Hossain M Z, Jagdale A B, Ao P, Boynton A L
Molecular Medicine, Northwest Hospital, Seattle, Washington 98125, USA.
J Cell Physiol. 1999 Apr;179(1):87-96. doi: 10.1002/(SICI)1097-4652(199904)179:1<87::AID-JCP11>3.0.CO;2-K.
Disruption of gap junctional communication (GJC) by various compounds, including growth factors and tumor promoters, is believed to be modulated by the phosphorylation of a gap junctional protein, connexin43 (Cx43). We have previously demonstrated a platelet-derived growth factor (PDGF)-induced blockade of GJC and phosphorylation of Cx43 in T51B rat liver epithelial cells expressing wild-type PDGF receptor beta (PDGFr beta). Both of these actions of PDGF required participation of protein kinase C (PKC) and mitogen-activated protein kinase (MAPK). Similar requirements of MAPK were suggested in the modulation of GJC by other agents, including epidermal growth factor (EGF) and lysophosphatidic acid (LPA). Since many of these agents activate additional protein kinases, our present study examined whether activation of MAPK was sufficient for Cx43 phosphorylation and GJC blockade. By utilizing a variety of MAPK activators, we now show that activation of MAPK is not always associated with either Cx43 phosphorylation or disruption of GJC, which suggests a requirement for additional factors. Furthermore, pretreatment with hydrogen peroxide (H2O2), a potent MAPK activator but inefficient GJC/Cx43 modulator, abrogated PDGF- or TPA-induced disruption of GJC. While a 5 min H2O2 pretreatment abolished both PDGF- and TPA-induced Cx43 phosphorylation and GJC blockade, a simultaneous H2O2 treatment interfered only with GJC closure but not with the phosphorylation of Cx43 induced by PDGF and TPA. This finding indicates that, in addition to the Cx43 phosphorylation step, inhibition of GJC requires interaction with other components. H2O2-mediated abrogation of PDGF/TPA signaling can be neutralized by the antioxidant N-acetylcysteine (NAC) or by the tyrosine kinase inhibitor genistein. Taken together, our results suggest that disruption of GJC is not solely mediated by either activated MAPK or Cx43 phosphorylation but requires the participation of additional kinases and regulatory components. This complex mode of regulation is perhaps essential for the proposed functional role of GJC.
包括生长因子和肿瘤启动子在内的多种化合物对间隙连接通讯（GJC）的破坏，被认为是由间隙连接蛋白连接蛋白43（Cx43）的磷酸化所调节。我们之前已经证明，血小板衍生生长因子（PDGF）可诱导表达野生型PDGF受体β（PDGFrβ）的T51B大鼠肝上皮细胞中GJC的阻断以及Cx43的磷酸化。PDGF的这两种作用都需要蛋白激酶C（PKC）和丝裂原活化蛋白激酶（MAPK）的参与。其他因子（包括表皮生长因子（EGF）和溶血磷脂酸（LPA））对GJC的调节也提示了对MAPK有类似的需求。由于这些因子中的许多都会激活其他蛋白激酶，我们目前的研究探讨了MAPK的激活是否足以导致Cx43磷酸化和GJC阻断。通过使用多种MAPK激活剂，我们现在表明，MAPK的激活并不总是与Cx43磷酸化或GJC破坏相关，这表明还需要其他因素。此外，用过氧化氢（H2O2）预处理，过氧化氢是一种有效的MAPK激活剂但对GJC/Cx43调节效率不高，可消除PDGF或佛波酯（TPA）诱导的GJC破坏。虽然5分钟的H2O2预处理消除了PDGF和TPA诱导的Cx43磷酸化和GJC阻断，但同时进行H2O2处理仅干扰GJC关闭，而不影响PDGF和TPA诱导的Cx43磷酸化。这一发现表明，除了Cx43磷酸化步骤外，GJC的抑制还需要与其他成分相互作用。H2O2介导的对PDGF/TPA信号的消除可被抗氧化剂N-乙酰半胱氨酸（NAC）或酪氨酸激酶抑制剂染料木黄酮中和。综上所述，我们的结果表明，GJC的破坏并非仅由激活的MAPK或Cx43磷酸化介导，而是需要其他激酶和调节成分的参与。这种复杂的调节模式可能对于GJC所提出的功能作用至关重要。
