PLEKHM1/DEF8/RAB7 complex regulates lysosome positioning and bone homeostasis
Toshifumi Fujiwara, … , Stavros C. Manolagas, Haibo Zhao
Toshifumi Fujiwara, … , Stavros C. Manolagas, Haibo Zhao
Published October 20, 2016
Citation Information: JCI Insight. 2016;1(17):e86330. https://doi.org/10.1172/jci.insight.86330.
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Research Article Bone biology

PLEKHM1/DEF8/RAB7 complex regulates lysosome positioning and bone homeostasis

Abstract

Mutations of the Plekhm1 gene in humans and rats cause osteopetrosis, an inherited bone disease characterized by diminished bone resorption by osteoclasts. PLEKHM1 binds to RAB7 and is critical for lysosome trafficking. However, the molecular mechanisms by which PLEKHM1 regulates lysosomal pathways remain unknown. Here, we generated germline and conditional Plekhm1-deficient mice. These mice displayed no overt abnormalities in major organs, except for an increase in trabecular bone mass. Furthermore, loss of PLEKHM1 abrogated the peripheral distribution of lysosomes and bone resorption in osteoclasts. Mechanistically, we indicated that DEF8 interacts with PLEKHM1 and promotes its binding to RAB7, whereas the binding of FAM98A and NDEL1 with PLEKHM1 connects lysosomes to microtubules. Importantly, suppression of these proteins results in lysosome positioning and bone resorption defects similar to those of Plekhm1-null osteoclasts. Thus, PLHKEM1, DEF8, FAM98A, and NDEL1 constitute a molecular complex that regulates lysosome positioning and secretion through RAB7.

Authors

Toshifumi Fujiwara, Shiqiao Ye, Thiago Castro-Gomes, Caylin G. Winchell, Norma W. Andrews, Daniel E. Voth, Kottayil I. Varughese, Samuel G. Mackintosh, Yunfeng Feng, Nathan Pavlos, Takashi Nakamura, Stavros C. Manolagas, Haibo Zhao

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Figure 1

Generation and characterization of Plekhm1 germline and conditional deletion mice.

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Generation and characterization of Plekhm1 germline and conditional dele...
(A) Strategy to generate a Plekhm1-flox allele. (B) Breeding strategy to generate Plekhm1 wild-type, germline deletion (KO), control (con), and conditional deletion (cKO) mice using Plekhm1-floxed mice and respective Cre-expressing mice. Hprt, hypoxanthine guanine phosphoribosyl transferase; Ctsk, cathepsin K. (C) Quantitative real-time PCR of exon 3 of murine Plekhm1 genomic DNA, normalized to the transferrin receptor 1 locus, using genomic DNA isolated from the indicated soft tissues and tibia (mean ± SD, n = 3). **P < 0.01 versus wild type by Student’s t test. (D) H&E staining of paraffin-embedded tissue sections of wild-type and Plekhm1 germline deletion mice. Representative images of each tissue from 3 different mice/genotype are shown. Original magnification: ×100. (E and F) Scatter dot plot presentations of body weight of 2-month-old male (WT, n = 17; KO, n = 17; con, n = 15; cKO, n = 12) and female mice (WT, n = 11; KO, n = 15; con, n = 14; cKO, n = 14). The mean and SD of each group are overlaid onto each column of dots.