Periodic mechanical stress activates MEK1/2-ERK1/2 mitogenic signals in rat chondrocytes through Src and PLCγ1

The mitogenic effects of periodic mechanical stress on chondrocytes have been studied extensively but the mechanisms whereby chondrocytes sense and respond to periodic mechanical stress remain a matter of debate. We explored the signal transduction pathways of chondrocyte proliferation and matrix synthesis under periodic mechanical stress. In particular, we sought to identify the role of the MEK1/2-ERK1/2 signaling pathway in chondrocyte proliferation and matrix synthesis following cyclic physiologic mechanical compression. Under periodic mechanical stress, both rat chondrocyte proliferation and matrix synthesis were significantly increased (P < 0.05) and were associated with increases in the phosphorylation of Src, PLCγ1, MEK1/2, and ERK1/2 (P < 0.05). Pretreatment with the MEK1/2-ERK1/2 selective inhibitor, PD98059, and shRNA targeted to ERK1/2 reduced periodic mechanical stress-induced chondrocyte proliferation and matrix synthesis (P < 0.05), while the phosphorylation levels of Src-Tyr418 and PLCγ1-Tyr783 were not inhibited. Proliferation, matrix synthesis and phosphorylation of MEK1/2-Ser217/221 and ERK1/2-Thr202/Tyr204 were inhibited after pretreatment with the PLCγ1 inhibitor U73122 in chondrocytes in response to periodic mechanical stress (P < 0.05), while the phosphorylation site of Src-Tyr418 was not affected. Inhibition of Src activity with PP2 and shRNA targeted to Src abrogated chondrocyte proliferation and matrix synthesis (P < 0.05) and attenuated PLCγ1, MEK1/2 and ERK1/2 activation in chondrocytes subjected to periodic mechanical stress (P < 0.05). These findings suggest that periodic mechanical stress promotes chondrocyte proliferation and matrix synthesis in part through the Src-PLCγ1-MEK1/2-ERK1/2 signaling pathway, which links these three important signaling molecules into a mitogenic cascade.

Phosphatidylinositol phosphates directly bind to neurofilament light chain (NF-L) for the regulation of NF-L self assembly

Phosphatidylinositol phosphates (PtdInsPs) are ubiquitous membrane phospholipids that play diverse roles in cell growth and differentiation. To clarify the regulation mechanism acting on neurofilament light chain (NF-L) self assembly, we examined the effects of various PtdInsPs on this process. We found that PtdInsPs, including PI(4,5)P2, directly bind to the positively charged Arg54 of murine NF-L, and this binding promotes NF-L self assembly in vitro. Mutant NF-L (R53A/R54A) proteins lacking binding affinity to PtdInsPs did not have the same effect, but the mutant NF-L proteins showed greater self assembly than the wild-type in the absence of any PtdInsP. These results collectively suggest that Arg54 plays a pivotal role in NF-L self assembly by binding with PtdInsPs.