GABP mediates insulin-increased prolactin gene transcription

Area of Science:

• Molecular Endocrinology
• Gene Regulation
• Signal Transduction

Background:

• The prolactin gene's insulin-response element (IRE) is crucial for insulin-stimulated transcription.
• Ets transcription factors are implicated in regulating gene expression, including the prolactin gene.

Purpose of the Study:

• To investigate the role of Ets-related transcription factors, specifically GABP, in mediating insulin's effect on prolactin gene expression.
• To elucidate the signaling pathway involving MAP kinase in the regulation of GABP activity by insulin.

Main Methods:

• Identification of Ets-related transcription factors in GH cell nuclear extracts using immunoblotting.
• Analysis of GABP binding to the prolactin promoter's IRE using electrophoretic mobility shift assays (EMSAs) and antibody supershifts.
• Assessment of GABP alpha and beta phosphorylation status and MAP kinase activity in response to insulin treatment.
• Functional studies involving GABP expression to assess its impact on insulin-induced prolactin gene expression.

Main Results:

• The Ets-binding site in the prolactin gene's IRE was identified.
• The Ets-related transcription factor GABP (GABP alpha and GABP beta 1) was found to bind the prolactin promoter's IRE.
• Insulin treatment led to a 3-fold increase in GABP alpha phosphorylation, but not GABP beta.
• Mitogen-activated protein (MAP) kinase activity increased 10-fold with insulin, and this activated MAP kinase phosphorylated GABP alpha.
• Overexpression of GABP alpha and GABP beta 1 inhibited insulin-induced prolactin gene expression.

Conclusions:

• GABP acts as a key mediator of insulin-increased transcription of the prolactin gene.
• MAP kinase-dependent phosphorylation of GABP alpha is a significant regulatory mechanism for insulin's effect on prolactin gene expression.