Osteoblast differentiation and migration are regulated by dynamin GTPase activity

Abstract

Bone formation is controlled by osteoblasts but the signaling proteins that control osteoblast differentiation and function are still unclear. We examined if the dynamin GTPase, which is associated with actin remodeling and migration in other cells, plays a role in osteoblast differentiation and migration. Dynamin mRNA was expressed in primary osteoblasts throughout differentiation (0-21 days). However, alkaline phosphatase (ALP) activity, a marker of osteoblast differentiation, was decreased in osteoblasts over-expressing dynamin. Conversely, ALP activity was increased following shRNA-mediated knockdown of dynamin and in osteoblasts treated with the dynamin inhibitor, dynasore. Dynasore also reduced c-fos and osterix expression, markers of early osteoblasts, suggesting a role for dynamin in pre-osteoblast to osteoblast differentiation. Since dynamin GTPase activity is regulated by tyrosine phosphorylation, we examined the mechanism of dynamin dephosphorylation in osteoblasts. Dynamin formed a protein complex with the tyrosine phosphatase PTP-PEST and inhibition of phosphatase activity increased the level of phosphorylated dynamin. Further, PTP-PEST blocked the Src-mediated increase in the phosphorylation and GTPase activity of wild-type dynamin but not the phosphorylation mutant dynY231F/Y597F. Although ALP activity was increased in osteoblasts expressing GTPasedefective dynK44A, and to a lesser extent dynY231F/Y597F, osteoblast migration was significantly inhibited by dynK44A and dynY231F/Y597F. These studies demonstrate a novel role for dynamin GTPase activity and phosphorylation in osteoblast differentiation and migration, which may be important for bone formation.

Discovery of the migrasome, an organelle mediating release of cytoplasmic contents during cell migration

“…To further validate the hypothesis, we treated cells with the dynamin inhibitor 'Dynasore' and the myosin II inhibitor 'Blebbistatin', both of which have been widely used to block cell migration 9 , 10 . We found that these migration inhibitors markedly reduced the number of PLSs, and close analysis revealed that the formation of new PLSs was largely blocked ( Figure 4F and 4G , and Supplementary information, Movies S3-S5 ).…”

1,8-Naphthalimide derivatives: new leads against dynamin I GTPase activity

“…Sulfonation of 1,8-naphthalic anhydride (4) at 120ºC gave 5 in a 96% yield. With the substituted naphthalimides (2)(3)(4)(5) in hand, the Nimide substituted analogues were derived through condensation with the desired amine (Scheme 2). The anhydride-amine condensation procedure was amine dependent, with each amine requiring subtle protocol optimisation to afford useful yields.…”

Effects of dietary calcium to available phosphorus ratios on bone metabolism and osteoclast activity of the OPG /RANK/RANKL signalling pathway in piglets

“…Moreover, we showed that osteoblast activity is maximal when Ca/AP ratio reaches 2:1. ALP (Eleniste, Huang, Wayakanon, Largura, & Bruzzaniti, ) and OCN (Lenders et al, ) are markers of osteoblast activity. ALP hydrolyses inorganic pyrophosphates that inhibit bone mineralization, improves osteoblast activity, increases bone mass and enhances bone metabolism (Hurwitz, Fishman, & Talpaz, ).…”

“…ALP (Eleniste, Huang, Wayakanon, Largura, & Bruzzaniti, 2014) and OCN (Lenders et al, 2013) are markers of osteoblast activity.…”