The antiapoptotic protein Bcl-xL negatively regulates the bone-resorbing activity of osteoclasts in mice
Mitsuyasu Iwasawa, Tsuyoshi Miyazaki, Yuichi Nagase, Toru Akiyama, Yuho Kadono, Masaki Nakamura, Yasushi Oshima, Tetsuro Yasui, Takumi Matsumoto, Takashi Nakamura, Shigeaki Kato, Lothar Hennighausen, Kozo Nakamura, Sakae Tanaka
Mitsuyasu Iwasawa, Tsuyoshi Miyazaki, Yuichi Nagase, Toru Akiyama, Yuho Kadono, Masaki Nakamura, Yasushi Oshima, Tetsuro Yasui, Takumi Matsumoto, Takashi Nakamura, Shigeaki Kato, Lothar Hennighausen, Kozo Nakamura, Sakae Tanaka
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Research Article Bone biology

The antiapoptotic protein Bcl-xL negatively regulates the bone-resorbing activity of osteoclasts in mice

Abstract

The B cell lymphoma 2 (Bcl-2) family member Bcl-xL has a well-characterized antiapoptotic function in lymphoid cells. However, its functions in other cells — including osteoclasts, which are of hematopoietic origin — and other cellular processes remain unknown. Here we report an unexpected function of Bcl-xL in attenuating the bone-resorbing activity of osteoclasts in mice. To investigate the role of Bcl-xL in osteoclasts, we generated mice with osteoclast-specific conditional deletion of Bcl-x (referred to herein as Bcl-x cKO mice) by mating Bcl-xfl/fl mice with mice in which the gene encoding the Cre recombinase has been knocked into the cathepsin K locus and specifically expressed in mature osteoclasts. Although the Bcl-x cKO mice grew normally with no apparent morphological abnormalities, they developed substantial osteopenia at 1 year of age, which was caused by increased bone resorption. Bcl-x deficiency increased the bone-resorbing activity of osteoclasts despite their high susceptibility to apoptosis, whereas Bcl-xL overexpression produced the opposite effect. In addition, Bcl-x cKO osteoclasts displayed increased c-Src activity, which was linked to increased levels of vitronectin and fibronectin expression. These results suggest that Bcl-xL attenuates osteoclastic bone-resorbing activity through the decreased production of ECM proteins, such as vitronectin and fibronectin, and thus provide evidence for what we believe to be a novel cellular function of Bcl-xL.

Authors

Mitsuyasu Iwasawa, Tsuyoshi Miyazaki, Yuichi Nagase, Toru Akiyama, Yuho Kadono, Masaki Nakamura, Yasushi Oshima, Tetsuro Yasui, Takumi Matsumoto, Takashi Nakamura, Shigeaki Kato, Lothar Hennighausen, Kozo Nakamura, Sakae Tanaka

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Figure 5

Ras-Mek-Erk pathways are upstream of Bcl-xL in osteoclasts.

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Ras-Mek-Erk pathways are upstream of Bcl-xL in osteoclasts. (A) Bcl-xfl/...
(A) Bcl-xfl/fl osteoclasts were infected with AxGFP, AxCre, AxBcl-xL, AxMekCA, or AxRasDN. Bcl-xL overexpression by AxBcl-xL infection suppressed, and knockout of Bcl-x gene by AxCre infection increased, Erk activity, as determined by the amount of phospho-Erk in Bcl-xfl/fl osteoclasts. In contrast, AxMekCA infection increased, and AxRasDN infection decreased, Bcl-xL expression. Relative intensity of the bands on each gel, measured by densitometry, is shown above each lane. (B) Bcl-xfl/fl osteoclasts were infected with AxGFP, AxGFP plus AxRasDN, AxBcl-xL, or AxBcl-xL plus AxRasDN. Reduced osteoclast survival by RasDN overexpression was completely rescued by Bcl-xL overexpression. *P < 0.01 versus AxGFP-infected cells. (C) Bcl-xfl/fl osteoclasts were infected with AxGFP or AxBcl-xL, and then treated with the indicated concentrations of MEK inhibitor PD98059. PD98059 treatment dose-dependently suppressed the survival of osteoclasts, which was completely rescued by Bcl-xL overexpression. *P < 0.01 versus untreated osteoclasts. (D) Bcl-xfl/fl osteoclasts were infected with AxGFP or AxCre together with AxMekCA. Prosurvival effect of MekCA overexpression was partially suppressed by Bcl-x deletion. *P < 0.01, **P < 0.05 versus AxGFP-AxMekCA–infected cells. All results are mean ± SD of 6 cultures.