Dataset Information

The adaptor protein p62 is involved in RANKL-induced autophagy and osteoclastogenesis.

ABSTRACT: Previous studies have implicated autophagy in osteoclast differentiation. The aim of this study was to investigate the potential role of p62, a characterized adaptor protein for autophagy, in RANKL-induced osteoclastogenesis. Real-time quantitative PCR and western blot analyses were used to evaluate the expression levels of autophagy-related markers during RANKL-induced osteoclastogenesis in mouse macrophage-like RAW264.7 cells. Meanwhile, the potential relationship between p62/LC3 localization and F-actin ring formation was tested using double-labeling immunofluorescence. Then, the expression of p62 in RAW264.7 cells was knocked down using small-interfering RNA (siRNA), followed by detecting its influence on RANKL-induced autophagy activation, osteoclast differentiation, and F-actin ring formation. The data showed that several key autophagy-related markers including p62 were significantly altered during RANKL-induced osteoclast differentiation. In addition, the expression and localization of p62 showed negative correlation with LC3 accumulation and F-actin ring formation, as demonstrated by western blot and immunofluorescence analyses, respectively. Importantly, the knockdown of p62 obviously attenuated RANKL-induced expression of autophagy- and osteoclastogenesis-related genes, formation of TRAP-positive multinuclear cells, accumulation of LC3, as well as formation of F-actin ring. Our study indicates that p62 may play essential roles in RANKL-induced autophagy and osteoclastogenesis, which may help to develop a novel therapeutic strategy against osteoclastogenesis-related diseases.

SUBMITTER: Li RF

PROVIDER: S-EPMC4244301 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The adaptor protein p62 is involved in RANKL-induced autophagy and osteoclastogenesis.

Li Rui-Fang RF Chen Gang G Ren Jian-Gang JG Zhang Wei W Wu Zhong-Xing ZX Liu Bing B Zhao Yi Y Zhao Yi-Fang YF

The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 20140827 12

Previous studies have implicated autophagy in osteoclast differentiation. The aim of this study was to investigate the potential role of p62, a characterized adaptor protein for autophagy, in RANKL-induced osteoclastogenesis. Real-time quantitative PCR and western blot analyses were used to evaluate the expression levels of autophagy-related markers during RANKL-induced osteoclastogenesis in mouse macrophage-like RAW264.7 cells. Meanwhile, the potential relationship between p62/LC3 localization ...[more]

PMID: 25163928

Similar Datasets

Project description:BackgroundPhosphorylation modification of BCL2 is involved in receptor activator of nuclear factor-κB ligand (RANKL)-induced autophagy of osteoclast precursors (OCPs) and osteoclastogenesis. As an antiapoptotic molecule, the role of BCL2 phosphorylation in osteoclastogenesis is unknown. This study aimed to explore how BCL2 phosphorylation at specific sites regulates osteoclastogenesis.MethodsWe first examined the effects of RANKL on BCL2 phosphorylation at different sites (Ser70 and Ser87) in OCPs. In vivo, transgenic mice overexpressing RANKL (Tg-hRANKL mice) were used to observe the effects of RANKL on phosphorylated BCL2 at different sites in OCPs of trabecular bone. Subsequently, using site-directed mutagenesis, we observed the respective effect of BCL2 mutations at different phosphorylation sites in OCPs on osteoclastogenesis, apoptosis, autophagy and the affinity between BCL2 and Beclin1/BAX under RANKL intervention.ResultsRANKL promoted BCL2 phosphorylation at the Ser70 (S70) site, but not the Ser87 (S87) site, in OCPs. Moreover, Tg-hRANKL mice had stronger BCL2 phosphorylation capacity at S70, not S87, in the OCPs of trabecular bone than wild-type mice in the same nest. Furthermore, BCL2 mutation at S70, not S87, inhibited RANKL-induced osteoclast differentiation and bone resorption activity. In addition, BCL2 mutation at S70 promoted OCP apoptosis, while BCL2 mutation at S87 showed the opposite effect. Remarkably, the BCL2 mutation at S70, not S87, inhibited OCP autophagic activity. Furthermore, BCL2 mutation at S70 enhanced the coimmunoprecipitation of BCL2 and Beclin1, whereas BCL2 mutation at S87 enhanced the coimmunoprecipitation of BCL2 and BAX in OCPs. More importantly, OCP autophagy, osteoclast differentiation and resorption pits inhibited by BCL2 mutation at S70 could be reversed by Beclin1 upregulation with TAT-Beclin1.ConclusionRANKL activates OCP autophagy through BCL2 phosphorylation at S70, thereby promoting osteoclastogenesis, which indicates that the inactivation of BCL2 at S70 in OCPs may be a therapeutic strategy for pathological bone loss.

Dataset Information

The adaptor protein p62 is involved in RANKL-induced autophagy and osteoclastogenesis.

Publications

The adaptor protein p62 is involved in RANKL-induced autophagy and osteoclastogenesis.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets