Lewthwaite Jo C, Bastow Edward R, Lamb Katherine J, Blenis John, Wheeler-Jones Caroline P D, Pitsillides Andrew A
Department of Veterinary Basic Sciences, Royal Veterinary College, University of London, Royal College Street, London NW1 0TU, United Kingdom.
J Biol Chem. 2006 Apr 21;281(16):11011-8. doi: 10.1074/jbc.M510680200. Epub 2006 Feb 7.
Mechanisms regulating cell behavior and extracellular matrix composition in response to mechanical stimuli remain unresolved. Our previous studies have established that the MEK-ERK cascade plays a specific role in the mechano-dependent joint formation process by promoting the assembly of pericellular matrices reliant upon hyaluronan (HA) for their integrity. Here we demonstrate: (i) novel cross-talk between p38 MAPK and MEK-ERK signaling pathways that is specific for mechanical stimuli and (ii) a role for p38 MAPK in facilitating HA production by cells derived from the articular surface of embryonic chick tibiotarsal joints. We find that p38 MAPK blockade restricts pericellular assembly of HA-rich matrices and reduces basal as well as mechanical strain-induced release of HA. p38 MAPK blockers potentiated early strain-induced increases but restricted sustained increases in MEK/ERK phosphorylation at later times; c-Fos hyperphosphorylation at threonine 325 was found to parallel this p38 MAPK-mediated modulation of ERK activation. In contrast, p38 MAPK inhibitors had no detectable effect on the ERK activation induced by fibroblast growth factor 2 or pervanadate, a phosphatase inhibitor, and MEK inhibitors did not influence p38 MAPK phosphorylation, confirming both the specificity and unidirectionality of p38 MAPK-ERK cross-talk. Immunochemical and immunoblotting studies revealed constitutive p38 MAPK activation in cells at, or derived from, developing articular joint surfaces. Unlike the MEK-ERK pathway, however, p38 MAPK was not further stimulated by mechanical stimulation in vitro. Thus, p38 MAPK specifically facilitates ERK activation and downstream signaling in response to mechanical stimuli. These results suggest that constitutively active p38 MAPK serves an essential, permissive role in mechanically induced changes in ERK activation and in the accumulation of HA-rich extracellular matrices that serve a key role in joint development.
响应机械刺激时调节细胞行为和细胞外基质组成的机制仍未明确。我们之前的研究已经证实，MEK-ERK级联反应通过促进依赖透明质酸（HA）维持完整性的细胞周围基质的组装，在机械依赖的关节形成过程中发挥特定作用。在此，我们证明：（i）p38丝裂原活化蛋白激酶（MAPK）与MEK-ERK信号通路之间存在针对机械刺激的新型相互作用；（ii）p38 MAPK在促进胚胎鸡胫跗关节关节表面来源的细胞产生HA方面发挥作用。我们发现，p38 MAPK阻断会限制富含HA的基质在细胞周围的组装，并减少基础状态以及机械应变诱导的HA释放。p38 MAPK抑制剂增强了早期应变诱导的增加，但在后期限制了MEK/ERK磷酸化的持续增加；发现苏氨酸325处的c-Fos过度磷酸化与这种p38 MAPK介导的ERK激活调节平行。相反，p38 MAPK抑制剂对成纤维细胞生长因子2或磷酸酶抑制剂过氧钒酸盐诱导的ERK激活没有可检测到的影响，并且MEK抑制剂不影响p38 MAPK磷酸化，这证实了p38 MAPK-ERK相互作用的特异性和单向性。免疫化学和免疫印迹研究揭示了发育中的关节表面或其来源的细胞中存在组成性p38 MAPK激活。然而，与MEK-ERK途径不同，体外机械刺激不会进一步刺激p38 MAPK。因此，p38 MAPK特异性地促进ERK激活和下游信号传导以响应机械刺激。这些结果表明，组成性激活的p38 MAPK在机械诱导的ERK激活变化以及在关节发育中起关键作用的富含HA的细胞外基质积累中发挥重要的允许作用。
