Dataset Information

Gene silencing of phogrin unveils its essential role in glucose-responsive pancreatic beta-cell growth.

ABSTRACT:

Objective

Phogrin and IA-2, autoantigens in insulin-dependent diabetes, have been shown to be involved in insulin secretion in pancreatic beta-cells; however, implications at a molecular level are confusing from experiment to experiment. We analyzed biological functions of phogrin in beta-cells by an RNA interference technique.

Research design and methods

Adenovirus-mediated expression of short hairpin RNA specific for phogrin (shPhogrin) was conducted using cultured beta-cell lines and mouse islets. Both glucose-stimulated insulin secretion and cell proliferation rate were determined in the phogrin-knockdown cells. Furthermore, protein expression was profiled in these cells. To see the binding partner of phogrin in beta-cells, coimmunoprecipitation analysis was carried out.

Results

Adenoviral expression of shPhogrin efficiently decreased its endogenous expression in pancreatic beta-cells. Silencing of phogrin in beta-cells abrogated the glucose-mediated mitogenic effect, which was accompanied by a reduction in the level of insulin receptor substrate 2 (IRS2) protein, without any changes in insulin secretion. Phogrin formed a complex with insulin receptor at the plasma membrane, and their interaction was promoted by high-glucose stimulation that in turn led to stabilization of IRS2 protein. Corroboratively, phogrin knockdown had no additional effect on the proliferation of beta-cell line derived from the insulin receptor-knockout mouse.

Conclusions

Phogrin is involved in beta-cell growth via regulating stability of IRS2 protein by the molecular interaction with insulin receptor. We propose that phogrin and IA-2 function as an essential regulator of autocrine insulin action in pancreatic beta-cells.

SUBMITTER: Torii S

PROVIDER: S-EPMC2646067 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Gene silencing of phogrin unveils its essential role in glucose-responsive pancreatic beta-cell growth.

Torii Seiji S Saito Naoya N Kawano Ayumi A Hou Ni N Ueki Kohjiro K Kulkarni Rohit N RN Takeuchi Toshiyuki T

Diabetes 20081210 3

<h4>Objective</h4>Phogrin and IA-2, autoantigens in insulin-dependent diabetes, have been shown to be involved in insulin secretion in pancreatic beta-cells; however, implications at a molecular level are confusing from experiment to experiment. We analyzed biological functions of phogrin in beta-cells by an RNA interference technique.<h4>Research design and methods</h4>Adenovirus-mediated expression of short hairpin RNA specific for phogrin (shPhogrin) was conducted using cultured beta-cell lin ...[more]

PMID: 19073770

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Project description:Energy sensing is indispensable to balance anabolic and catabolic processes for the maintenance of cell viability. Pancreatic β cells are especially relevant because of their involvement in the coordination of insulin secretion when glucose concentration arises in the local milieu. In this work, we uncover the increased susceptibility of pancreatic β cells to cell death in response to different energy stressors. Upon glucose decline, from 25 to 5 mM, caused by stimulation with either 2-deoxyglucose or metformin, only pancreatic β cells showed an increase in cell death. Very interestingly, when we transfected either mouse insulinoma cell or human embryo kidney cells with a phospho-mutant form of B cell lymphoma 2 associated agonist of cell death at serine 155 (BAD S155D), an increase in the pro-survival factor B cell lymphoma 2 was detected in pancreatic β cells and not in human embryonic kidney cells in the presence of the energetic stressors. This data suggests that the protective capacity of this mutant form is only present in cells that present glucokinase. In contrast, upon hyperactivation of mechanistic target of rapamycin complex 1 signaling by knocking-down tuberous sclerosis complex protein, we observed increased susceptibility to cell death in response to energy stress in both pancreatic and non-pancreatic β cells. Therefore, mechanistic target of rapamycin complex 1 signaling presents a dual effect on cell viability. On the one hand, a chronic inhibition of mechanistic target of rapamycin complex 1 activity in response to the energy status is deleterious for pancreatic β cells, being attenuated by the overexpression of B cell lymphoma 2 associated agonist of cell death S155D. On the other hand, mechanistic target of rapamycin complex 1 hyperactivity provokes a susceptibility to energetic stress-induced cell death. Taken together, these results may open potential implications for the use of glucokinase activators or mechanistic target of rapamycin complex 1 modulators for the maintenance of pancreatic β cells for longer periods of time avoiding its loss in different pathologies such as type 2 diabetes mellitus.

Dataset Information

Gene silencing of phogrin unveils its essential role in glucose-responsive pancreatic beta-cell growth.

Objective

Research design and methods

Results

Conclusions

Publications

Gene silencing of phogrin unveils its essential role in glucose-responsive pancreatic beta-cell growth.

Similar Datasets

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