Rab27a and Rab27b are involved in stimulation-dependent RANKL release from secretory lysosomes in osteoblastic cells

Abstract

The quantity of the receptor activator of NF-kB ligand (RANKL) expressed at the cell surface of osteoblastic cells is an important factor regulating osteoclast activation. Previously, RANKL was found to be localized to secretory lysosomes in osteoblastic cells and to translocate to the cell surface in response to stimulation with RANK-Fc-conjugated beads. However, the in vivo significance of stimulation-dependent RANKL release has not been elucidated. In this study we show that small GTPases Rab27a and Rab27b are involved in the stimulation-dependent RANKL release pathway in osteoblastic cells. Suppression of either Rab27a or Rab27b resulted in a marked reduction in RANKL release after stimulation. Slp4-a, Slp5, and Munc13-4 acted as effector molecules that coordinated Rab27a/b activity in this pathway. Suppression of Rab27a/b or these effector molecules did not inhibit accumulation of RANKL in lysosomal vesicles around the stimulated sites but did inhibit the fusion of these vesicles to the plasma membrane. In osteoblastic cells, suppression of the effector molecules resulted in reduced osteoclastogenic ability. Furthermore, Jinx mice, which lack a functional Munc13-4 gene, exhibited a phenotype characterized by increased bone volume near the tibial metaphysis caused by low bone resorptive activity. In conclusion, stimulation-dependent RANKL release is mediated by Rab27a/b and their effector molecules, and this mechanism may be important for osteoclast activation in vivo. ß

Rab27A Regulates Transport of Cell Surface Receptors Modulating Multinucleation and Lysosome-Related Organelles in Osteoclasts

“…Suppression of Rab27A or RAb27B by RNA interference causes markedly reduced secretion of RANKL in murine osteoblastic cell line ST2 cells41. These results also suggest that Rab27deficiency in osteoblasts results in reduced OCL activation.…”

MiR-124 Attenuates Osteoclastogenic Differentiation of Bone Marrow Monocytes Via Targeting Rab27a

“…In a previous report, the researchers found that Rab27a-deficient osteoclasts showed a markedly impaired bone resorption activity, indicating the important role of Rab27a in maintaining the normal functions of osteoclasts [35,36]. They also showed that depletion of Rab27a results in abnormal phenotypes in osteoclasts.…”

Membrane depolarization regulates intracellular RANKL transport in non-excitable osteoblasts

“…Lysosomal vesicles transport RANKL molecules to the cell membrane [8,9], a process that involves lysosomal molecules such as Rab27 and Vps33; however, the mechanism of RANKL-lv fusion to the cell membrane is unknown. We suggest that membrane depolarization is the first trigger for vesicle fusion in RANKLiT.…”

“…Parathyroid hormone (PTH) and 1α,25-dihydroxyvitamin D 3 (VD 3 ) are osteogenic factors in bone remodeling and also major regulators of osteoclastogenesis [4,5], promoting osteoclast differentiation by increasing RANKL mRNA expression [6,7]. Previous reports have suggested that RANKLiT is lysosome-dependent and regulated by lysosomal molecules, including those of the Rab protein family [8,9]. Although the transport route has been partly described, an outstanding question is how RANKL lysosomal vesicles (RANKL-lvs) fuse to the cell membrane.…”