Molecular cloning, expression and localization of human 105 kDa heat shock protein, hsp105

Abstract

We have shown that the 105 kDa heat shock protein (hsp105alpha) and hsp105beta (42 degreesC-specific heat shock protein) constitute high molecular mass (HMM) heat shock proteins (HSPs) in mouse cells. However, since HMM HSPs have not been identified in human cells, we screened a cDNA library constructed with poly(A)+ RNA derived from heat-shocked human HeLa cells using mouse hsp105alpha cDNA. Two full-length cDNA clones were obtained: the pBH105-1 insert encoded an 858-amino-acid protein, and the pBH105-2 insert encoded an 814-amino-acid protein which lacked 44 amino acids from pBH105-1. The deduced amino acid sequences of pBH105-1 and pBH105-2 inserts were highly homologous to mouse hsp105alpha (96%) and hamster hsp110 (92%), and to mouse hsp105beta (93%), respectively. The transcript of pBH105-1 was induced by various stresses in HeLa cells, but the transcript of pBH105-2 was only induced during heat shock at 42 degreesC. These results indicated that pBH105-1 and pBH105-2 encoded human hsp105alpha and hsp105beta, respectively. Furthermore, a rabbit antibody was raised against recombinant human hsp105alpha, and immunofluorescence study also confirmed that hsp105 was present in the cytoplasm but was not found in the nucleoli of mammalian cells under both nonstressed and stressed conditions.