Polypyrimidine tract binding protein 2 stabilizes phosphoglycerate kinase 2 mRNA in murine male germ cells by binding to its 3'UTR

Biology of Reproduction
Mingang Xu, Norman B Hecht

Abstract

The mRNA that encodes the testis-specific protein phosphoglycerate kinase (PGK2) is a long-lived mRNA that is transcribed in meiotic and postmeiotic male germ cells. Pgk2 mRNA is present in germ cells for up to 2 wk before its protein product is detected. Using affinity chromatography with the 3'-UTR of the Pgk2 mRNA, several proteins, including the RNA-binding protein, polypyrimidine tract binding protein 2 (PTBP2), were identified in mouse testis extracts. Coimmunoprecipitation experiments confirmed that PTBP2 binds to Pgk2 mRNA in the testis and RNA gel shifts demonstrated that PTBP2, but not PTBP1, binds to a specific region of the Pgk2 3'-UTR. Recombinant PTBP2 increased the stability of reporter constructs that contained the 3'-UTR Pgk2 sequence element in both testis extracts and transfected HeLa cells. We propose that PTBP2 is a trans-acting factor that helps to stabilize Pgk2 mRNA in male mouse germ cells.

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Methods Mentioned

BETA
in vitro transcription
PCR
Gel Shift
Affinity purification
Immunoprecipitation
Assay
immunoprecipitation assay
Protein Assay
transfection
electrophoresis

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