cells led to its localization at focal adhesions. Clustering of this integrin isoform on the cell surface stimulated tyrosine phosphorylation of pp125FAK (focal adhesion kinase) and caused transient activation of mitogen-activated protein (MAP) kinases. These data indicate that beta 1D and beta 1A integrin isoforms are functionally similar with regard to integrin-mediated signaling.
整合素的细胞质结构域使这些细胞外基质受体与细胞骨架相连，并在整合素介导的信号转导中发挥关键作用。在本报告中，我们描述了一种新型β1整合素（称为β1D）的鉴定、表达、定位及初步功能特性。这种异构体包含一个独特的由50个氨基酸组成的可变剪接细胞质结构域，其最后24个氨基酸由一个额外的外显子编码。与β1A异构体相比，这24个氨基酸中有11个是保守的，但有13个是独特的（Zhidkova, N. I., A. M. Belkin, and R. Mayne. 1995. Biochem. Biophys. Res. Commun. 214:279 - 285; van der Flier, A., I. Kuikman, C. Baudoin, R, van der Neuf, and A. Sonnenberg. 1995. FEBS Lett. 369:340 - 344）。使用针对β1D整合素亚基的抗肽抗体，我们证明β1D异构体仅在骨骼肌和心肌中合成，而通过免疫印迹在横纹肌中检测到的β1A含量非常低。通过免疫荧光在成年骨骼肌纤维和心肌细胞中无法检测到β1A，而β1D定位于这两种细胞类型的肌膜。在骨骼肌中，β1D集中在肌小节、肌纤维与肌腱连接点以及神经肌肉接头处。在心肌中，这种β1异构体存在于肌小节和闰盘中。在成年骨骼肌中，β1D与α7A和α7B相关联。在成年心脏的心肌细胞中，α7B是β1D异构体的主要结合伙伴。在增殖的C2C12成肌细胞中无法检测到β1D，但在成肌细胞融合后立即出现，并且其含量在肌管生长和成熟过程中持续增加。相反，β1A异构体的表达在培养的肌分化过程中下调，并且在成熟分化的肌管中完全被β1D取代。我们还分析了β1D整合素亚基的一些功能特性。人β1D在CHO细胞中的表达导致其定位于粘着斑。这种整合素异构体在细胞表面的聚集刺激了pp125FAK（粘着斑激酶）的酪氨酸磷酸化，并引起丝裂原活化蛋白（MAP）激酶的瞬时激活。这些数据表明，就整合素介导的信号传导而言，β1D和β1A整合素异构体在功能上相似。
