Signal transducer and activator of transcription 5 activation is sufficient to drive transcriptional induction of cyclin D2 gene and proliferation of rat pancreatic beta-cells

TY - JOUR

T1 - Signal transducer and activator of transcription 5 activation is sufficient to drive transcriptional induction of cyclin D2 gene and proliferation of rat pancreatic beta-cells

AU - Friedrichsen, Birgitte N

AU - Richter, Henrijette E

AU - Hansen, Johnny A

AU - Rhodes, Christopher J

AU - Nielsen, Jens Høiriis

AU - Billestrup, Nils

AU - Møldrup, Annette

PY - 2003/5

Y1 - 2003/5

N2 - Signal transducer and activator of transcription 5 (STAT5) activation plays a central role in GH- and prolactin-mediated signal transduction in the pancreatic beta-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 beta-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, STAT5aDelta749, 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 beta-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 beta-cells and that cyclin D2 may be a critical target gene for STAT5 in this process.

AB - Signal transducer and activator of transcription 5 (STAT5) activation plays a central role in GH- and prolactin-mediated signal transduction in the pancreatic beta-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 beta-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, STAT5aDelta749, 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 beta-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 beta-cells and that cyclin D2 may be a critical target gene for STAT5 in this process.

KW - Animals

KW - Binding Sites

KW - Cell Division

KW - Cells, Cultured

KW - Cyclin D2

KW - Cyclins

KW - DNA-Binding Proteins

KW - Doxycycline

KW - Gene Expression Regulation

KW - Human Growth Hormone

KW - Islets of Langerhans

KW - Mice

KW - Milk Proteins

KW - Mutation

KW - Promoter Regions, Genetic

KW - Rats

KW - STAT5 Transcription Factor

KW - Trans-Activators

KW - Transcription, Genetic

KW - Transcriptional Activation

U2 - 10.1210/me.2002-0356

DO - 10.1210/me.2002-0356

M3 - Journal article

C2 - 12586844

SN - 0888-8809

VL - 17

SP - 945

EP - 958

JO - Molecular endocrinology (Baltimore, Md.)

JF - Molecular endocrinology (Baltimore, Md.)

IS - 5

ER -