Lynch Grit, Kohler Sandra, Leser Juergen, Beil Michael, Garcia-Marin Luis J, Lutz Manfred P
Department of Internal Medicine I, University of Ulm, Ulm, Germany.
Pflugers Arch. 2004 Jan;447(4):445-51. doi: 10.1007/s00424-003-1188-7. Epub 2003 Nov 21.
Acinar cells require a functional apical actin web for secretion. During stimulation with supraphysiological concentrations of cholecystokinin (CCK), a condition that mimics acute pancreatitis, the actin filaments disintegrate. This leads to retention of secretory enzymes and, together with their premature activation, results in cell injury. Actin filaments are anchored through membrane-associated protein complexes that can be regulated through Src-family kinases in some model systems. Here we show that the Src-family kinases Yes and Lyn, but not Src and Fyn, are expressed in isolated pancreatic acini of Wistar rats. Upon stimulation with supramaximal secretory CCK (10(-8) M), Yes became reversibly tyrosine-phosphorylated and activated within 2 min. Immunocytochemical and subcellular fractionation studies showed reversible redistribution of Yes to the apical actin web and to the membrane fraction within 5 min. Coimmunoprecipitation demonstrated that Yes forms a complex with the focal adhesion protein Pyk2, which increased with CCK stimulation. In functional studies, inhibition of Src-kinase activity with PP2 partially reversed actin disintegration and also restored amylase secretion. We conclude that Yes participates in the regulation of the acinar cell actin, probably by interaction with Pyk2.
腺泡细胞分泌需要一个功能性的顶端肌动蛋白网。在用超生理浓度的胆囊收缩素（CCK）刺激时，这种情况模拟了急性胰腺炎，肌动蛋白丝会解体。这导致分泌酶的潴留，并与其过早激活一起导致细胞损伤。在一些模型系统中，肌动蛋白丝通过与膜相关的蛋白复合物锚定，这些复合物可通过Src家族激酶进行调节。在这里，我们表明Src家族激酶Yes和Lyn，而不是Src和Fyn，在Wistar大鼠的分离胰腺腺泡中表达。在用超最大分泌性CCK（10^(-8) M）刺激后，Yes在2分钟内可逆地酪氨酸磷酸化并被激活。免疫细胞化学和亚细胞分级分离研究表明，Yes在5分钟内可逆地重新分布到顶端肌动蛋白网和膜部分。免疫共沉淀表明，Yes与粘着斑蛋白Pyk2形成复合物，该复合物随CCK刺激而增加。在功能研究中，用PP2抑制Src激酶活性部分逆转了肌动蛋白解体，并恢复了淀粉酶分泌。我们得出结论，Yes可能通过与Pyk2相互作用参与腺泡细胞肌动蛋白的调节。
