scholarly journals The Role of TRAPγ/SSR3 in Preproinsulin Translocation into the Endoplasmic Reticulum

2021 ◽  
Author(s):  
Xiaoxi Xu ◽  
Yumeng Huang ◽  
Xin Li ◽  
Peter Arvan ◽  
Ming Liu
Keyword(s):  
Endoplasmic Reticulum ◽  
Signal Sequence ◽  
Human Islets ◽  
Insulin Biosynthesis ◽  
Mrna Levels ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Protein Levels ◽  
Proinsulin Biosynthesis ◽  
Extracellular Glucose

In the endoplasmic reticulum (ER), the Translocation-Associated Protein complex (TRAP, also called Signal sequence receptor, SSR) includes four integral membrane proteins TRAPα/SSR1, TRAPβ/SSR2 and TRAPδ/SSR4 with the bulk of their extramembranous portions primarily in the ER lumen, whereas the extramembranous portion of TRAPγ/SSR3 is primarily cytosolic. Individually diminished expression of either TRAPα/SSR1, TRAPβ/SSR2 or TRAPδ/SSR4 mRNA is known in each case to lower TRAPα/SSR1 protein levels leading to impaired proinsulin biosynthesis, whereas forced expression of TRAPα/SSR1 at least partially suppresses the proinsulin biosynthetic defect. Here we report that diminished TRAPγ/SSR3 expression in pancreatic β-cells leaves TRAPα/SSR1 levels unaffected while nevertheless inhibiting co-translational and post-translational translocation of preproinsulin into the ER. Crucially, acute exposure to high glucose leads to a rapid upregulation of both TRAPγ/SSR3 and proinsulin protein without change in the respective mRNA levels — observed in cultured rodent β-cell lines and confirmed in human islets. Strikingly, pancreatic β-cells with suppressed TRAPγ/SSR3 expression are blocked in glucose-dependent upregulation of proinsulin (or insulin) biosynthesis. Most remarkable, overexpression of TRAPγ/SSR3 in control β-cells raises proinsulin levels even without boosting extracellular glucose. The data suggest the possibility that TRAPγ/SSR3 may fulfill a rate-limiting function in preproinsulin translocation across the ER membrane for proinsulin biosynthesis.

The Role of TRAPγ/SSR3 in Preproinsulin Translocation into the Endoplasmic Reticulum

2021 ◽  
Author(s):  
Xiaoxi Xu ◽  
Yumeng Huang ◽  
Xin Li ◽  
Peter Arvan ◽  
Ming Liu
Keyword(s):  
Endoplasmic Reticulum ◽  
Signal Sequence ◽  
Human Islets ◽  
Insulin Biosynthesis ◽  
Mrna Levels ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Protein Levels ◽  
Proinsulin Biosynthesis ◽  
Extracellular Glucose

In the endoplasmic reticulum (ER), the Translocation-Associated Protein complex (TRAP, also called Signal sequence receptor, SSR) includes four integral membrane proteins TRAPα/SSR1, TRAPβ/SSR2 and TRAPδ/SSR4 with the bulk of their extramembranous portions primarily in the ER lumen, whereas the extramembranous portion of TRAPγ/SSR3 is primarily cytosolic. Individually diminished expression of either TRAPα/SSR1, TRAPβ/SSR2 or TRAPδ/SSR4 mRNA is known in each case to lower TRAPα/SSR1 protein levels leading to impaired proinsulin biosynthesis, whereas forced expression of TRAPα/SSR1 at least partially suppresses the proinsulin biosynthetic defect. Here we report that diminished TRAPγ/SSR3 expression in pancreatic β-cells leaves TRAPα/SSR1 levels unaffected while nevertheless inhibiting co-translational and post-translational translocation of preproinsulin into the ER. Crucially, acute exposure to high glucose leads to a rapid upregulation of both TRAPγ/SSR3 and proinsulin protein without change in the respective mRNA levels — observed in cultured rodent β-cell lines and confirmed in human islets. Strikingly, pancreatic β-cells with suppressed TRAPγ/SSR3 expression are blocked in glucose-dependent upregulation of proinsulin (or insulin) biosynthesis. Most remarkable, overexpression of TRAPγ/SSR3 in control β-cells raises proinsulin levels even without boosting extracellular glucose. The data suggest the possibility that TRAPγ/SSR3 may fulfill a rate-limiting function in preproinsulin translocation across the ER membrane for proinsulin biosynthesis.

scholarly journals Signal Transducer and Activator of Transcription 5 Activation Is Sufficient to Drive Transcriptional Induction of Cyclin D2 Gene and Proliferation of Rat Pancreatic β-Cells

2003 ◽  
Vol 17 (5) ◽  
pp. 945-958 ◽  
Author(s):  
Birgitte N. Friedrichsen ◽  
Henrijette E. Richter ◽  
Johnny A. Hansen ◽  
Christopher J. Rhodes ◽  
Jens H. Nielsen ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Fold Increase ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Signal Transducer ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Cyclin D2 ◽  
Protein Levels

Abstract Signal transducer and activator of transcription 5 (STAT5) activation plays a central role in GH- and prolactin-mediated signal transduction in the pancreatic β-cells. In previous experiments we demonstrated that STAT5 activation is necessary for human (h)GH-stimulated proliferation of INS-1 cells and hGH-induced increase of mRNA-levels of the cell cycle regulator cyclin D2. In this study we have further characterized the role of STAT5 in the regulation of cyclin D expression and β-cell proliferation by hGH. Cyclin D2 mRNA and protein levels (but not cyclin D1 and D3) were induced in a time-dependent manner by hGH in INS-1 cells. Inhibition of protein synthesis by coincubation with cycloheximide did not affect the hGH-induced increase of cyclin D2 mRNA levels at 4 h. Expression of a dominant negative STAT5 mutant, STAT5aΔ749, partially inhibited cyclin D2 protein levels. INS-1 cells transiently transfected with a cyclin D2 promoter-reporter construct revealed a 3- to 5-fold increase of transcriptional activity in response to hGH stimulation. Furthermore, coexpression of a constitutive active STAT5 mutant (either CA-STAT5a or CA-STAT5b) was sufficient to drive transactivation of the promoter. CA-STAT5b was stably expressed in INS-1 cells under the control of a doxycycline-inducible promoter. Gel retardation experiments using a probe representing a putative STAT5 binding site in the cyclin D2 promoter revealed binding of the doxycycline-induced CA-STAT5b. Furthermore, induction of CA-STAT5b stimulated transcriptional activation of the cyclin D2 promoter and induced hGH-independent proliferation in these cells. In primary β-cells, adenovirus-mediated expression of CA-STAT5b profoundly stimulated DNA-synthesis (5.3-fold over control) in the absence of hGH. Our studies indicate that STAT5 activation is sufficient to drive proliferation of the β-cells and that cyclin D2 may be a critical target gene for STAT5 in this process.

scholarly journals Upstream stimulatory factor regulates Pdx-1 gene expression in differentiated pancreatic β-cells

1999 ◽  
Vol 341 (2) ◽  
pp. 315-322 ◽  
Author(s):  
Jin QIAN ◽  
Elizabeth N. KAYTOR ◽  
Howard C. TOWLE ◽  
L. Karl OLSON
Keyword(s):  
Gene Expression ◽  
Gene Promoter ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Upstream Stimulatory Factor ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Protein Levels ◽  
E Box ◽  
Stimulatory Factor

The homeobox gene Pdx-1 plays a key role in the development of the pancreas. In the adult, however, expression of the Pdx-1 gene is restricted to pancreatic β-cells and endocrine cells of duodenal epithelium. Recently, the transcription factor, upstream stimulatory factor (USF), has been shown to bind invitro to a mutationally sensitive E-box motif within the 5′-flanking region of the Pdx-1 gene [Sharma, Leonard, Lee, Chapman, Leiter and Montminy (1996) J. Biol. Chem. 271, 2294-2299]. In the present study, we show that USF not only binds to the Pdx-1 gene promoter but also functionally regulates the expression of the Pdx-1 gene in differentiated pancreatic β-cells. Adenovirus-mediated overexpression of a dominant negative form of USF2 decreased binding of endogenous USF to the E-box element by ~ 90%. This reduction in endogenous USF binding led to a greater than 50% decrease in Pdx-1 gene promoter activity, which, in turn, resulted in marked reductions in Pdx-1 mRNA and protein levels. Importantly, the lower Pdx-1 protein levels led to a greater than 50% reduction in Pdx-1 binding activity to the A3 element on the insulin gene promoter, and a significant reduction in insulin mRNA levels. Overall, our results show that USF functionally regulates Pdx-1 gene expression in differentiated pancreatic β-cells and provide the first functional data for a role of USF in the regulation of a normal cellular gene.

Conditional expression of the FTO gene product in rat INS-1 cells reveals its rapid turnover and a role in the profile of glucose-induced insulin secretion

2011 ◽  
Vol 120 (9) ◽  
pp. 403-413 ◽  
Author(s):  
Mark A. Russell ◽  
Noel G. Morgan
Keyword(s):  
Insulin Secretion ◽  
Expression System ◽  
Proteasome Inhibitors ◽  
Inducible Promoter ◽  
Insulin Biosynthesis ◽  
Second Phase ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Peripheral Tissues ◽  
Conditional Expression

Common polymorphisms within the FTO (fat mass and obesity-associated) gene correlate with increased BMI (body mass index) and a rising risk of Type 2 diabetes. FTO is highly expressed in the brain but has also been detected in peripheral tissues, including the endocrine pancreas, although its function there is unclear. The aim of the present study was to investigate the role of FTO protein in pancreatic β-cells using a conditional expression system developed in INS-1 cells. INS-1 cells were stably transfected with FTO–HA (haemagluttinin) incorporated under the control of a tetracycline-inducible promoter. Induction of FTO protein resulted in localization of the tagged protein to the nucleus. The level of FTO–HA protein achieved in transfected cells was tightly regulated, and experiments with selective inhibitors revealed that FTO–HA is rapidly degraded via the ubiquitin/proteasome pathway. The nuclear localization was not altered by proteasome inhibitors, although following treatment with PYR-41, an inhibitor of ubiquitination, some of the protein adopted a perinuclear localization. Unexpectedly, modestly increased expression of FTO–HA selectively enhanced the first phase of insulin secretion when INS-1 monolayers or pseudoislets were stimulated with 20 mM glucose, whereas the second phase remained unchanged. The mechanism responsible for the potentiation of glucose-induced insulin secretion is unclear; however, further experiments revealed that it did not involve an increase in insulin biosynthesis or any changes in STAT3 (signal transducer and activator of transcription 3) expression. Taken together, these results suggest that the FTO protein may play a hitherto unrecognized role in the control of first-phase insulin secretion in pancreatic β-cells.

scholarly journals Mitochondrial–Endoplasmic Reticulum Interplay: A Lifelong On–Off Relationship?

Contact ◽  
2019 ◽  
Vol 2 ◽  
pp. 251525641986122 ◽  
Author(s):  
Corina T. Madreiter-Sokolowski ◽  
Roland M. Malli ◽  
Wolfgang F. Graier
Keyword(s):  
Endoplasmic Reticulum ◽  
Adenosine Triphosphate ◽  
Therapeutic Strategies ◽  
Mitochondrial Activity ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Potential Target ◽  
Pathological Conditions ◽  
Elevated Glucose ◽  
Stimulation Of

This article comments recent publications that highlight an intriguing importance of specific settings in the interaction between the mitochondria and the endoplasmic reticulum to ensure cell-specific functions like the responsiveness to elevated glucose in pancreatic β-cells. Hence, alterations of the mitochondria–endoplasmic reticulum communications under various pathological conditions like aging or cancer often come with enhanced Ca2+ transfer that, in turn, yields stimulation of basal mitochondrial activity to meet the increasing adenosine triphosphate demand of the very cell. Such observations identify mitochondria-associated membranes as potential target for new therapeutic strategies against aging or cancer.

scholarly journals Protein Kinase CK2 Controls CaV2.1-Dependent Calcium Currents and Insulin Release in Pancreatic β-cells

2020 ◽  
Vol 21 (13) ◽  
pp. 4668
Author(s):  
Rebecca Scheuer ◽  
Stephan Ernst Philipp ◽  
Alexander Becker ◽  
Lisa Nalbach ◽  
Emmanuel Ampofo ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Calcium Currents ◽  
Insulin Biosynthesis ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Voltage Dependent ◽  
Regulatory Impact ◽  
Kinase Ck2

The regulation of insulin biosynthesis and secretion in pancreatic β-cells is essential for glucose homeostasis in humans. Previous findings point to the highly conserved, ubiquitously expressed serine/threonine kinase CK2 as having a negative regulatory impact on this regulation. In the cell culture model of rat pancreatic β-cells INS-1, insulin secretion is enhanced after CK2 inhibition. This enhancement is preceded by a rise in the cytosolic Ca2+ concentration. Here, we identified the serine residues S2362 and S2364 of the voltage-dependent calcium channel CaV2.1 as targets of CK2 phosphorylation. Furthermore, co-immunoprecipitation experiments revealed that CaV2.1 binds to CK2 in vitro and in vivo. CaV2.1 knockdown experiments showed that the increase in the intracellular Ca2+ concentration, followed by an enhanced insulin secretion upon CK2 inhibition, is due to a Ca2+ influx through CaV2.1 channels. In summary, our results point to a modulating role of CK2 in the CaV2.1-mediated exocytosis of insulin.

GSTO1 regulates insulin biosynthesis in pancreatic β cells

2020 ◽  
Vol 524 (4) ◽  
pp. 936-942
Author(s):  
Linlin Wang ◽  
Lei Lei ◽  
Tao Xu ◽  
You Wang
Keyword(s):  
Insulin Biosynthesis ◽  
Β Cells ◽  
Pancreatic Β Cells
Sign in / Sign up

Export Citation Format

Share Document