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Identification of 14‐3‐3 Binding Proteins in Mouse Testis by Tandem Affinity Tag Purification
Author(s) -
Puri Pawan,
Snow Alan J,
kline Douglas W,
Vijayaraghavan Srinivasan
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1025.2
Subject(s) - biology , microbiology and biotechnology , germ cell , tandem affinity purification , spermiogenesis , spermatogenesis , gene , phosphorylation , alternative splicing , genetics , messenger rna , biochemistry , nucleus , endocrinology , affinity chromatography , enzyme
The 14‐3‐3 proteins interact with various cellular proteins in a phosphorylation dependent manner. The ability of 14‐3‐3 to bind phosphorylated proteins implicates 14‐3‐3 proteins in a variety of cell signaling pathways. 14‐3‐3 proteins play important regulatory role in mitosis and meiosis. Sperm are produced in testis by a complex process of spermatogenesis. While 14‐3‐3 is present in testis and sperm its role in male gametes is relatively unknown. Tandem affinity tagged (TAP) 14‐3‐3 was expressed in transgenic mice driven by an ubiquitin promoter (Angard et al 2006). TAP‐tagged 14‐3‐3 was purified from testis of the transgenic mice. A total of 150 proteins were found to be associated with 14‐3‐3. These include proteins involved in cell signaling, transcription, translation, splicing, metabolism and protein trafficking. Some of the testis 14‐3‐3 binding proteins were protein were: protein phosphatase1γ2, GSK‐3, testis specific gene 2, SH3 domain GRB2 like B1, gravin, WLK1, Y box protein 2, phosphatidyl ethanolamine binding protein, and piwi homolog 1. PP1γ2 and GSK‐3 binding with 14‐3‐3 was further confirmed by GST pull down assays. Mice lacking PP1 gene are infertile due to impaired spermiogenesis. Efforts are underway to characterize the 14‐3‐3 /PP1γ2 complex to elucidate its function in spermatogenesis. Our results suggest 14‐3‐3 may have important regulatory role in male germ cell maturation in the testis.