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Regulation of Insulin Receptor Trafficking by Bardet Biedl Syndrome Proteins.

Starks RD, Beyer AM, Guo DF, Boland L, Zhang Q, Sheffield VC, Rahmouni K - PLoS Genet. (2015)

Bottom Line: We demonstrate that the IR interacts physically with BBS proteins, and reducing the expression of BBS proteins perturbs IR expression in the cell surface.Our data identify defects in trafficking and localization of the IR as a novel mechanism accounting for the insulin resistance commonly associated with human BBS.This is supported by the reduced surface expression of the IR in fibroblasts derived from patients bearing the M390R mutation in the BBS1 gene.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Iowa College of Medicine, Iowa City, Iowa, United States of America.

ABSTRACT
Insulin and its receptor are critical for the regulation of metabolic functions, but the mechanisms underlying insulin receptor (IR) trafficking to the plasma membrane are not well understood. Here, we show that Bardet Biedl Syndrome (BBS) proteins are necessary for IR localization to the cell surface. We demonstrate that the IR interacts physically with BBS proteins, and reducing the expression of BBS proteins perturbs IR expression in the cell surface. We show the consequence of disrupting BBS proteins for whole body insulin action and glucose metabolism using mice lacking different BBS genes. These findings demonstrate the importance of BBS proteins in underlying IR cell surface expression. Our data identify defects in trafficking and localization of the IR as a novel mechanism accounting for the insulin resistance commonly associated with human BBS. This is supported by the reduced surface expression of the IR in fibroblasts derived from patients bearing the M390R mutation in the BBS1 gene.

No MeSH data available.


Related in: MedlinePlus

Physical interaction between BBS proteins and IR.A) Sedimentation analysis showing the β subunit of the IR in complex with the BBSome (corresponding to fraction 9). B-C) Reciprocal co-immunoprecipitation with GFP-tagged BBS17 and the β subunit of the IR in protein lysates from HEK293T cells. The interaction between the IRβ subunit and BBS17 can be detected whether IR antibody is used for immunoprecipitation (A) or GFP-BBS17 is used to precipitate IR (Ctrl line in C). Silencing the Bbs1 gene with shRNA disrupt the interaction between BBS17 and IRβ (right line in C).
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pgen.1005311.g001: Physical interaction between BBS proteins and IR.A) Sedimentation analysis showing the β subunit of the IR in complex with the BBSome (corresponding to fraction 9). B-C) Reciprocal co-immunoprecipitation with GFP-tagged BBS17 and the β subunit of the IR in protein lysates from HEK293T cells. The interaction between the IRβ subunit and BBS17 can be detected whether IR antibody is used for immunoprecipitation (A) or GFP-BBS17 is used to precipitate IR (Ctrl line in C). Silencing the Bbs1 gene with shRNA disrupt the interaction between BBS17 and IRβ (right line in C).

Mentions: To investigate the possibility that BBS proteins are involved in IR trafficking, we first examined whether BBS proteins interact with the IR in a sucrose gradient fractionation experiment. Interestingly, a portion of the β subunit of the IR was identified in the same pool of proteins that contained the BBSome complex (Fig 1A). This fraction also contained other known partners of the BBSome such as Ptc1 and Smo proteins [20]. Next, we performed a pairwise reciprocal immunoprecipitation assay to investigate whether BBS proteins interact directly with the IR. Among all the BBS proteins tested, a physical interaction between the IR and BBS17 was identified based on the ability of the endogenous IR and GFP- or Flag-BBS17 to pull down each other in cells (Figs 1B and S1A). Similar results were obtained with the endogenous proteins (IR and BBS17) using mouse tissue (S1B Fig). Of note, shRNA-mediated reduction in the expression of BBS1 (S1C Fig), a key component of the BBSome, disrupted the interaction between BBS17 and the IR (Fig 1C). Together, these data indicate a direct interaction between BBS proteins and IR.


Regulation of Insulin Receptor Trafficking by Bardet Biedl Syndrome Proteins.

Starks RD, Beyer AM, Guo DF, Boland L, Zhang Q, Sheffield VC, Rahmouni K - PLoS Genet. (2015)

Physical interaction between BBS proteins and IR.A) Sedimentation analysis showing the β subunit of the IR in complex with the BBSome (corresponding to fraction 9). B-C) Reciprocal co-immunoprecipitation with GFP-tagged BBS17 and the β subunit of the IR in protein lysates from HEK293T cells. The interaction between the IRβ subunit and BBS17 can be detected whether IR antibody is used for immunoprecipitation (A) or GFP-BBS17 is used to precipitate IR (Ctrl line in C). Silencing the Bbs1 gene with shRNA disrupt the interaction between BBS17 and IRβ (right line in C).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4478011&req=5

pgen.1005311.g001: Physical interaction between BBS proteins and IR.A) Sedimentation analysis showing the β subunit of the IR in complex with the BBSome (corresponding to fraction 9). B-C) Reciprocal co-immunoprecipitation with GFP-tagged BBS17 and the β subunit of the IR in protein lysates from HEK293T cells. The interaction between the IRβ subunit and BBS17 can be detected whether IR antibody is used for immunoprecipitation (A) or GFP-BBS17 is used to precipitate IR (Ctrl line in C). Silencing the Bbs1 gene with shRNA disrupt the interaction between BBS17 and IRβ (right line in C).
Mentions: To investigate the possibility that BBS proteins are involved in IR trafficking, we first examined whether BBS proteins interact with the IR in a sucrose gradient fractionation experiment. Interestingly, a portion of the β subunit of the IR was identified in the same pool of proteins that contained the BBSome complex (Fig 1A). This fraction also contained other known partners of the BBSome such as Ptc1 and Smo proteins [20]. Next, we performed a pairwise reciprocal immunoprecipitation assay to investigate whether BBS proteins interact directly with the IR. Among all the BBS proteins tested, a physical interaction between the IR and BBS17 was identified based on the ability of the endogenous IR and GFP- or Flag-BBS17 to pull down each other in cells (Figs 1B and S1A). Similar results were obtained with the endogenous proteins (IR and BBS17) using mouse tissue (S1B Fig). Of note, shRNA-mediated reduction in the expression of BBS1 (S1C Fig), a key component of the BBSome, disrupted the interaction between BBS17 and the IR (Fig 1C). Together, these data indicate a direct interaction between BBS proteins and IR.

Bottom Line: We demonstrate that the IR interacts physically with BBS proteins, and reducing the expression of BBS proteins perturbs IR expression in the cell surface.Our data identify defects in trafficking and localization of the IR as a novel mechanism accounting for the insulin resistance commonly associated with human BBS.This is supported by the reduced surface expression of the IR in fibroblasts derived from patients bearing the M390R mutation in the BBS1 gene.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Iowa College of Medicine, Iowa City, Iowa, United States of America.

ABSTRACT
Insulin and its receptor are critical for the regulation of metabolic functions, but the mechanisms underlying insulin receptor (IR) trafficking to the plasma membrane are not well understood. Here, we show that Bardet Biedl Syndrome (BBS) proteins are necessary for IR localization to the cell surface. We demonstrate that the IR interacts physically with BBS proteins, and reducing the expression of BBS proteins perturbs IR expression in the cell surface. We show the consequence of disrupting BBS proteins for whole body insulin action and glucose metabolism using mice lacking different BBS genes. These findings demonstrate the importance of BBS proteins in underlying IR cell surface expression. Our data identify defects in trafficking and localization of the IR as a novel mechanism accounting for the insulin resistance commonly associated with human BBS. This is supported by the reduced surface expression of the IR in fibroblasts derived from patients bearing the M390R mutation in the BBS1 gene.

No MeSH data available.


Related in: MedlinePlus