Vps33a mediates RANKL storage in secretory lysosomes in osteoblastic cells.
Previous studies have indicated that the amount of RANKL expressed on the cell surface of osteoblasts or bone marrow stromal cells (BMSCs) is considered an important factor determining the extent of osteoclast activation. However, subcellular trafficking of RANKL and its regulatory mechanisms in osteoblastic cells is still unclear. In this study, we showed that RANKL is predominantly localized in lysosomal organelles, but little is found on the cell surface of osteoblastic cells. We also showed that RANKL is relocated to the plasma membrane in response to stimulation with RANK-Fc-coated beads, indicating that the lysosomal organelles where RANKL is localized function as secretory lysosomes. In addition, using a protein pull-down method, we identified vacuolar protein sorting (Vps)33a as interacting with the cytoplasmic tail of RANKL. Furthermore, knockdown of Vps33a expression reduced the lysosomal storage of RANKL and caused the accumulation of newly synthesized RANKL in the Golgi apparatus, indicating that Vps33a is involved in transporting RANKL from the Golgi apparatus to secretory lysosomes. We also showed that suppression of Vps33a affects the cell surface expression level of RANKL and disrupts the regulated behavior of RANKL. These results suggest that RANKL storage in secretory lysosomes is important to control osteoclast activation and to maintain bone homeostasis.
Department of Pharmacy, The University of Tokyo Hospital, Faculty of Medicine, The University of Tokyo, Tokyo, Japan.
Yoshiaki
Y
Kariya
Masashi
M
Honma
Shigeki
S
Aoki
Atsushi
A
Chiba
Hiroshi
H
Suzuki
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006056:Golgi Apparatus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D010006:Osteoblasts / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D053245:RANK Ligand / Q000737:chemistry / Q000378:metabolism*; D055571:Secretory Pathway; D022142:Secretory Vesicles / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000378:metabolism*
D053245:RANK Ligand; D033921:Vesicular Transport Proteins; C472129:Vps33a protein, mouse