Guan Shengxi, Fan Jianhua, Han Arum, Chen Mei, Woodley David T, Li Wei
Department of Dermatology and USC-Norris Comprehensive Cancer Center, Keck School of Medicine, The University of Southern California, Los Angeles, California 90033, USA.
J Invest Dermatol. 2009 Aug;129(8):1909-20. doi: 10.1038/jid.2008.457. Epub 2009 Feb 26.
Platelet-derived growth factor BB (PDGF-BB) is a Food and Drug Administration (FDA)-approved growth factor, acting as a mitogen and motogen of dermal fibroblasts (DFs), for skin wound healing. The two closely related SH2/SH3 adapter proteins, Nckalpha and Nckbeta, connect PDGF-BB signaling to the actin cytoskeleton and cell motility. The mechanism has not been fully understood. In this study, we investigated, side by side, the roles of Nckalpha and Nckbeta in PDGF-BB-stimulated DF migration. We found that cells expressing the PDGFRbeta-Y751F mutant (preventing Nckalpha binding) or PDGFRbeta-Y1009F mutant (preventing Nckbeta binding), DF cells isolated from Nckalpha- or Nckbeta-knockout mice, and primary human DF cells with RNA interference (RNAi) knockdown of the endogenous Nckalpha or Nckbeta all failed to migrate in response to PDGF-BB. Overexpression of the middle SH3 domain of Nckalpha or Nckbeta alone in human DFs also blocked PDGF-BB-induced cell migration. However, neither Nckalpha nor Nckbeta was required for the activation of the PDGF receptor, p21-activated protein kinase (Pak1), AKT, extracellular signal-regulated kinase (ERK) 1/2, or p38MAP by PDGF-BB. Although PDGF-BB stimulated the membrane translocation of both Nckalpha and Nckbeta, Nckalpha appeared to mediate Cdc42 signaling for filopodium formation, whereas Nckbeta mediated Rho signaling to induce stress fibers. Thus, this study has elucidated the independent roles and mechanisms of action of Nckalpha and Nckbeta in DF migration, which is critical for wound healing.
血小板衍生生长因子BB（PDGF - BB）是一种经美国食品药品监督管理局（FDA）批准的生长因子，在皮肤伤口愈合过程中作为真皮成纤维细胞（DFs）的促分裂原和促细胞运动因子。两种密切相关的SH2/SH3衔接蛋白Nckα和Nckβ，将PDGF - BB信号传导与肌动蛋白细胞骨架及细胞运动联系起来。其机制尚未完全明确。在本研究中，我们同时探究了Nckα和Nckβ在PDGF - BB刺激的DF迁移中的作用。我们发现，表达PDGFRβ - Y751F突变体（阻止Nckα结合）或PDGFRβ - Y1009F突变体（阻止Nckβ结合）的细胞、从Nckα或Nckβ基因敲除小鼠分离得到的DF细胞，以及通过RNA干扰（RNAi）敲低内源性Nckα或Nckβ的原代人DF细胞，均无法对PDGF - BB作出迁移反应。仅在人DF细胞中过表达Nckα或Nckβ的中间SH3结构域，也会阻断PDGF - BB诱导的细胞迁移。然而，PDGF - BB激活PDGF受体、p21激活蛋白激酶（Pak1）、AKT、细胞外信号调节激酶（ERK）1/2或p38丝裂原活化蛋白激酶（p38MAP）时，既不需要Nckα也不需要Nckβ。虽然PDGF - BB刺激了Nckα和Nckβ两者向膜的转位，但Nckα似乎介导Cdc42信号传导以形成丝状伪足，而Nckβ介导Rho信号传导以诱导应力纤维。因此，本研究阐明了Nckα和Nckβ在DF迁移中的独立作用及作用机制，这对伤口愈合至关重要。
