Src kinases mediate VEGFR2 transactivation by the osteostatin domain of PTHrP to modulate osteoblastic function
Src kinases mediate VEGFR2 transactivation by the osteostatin domain of PTHrP to modulate osteoblastic function
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DOI：
10.1002/jcb.24482
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发表时间：
2013-06-01
影响因子：
4
通讯作者：
Esbrit, Pedro
中科院分区：
文献类型：
--
作者：
Garcia-Martin, Adela;Acitores, Alicia;Esbrit, Pedro
甲状旁腺相关蛋白（PTHrP）通过其N-和C-末端结构域刺激成骨细胞功能。由于后者域的成骨作用似乎至少部分取决于其与血管内皮生长因子（VEGF）系统的相互作用，我们的目的是探索这种相互作用在成骨细胞的假定机制。使用天然条件进行蛋白质提取和免疫印迹，我们发现PTHrP（107139）和较短的PTHrP（107111）肽（称为骨抑制素），在100 nM时，促进表观Mr. wt. 230 kDa，这可能代表其在小鼠成骨细胞MC 3 T3-E1细胞中通过二聚体形成而活化。此外，在MC 3 T3-E1和大鼠成骨细胞骨肉瘤UMR-106细胞中，osteostatin（100 nM）在510 min内最大限度地增加VEGFR 2在Tyr-1059处的磷酸化。这种由骨抑制素引起的磷酸化似乎是VEGF非依赖性的，但被VEGFR 2活化抑制剂SU 1498以及Src激酶抑制剂SU 6656和PP 1阻止。此外，骨抑制素诱导Src、细胞外信号调节激酶（ERK）和Akt的磷酸化，其时间过程与在这些成骨细胞中观察到的VEGFR 2活化相似。SU 6656消除了这种骨抑制素依赖性的ERK和Akt激活诱导。在这些成骨细胞中，SU 1498和SU 6656都阻止了VEGF和骨保护素基因表达的上调以及骨抑制素治疗诱导的促生存作用。总的来说，这些发现表明C末端PTHrP的骨抑制素结构域通过Src激活使VEGFR 2磷酸化，这代表了调节成骨细胞功能的机制。J.细胞。114：14041413，2013。(c)2012 Wiley Periodicals，Inc.
Parathyroid hormone-related protein (PTHrP) stimulates osteoblastic function through its N- and C-terminal domains. Since the osteogenic action of the latter domain appears to depend at least in part on its interaction with the vascular endothelial growth factor (VEGF) system, we aimed to explore the putative mechanism underlying this interaction in osteoblasts. Using native conditions for protein extraction and immunoblotting, we found that both PTHrP (107139) and the shorter PTHrP (107111) peptide (known as osteostatin), at 100nM, promoted the appearance of a VEGF receptor (VEGFR) 2 protein band of apparent Mr. wt. 230kDa, which likely represents its activation by dimer formation, in mouse osteoblastic MC3T3-E1 cells. Moreover, osteostatin (100nM) maximally increased VEGFR2 phosphorylation at Tyr-1059 within 510min in both MC3T3-E1 and rat osteoblastic osteosarcoma UMR-106 cells. This phosphorylation elicited by osteostatin appears to be VEGF-independent, but prevented by the VEGFR2 activation inhibitor SU1498 and also by the Src kinase inhibitors SU6656 and PP1. Furthermore, osteostatin induced phosphorylation of Src, extracellular signal-regulated kinase (ERK) and Akt with a similar time course to that observed for VEGFR2 activation in these osteoblastic cells. This osteostatin-dependent induction of ERK and Akt activation was abrogated by SU6656. Up-regulation of VEGF and osteoprotegerin gene expression as well as the pro-survival effect induced by osteostatin treatment were all prevented by both SU1498 and SU6656 in these osteoblastic cells. Collectively, these findings demonstrate that the osteostatin domain of C-terminal PTHrP phosphorylates VEGFR2 through Src activation, which represents a mechanism for modulating osteoblastic function. J. Cell. Biochem. 114: 14041413, 2013. (c) 2012 Wiley Periodicals, Inc.