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Characterization of the human skeletal muscle glycogen synthase gene (GYS1) promoter.

Fredriksson, Jenny LU ; Ridderstråle, Martin LU ; Groop, Leif LU and Orho-Melander, Marju LU (2004) In European Journal of Clinical Investigation 34(2). p.113-121
Abstract
Background Impaired activation of the human skeletal muscle glycogen synthase by insulin is typical for type 2 diabetic patients. Regulation of glycogen synthase occurs mainly by phosphorylation/dephoshorylation but little is known whether there also is transcriptional regulation. Therefore we studied transcriptional regulation of the human skeletal muscle glycogen synthase gene (GYS1) and evaluated the effects of insulin and forskolin on the promoter activity.



Methods Seven promoter fragments were expressed in C2C12 myoblasts and myotubes and in HEK293 cells, and the luciferase assay was used to determine transcriptional activity.



Results The highest luciferase activity, 350-fold of the promoterless... (More)
Background Impaired activation of the human skeletal muscle glycogen synthase by insulin is typical for type 2 diabetic patients. Regulation of glycogen synthase occurs mainly by phosphorylation/dephoshorylation but little is known whether there also is transcriptional regulation. Therefore we studied transcriptional regulation of the human skeletal muscle glycogen synthase gene (GYS1) and evaluated the effects of insulin and forskolin on the promoter activity.



Methods Seven promoter fragments were expressed in C2C12 myoblasts and myotubes and in HEK293 cells, and the luciferase assay was used to determine transcriptional activity.



Results The highest luciferase activity, 350-fold of the promoterless vector, was obtained with nucleotides -692 to +59 in myotubes (P < 0·001), while the nucleotides -250 to +59 provided the highest, 45-fold, activity in the HEK293 cells (P < 0·001). Longer promoter constructs (nucleotides -971, -1707 and -2158 to +59, respectively) had low promoter activity in both cell types. Forskolin treatment for 24 h resulted in approximately 30% decreased promoter activity in myotubes (P < 0·05). Insulin treatment for 0·5-3 h did not increase GYS1 promoter activity; instead the activity was slightly but significantly decreased after 24 h in myotubes (P < 0·005).



Conclusions From our results we conclude that basal GYS1 promoter activity is obtained from the first 250 nucleotides of the promoter, while the nucleotides -692 to -544 seem to be responsible for muscle-specific expression, and nucleotides -971 to -692 for negative regulation. In myotubes, the GYS1 promoter was sensitive to negative regulation by forskolin, whereas insulin did not increase GYS1 transcription. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
European Journal of Clinical Investigation
volume
34
issue
2
pages
113 - 121
publisher
Wiley-Blackwell
external identifiers
  • wos:000188811400006
  • pmid:14764074
  • scopus:1342331364
ISSN
0014-2972
DOI
10.1111/j.1365-2362.2004.01299.x
language
English
LU publication?
yes
id
d347b378-a1e6-4576-9640-611d5ac8a995 (old id 120706)
date added to LUP
2007-07-19 12:10:36
date last changed
2017-01-22 03:35:02
@article{d347b378-a1e6-4576-9640-611d5ac8a995,
  abstract     = {Background Impaired activation of the human skeletal muscle glycogen synthase by insulin is typical for type 2 diabetic patients. Regulation of glycogen synthase occurs mainly by phosphorylation/dephoshorylation but little is known whether there also is transcriptional regulation. Therefore we studied transcriptional regulation of the human skeletal muscle glycogen synthase gene (GYS1) and evaluated the effects of insulin and forskolin on the promoter activity.<br/><br>
<br/><br>
Methods Seven promoter fragments were expressed in C2C12 myoblasts and myotubes and in HEK293 cells, and the luciferase assay was used to determine transcriptional activity.<br/><br>
<br/><br>
Results The highest luciferase activity, 350-fold of the promoterless vector, was obtained with nucleotides -692 to +59 in myotubes (P &lt; 0·001), while the nucleotides -250 to +59 provided the highest, 45-fold, activity in the HEK293 cells (P &lt; 0·001). Longer promoter constructs (nucleotides -971, -1707 and -2158 to +59, respectively) had low promoter activity in both cell types. Forskolin treatment for 24 h resulted in approximately 30% decreased promoter activity in myotubes (P &lt; 0·05). Insulin treatment for 0·5-3 h did not increase GYS1 promoter activity; instead the activity was slightly but significantly decreased after 24 h in myotubes (P &lt; 0·005).<br/><br>
<br/><br>
Conclusions From our results we conclude that basal GYS1 promoter activity is obtained from the first 250 nucleotides of the promoter, while the nucleotides -692 to -544 seem to be responsible for muscle-specific expression, and nucleotides -971 to -692 for negative regulation. In myotubes, the GYS1 promoter was sensitive to negative regulation by forskolin, whereas insulin did not increase GYS1 transcription.},
  author       = {Fredriksson, Jenny and Ridderstråle, Martin and Groop, Leif and Orho-Melander, Marju},
  issn         = {0014-2972},
  language     = {eng},
  number       = {2},
  pages        = {113--121},
  publisher    = {Wiley-Blackwell},
  series       = {European Journal of Clinical Investigation},
  title        = {Characterization of the human skeletal muscle glycogen synthase gene (GYS1) promoter.},
  url          = {http://dx.doi.org/10.1111/j.1365-2362.2004.01299.x},
  volume       = {34},
  year         = {2004},
}