Dataset Information

Loss of mTORC1 signalling impairs ?-cell homeostasis and insulin processing.

ABSTRACT: Deregulation of mTOR complex 1 (mTORC1) signalling increases the risk for metabolic diseases, including type 2 diabetes. Here we show that ?-cell-specific loss of mTORC1 causes diabetes and ?-cell failure due to defects in proliferation, autophagy, apoptosis and insulin secretion by using mice with conditional (?raKO) and inducible (MIP-?raKO^f/f) raptor deletion. Through genetic reconstitution of mTORC1 downstream targets, we identify mTORC1/S6K pathway as the mechanism by which mTORC1 regulates ?-cell apoptosis, size and autophagy, whereas mTORC1/4E-BP2-eIF4E pathway regulates ?-cell proliferation. Restoration of both pathways partially recovers ?-cell mass and hyperglycaemia. This study also demonstrates a central role of mTORC1 in controlling insulin processing by regulating cap-dependent translation of carboxypeptidase E in a 4EBP2/eIF4E-dependent manner. Rapamycin treatment decreases CPE expression and insulin secretion in mice and human islets. We suggest an important role of mTORC1 in ?-cells and identify downstream pathways driving ?-cell mass, function and insulin processing.

SUBMITTER: Blandino-Rosano M

PROVIDER: S-EPMC5510183 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Loss of mTORC1 signalling impairs β-cell homeostasis and insulin processing.

Blandino-Rosano Manuel M Barbaresso Rebecca R Jimenez-Palomares Margarita M Bozadjieva Nadejda N Werneck-de-Castro Joao Pedro JP Hatanaka Masayuki M Mirmira Raghavendra G RG Sonenberg Nahum N Liu Ming M Rüegg Markus A MA Hall Michael N MN Bernal-Mizrachi Ernesto E

Nature communications 20170712

Deregulation of mTOR complex 1 (mTORC1) signalling increases the risk for metabolic diseases, including type 2 diabetes. Here we show that β-cell-specific loss of mTORC1 causes diabetes and β-cell failure due to defects in proliferation, autophagy, apoptosis and insulin secretion by using mice with conditional (βraKO) and inducible (MIP-βraKO<sup>f/f</sup>) raptor deletion. Through genetic reconstitution of mTORC1 downstream targets, we identify mTORC1/S6K pathway as the mechanism by which mTORC ...[more]

PMID: 28699639

Similar Datasets

Project description:BackgroundCaveolins are the principal structural components of plasma membrane caveolae. Dominant pathogenic mutations in the muscle-specific caveolin-3 (Cav3) gene isoform, such as the limb girdle muscular dystrophy type 1C (LGMD-1C) P104L mutation, result in dramatic loss of the Cav3 protein and pathophysiological muscle weakness/wasting. We hypothesize that such muscle degeneration may be linked to disturbances in signalling events that impact protein turnover. Herein, we report studies assessing the effects of Cav3 deficiency on mammalian or mechanistic target of rapamycin complex 1 (mTORC1) signalling in skeletal muscle cells.MethodsL6 myoblasts were stably transfected with Cav3P104L or expression of native Cav3 was abolished by CRISPR/Cas9 genome editing (Cav3 knockout [Cav3KO]) prior to performing subcellular fractionation and immunoblotting, analysis of real-time mitochondrial respiration or fixed cell immunocytochemistry. Skeletal muscle from wild-type and Cav3-/- mice was processed for immunoblot analysis of downstream mTORC1 substrate phosphorylation.ResultsCav3 was detected in lysosomal-enriched membranes isolated from L6 myoblasts and observed by confocal microscopy to co-localize with lysosomal-specific markers. Cav3P104L expression, which results in significant (~95%) loss of native Cav3, or CRISPR/Cas9-mediated Cav3KO, reduced amino acid-dependent mTORC1 activation. The decline in mTORC1-directed signalling was detected by immunoblot analysis of L6 muscle cells and gastrocnemius Cav3-/- mouse muscle as judged by reduced phosphorylation of mTORC1 substrates that play key roles in the initiation of protein synthesis (4EBP1S65 and S6K1T389 ). S6K1T389 and 4EBP1S65 phosphorylation reduced by over 75% and 80% in Cav3KO muscle cells and by over 90% and 30% in Cav3-/- mouse skeletal muscle, respectively. The reduction in protein synthetic capacity in L6 muscle cells was confirmed by analysis of puromycylated peptides using the SUnSET assay. Cav3 loss was also associated with a 26% increase in lysosomal cholesterol, and pharmacological manipulation of lysosomal cholesterol was effective in replicating the reduction in mTORC1 activity observed in Cav3KO cells. Notably, re-expression of Cav3 in Cav3KO myoblasts normalized lysosomal cholesterol content, which coincided with a recovery in protein translation and an associated increase in mTORC1-directed phosphorylation of downstream targets.ConclusionsOur findings indicate that Cav3 can localize on lysosomal membranes and is a novel regulator of mTORC1 signalling in muscle. Cav3 deficiency associated with the Cav3P104L mutation impairs mTORC1 activation and protein synthetic capacity in skeletal muscle cells, which may be linked to disturbances in lysosomal cholesterol trafficking and contribute to the pathology of LGMD-1C.

Dataset Information

Loss of mTORC1 signalling impairs ?-cell homeostasis and insulin processing.

Publications

Loss of mTORC1 signalling impairs β-cell homeostasis and insulin processing.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets