title: Pharmacological inhibition of lysosomes activates the MTORC1 signaling pathway in chondrocytes in an autophagy-independent manner.
abstract: Mechanistic target of rapamycin (serine/threonine kinase) complex 1 (MTORC1) is a protein-signaling complex at the fulcrum of anabolic and catabolic processes, which acts depending on wide-ranging environmental cues. It is generally accepted that lysosomes facilitate MTORC1 activation by generating an internal pool of amino acids. Amino acids activate MTORC1 by stimulating its translocation to the lysosomal membrane where it forms a super-complex involving the lysosomal-membrane-bound vacuolar-type H(+)-ATPase (v-ATPase) proton pump. This translocation and MTORC1 activation require functional lysosomes. Here we found that, in contrast to this well-accepted concept, in epiphyseal chondrocytes inhibition of lysosomal activity by v-ATPase inhibitors bafilomycin A1 or concanamycin A potently activated MTORC1 signaling. The activity of MTORC1 was visualized by phosphorylated forms of RPS6 (ribosomal protein S6) and EIF4EBP1, 2 well-known downstream targets of MTORC1. Maximal RPS6 phosphorylation was observed at 48-h treatment and reached as high as a 12-fold increase (p < 0.018). This activation of MTORC1 was further confirmed in bone organ culture and promoted potent stimulation of longitudinal growth (p < 0.001). Importantly, the same effect was observed in ATG5 (autophagy-related 5)-deficient bones suggesting a macroautophagy-independent mechanism of MTORC1 inhibition by lysosomes. Thus, our data show that in epiphyseal chondrocytes lysosomes inhibit MTORC1 in a macroautophagy-independent manner and this inhibition likely depends on v-ATPase activity.
authors.affiliation: a Department of Physiology and Pharmacology ; Karolinska Institutet ; Stockholm , Sweden.|b Department of Women's and Children's Health ; Astrid Lindgren Children's Hospital; Karolinska University Hospital ; Stockholm , Sweden.
authors.forename: Phillip T
authors.initials: PT
authors.lastname: Newton
authors.affiliation: a Department of Physiology and Pharmacology ; Karolinska Institutet ; Stockholm , Sweden.
authors.forename: Karuna K
authors.initials: KK
authors.lastname: Vuppalapati
authors.affiliation: a Department of Physiology and Pharmacology ; Karolinska Institutet ; Stockholm , Sweden.
authors.forename: Thibault
authors.initials: T
authors.lastname: Bouderlique
authors.affiliation: a Department of Physiology and Pharmacology ; Karolinska Institutet ; Stockholm , Sweden.|b Department of Women's and Children's Health ; Astrid Lindgren Children's Hospital; Karolinska University Hospital ; Stockholm , Sweden.
authors.forename: Andrei S
authors.initials: AS
authors.lastname: Chagin
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D001846:Bone Development / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006984:Hypertrophy; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D018942:Macrolides / Q000494:pharmacology*; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D046912:Multiprotein Complexes / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051381:Rats; D038601:Ribosomal Protein S6 / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism
chemical_list: C526060:Bhd protein, mouse; D018942:Macrolides; D046912:Multiprotein Complexes; D011518:Proto-Oncogene Proteins; D038601:Ribosomal Protein S6; D025521:Tumor Suppressor Proteins; C448579:ribosomal protein S6, mouse; C040929:bafilomycin A1; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D025262:Vacuolar Proton-Translocating ATPases
keywords: MTOR; MTORC1; autophagy; bafilomycin; bone; chondrocyte; lysosome