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Glucose- and hypoxia-regulated insulin gene expression
Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
2002 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Although extensively studied, there are still many unanswered questions regarding the exact regulation of insulin gene expression. This is important to further investigate since it will hopefully help us understand the pathophysiology of some types of diabetes. The insulin mRNA has a long half-life and it is therefore likely to be regulated through specific mechanisms. The aim of the present work was to evaluate the importance of the 3´untranslated region (UTR) of the insulin mRNA in the regulation of insulin mRNA stability. The number of insulin mRNA molecules/rat β-cell was first determined to be 20 000 - 100 000, depending on the glucose concentration of the culture medium. However, the insulin mRNA level was neither increased by glucose nor insulin when stimulated for only one hour. Glucose withdrawal for 16 h caused only a small decrease in insulin mRNA levels. This decrease was of the same magnitude in the presence of a transcriptional inhibitor, indicating that the insulin mRNA stability significantly contributes to the regulation of insulin gene expression. To study the mechanisms by which glucose regulates insulin mRNA stability, we probed for protein binding to the rat and human 3´UTR insulin mRNA (ins-PRS) using the electrophoresis mobility shift assay. Specific binding was observed and the protein was identified as the polypyrimidine tract-binding protein (PTB). Introduction of the ins-PRS in to the 3´UTR of a reporter gene mRNA did not significantly affect the mRNA stability. However, mutation of the ins-PRS and disruption of PTB binding resulted in a marked decrease in expression of the reporter gene mRNA. The binding activity was increased in islets in response to glucose, hypoxia, sphingomyelinase (an inducer of endogenous ceramide) and in vitro by the addition of the reducing agent dithiothreitol (DTT). The hypoxia-induced binding activity was counteracted by exogenous administration of hydrogen peroxide, while the glucose-induced PTB binding activity was inhibited by rapamycin, which is an inhibitor of mTOR and p70s6 kinase. The insulin mRNA levels were increased in response to hypoxia, equal to the increase in response to glucose. In conclusion, PTB binding to the ins-PRS is an important, but not sufficient, event in regulation of insulin mRNA stability. Binding activity is increased in response to glucose and hypoxia, possibly mediated through pathways involving the redox status and/or reactive oxygen species (ROS), ceramide and mTOR/p70s6k.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2002. , 62 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1179
Keyword [en]
Cell biology, islet, beta-cell, insulin, mRNA stability, PTB, glucose, hypoxia, mTOR, p70s6k, ceramide, redox, ROS
Keyword [sv]
Cellbiologi
National Category
Cell and Molecular Biology
Research subject
Medical Cell Biology
Identifiers
URN: urn:nbn:se:uu:diva-2546ISBN: 91-554-5393-7 (print)OAI: oai:DiVA.org:uu-2546DiVA: diva2:161879
Public defence
2002-10-19, lecture hall B21, Biomedical Centre (BMC), Uppsala, 13:15
Opponent
Available from: 2002-09-25 Created: 2002-09-25Bibliographically approved
List of papers
1. Control of insulin mRNA stability in rat pancreatic islets: Regulatory role of a 3'-untranslated region pyrimidine-rich sequence
Open this publication in new window or tab >>Control of insulin mRNA stability in rat pancreatic islets: Regulatory role of a 3'-untranslated region pyrimidine-rich sequence
2002 In: J Biol Chem, Vol. 277, no 2, 1099-1106 p.Article in journal (Refereed) Published
Abstract
Identifiers
urn:nbn:se:uu:diva-89940 (URN)
Available from: 2002-09-25 Created: 2002-09-25 Last updated: 2017-01-04Bibliographically approved
2. Hypoxia may increase rat insulin mRNA levels by promoting binding to the polypyrimidine tract-binding protein (PTB) to the pyrimidine-rich insulin mRNA 3´-untranslated region
Open this publication in new window or tab >>Hypoxia may increase rat insulin mRNA levels by promoting binding to the polypyrimidine tract-binding protein (PTB) to the pyrimidine-rich insulin mRNA 3´-untranslated region
2002 In: Mol Med.Article in journal (Refereed) Published
Identifiers
urn:nbn:se:uu:diva-89941 (URN)
Available from: 2002-09-25 Created: 2002-09-25Bibliographically approved
3. Glucose-induced polypyrimidine-tract-binding protein (PTB) to the 3´-untranslated region of the insulin mRNA is inhibited by rapamycin
Open this publication in new window or tab >>Glucose-induced polypyrimidine-tract-binding protein (PTB) to the 3´-untranslated region of the insulin mRNA is inhibited by rapamycin
Manuscript (Other academic)
Identifiers
urn:nbn:se:uu:diva-89942 (URN)
Available from: 2002-09-25 Created: 2002-09-25 Last updated: 2010-01-13Bibliographically approved

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