Identification and Characterization of a Novel Protein from Sertoli Cells, PASS1, That Associates with Mammalian Small Stress Protein hsp27

• Published in: The Journal of biological chemistry 2000-06, Vol.275 (25), p.18724-18731
• Main Authors: Liu, Chenghua, Gilmont, Robert R., Benndorf, Rainer, Welsh, Michael J.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Caenorhabditis elegans Proteins; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Line; Cloning, Molecular; DNA, Complementary; Heat-Shock Proteins; Heat-Shock Response - genetics; HSP27 Heat-Shock Proteins; Intracellular Signaling Peptides and Proteins; Male; Mice; Molecular Chaperones; Molecular Sequence Data; Neoplasm Proteins - metabolism; Protein Binding; Rats; Sertoli Cells - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M001981200

hsp27 is involved in development of tolerance to stress, possibly by its involvement in molecular chaperoning, maintenance of glutathione status, and/or modulation of microfilament structure and function. We hypothesize that hsp27 function depends on specific association with other proteins. To discover proteins that associate with hsp27, we made a differentiated rat Sertoli cell cDNA expression library and screened it using the yeast two-hybrid system. We obtained a cDNA coding for a novel protein of 428 amino acids that we have named PASS1 (proteinassociated with small stress proteins 1). BLAST searches did not reveal major similarity of PASS1 to any known protein, but the cDNA sequence matched several mouse EST clones and shares 34% homology with aCaenorhabditis elegans genomic sequence. In vitro, bacterially expressed glutathioneS-transferase-PASS1 fusion protein bound to hsp27, and hsp27 was co-immunoprecipitated with c-Myc-tagged PASS1 overexpressed in several cell lines. The region of PASS1 responsible for association with hsp27 was identified as existing predominantly between amino acids 108 and 208 of PASS1. Northern hybridization and Western blot analysis demonstrated that PASS1 is expressed in several tissues, with the highest expression occurring in testis, primarily in Sertoli cells. The presence of a 1.4-kilobase PASS1 mRNA in kidney as well as the 1.8-kilobase mRNA seen in other tissues suggests that alternate splicing may occur in this organ. Ectopic expression of PASS1 in two cultured cell lines was observed to inhibit the ability of hsp27 to protect cells against heat shock, indicating that PASS1 does interact with hsp27 in the live cell.