N-CoR (nuclear hormone receptor corepressor) was identified originally as a corepressor that binds to, and mediates transcriptional repression by, nuclear hormone receptors (Hö rlein et al. 1995). Thyroid-hormone and retinoic-acid receptors (TR and RAR) of the nuclear hormone receptor family actively repress the transcription of target genes in the absence of ligand (Chambon 1994;Mangelsdorf et al. 1995). Transcriptional repression is mediated by a conserved region in the aminoterminal part of the ligand-binding domain of TR (Baniahmad et al. 1995). N-CoR binds to the ligand-binding domain, termed the Co-R box, and, thereby, mediates transcriptional repression (Hö rlein et al. 1995). N-CoR is a large protein with a molecular mass of 270,000 (Mr 270K), and contains three repressor domains in its amino-terminal region (Hö rlein et al. 1995). Another corepressor, SMRT, which also binds to the Co-R box, shows striking homology to N-CoR (Chen and Evans 1995). N-CoR also forms a complex with mammalian Sin3 orthologs (mSin3A and mSin3B), which bind to another repressor, Mad (Alland et al. 1997;Hassing et al. 1997;Heinzel et al. 1997;Laherty et al. 1997;Nagy et al. 1997). The basic helix-loop-helix (bHLH) proteins of the Mad family act as transcriptional repressors after heterodimerization with Max (Ayer et al. 1993). N-CoR is required for Mad-induced transcriptional repression. The same target sequence of Mad/Max, the so-called E-box, is also recognized by a heterodimer of Myc/Max that activates transcription. It is believed that transcriptional activation of a group of target genes by Myc/Max enhances cellular proliferation or transformation, whereas transcriptional repression of the same target genes by Mad/Max leads to suppression of proliferation or induction of terminal differentiation in a wide range of cell types Chin et al. 1995;Roussel et al. 1996)

Mortalin, also known as mthsp70/GRP75/PBP74, interacts with the tumor suppressor protein p53 and inactivates its transcriptional activation and apoptotic functions. Here, we examined the level of mortalin expression in a large variety of tumor tissues, tumor-derived and in vitro immortalized human cells. It was elevated in many human tumors, and in all of the tumor-derived and in vitro immortalized cells. In human embryonic fibroblasts immortalized with an expression plasmid for hTERT, the telomerase catalytic subunit, with or without human papillomavirus E6 and E7 genes, we found that subclones with spontaneously increased mortalin expression levels became anchorage-independent and acquired the ability to form tumors in nude mice. Furthermore, overexpression of mortalin was sufficient to increase the malignancy of breast carcinoma cells. The study demonstrates that upregulation of mortalin contributes significantly to tumorigenesis, and thus is a good candidate target for cancer therapy. ' 2006 Wiley-Liss, Inc.

The mortalin genes, mot-1 and mot-2, are hsp70 family members that were originally cloned from normal and immortal murine cells, respectively. Their proteins differ by only two amino acid residues but exhibit different subcellular localizations, arise from two distinct genes, and have contrasting biological activities. We report here that the two proteins also differ in their interactions with the tumor suppressor protein p53. The pancytosolic mot-1 protein in normal cells did not show colocalization with p53; in contrast, nonpancytosolic mot-2 and p53 overlapped significantly in immortal cells. Transfection of mot-2 but not mot-1 resulted in the repression of p53-mediated transactivation in p53-responsive reporter assays. Inactivation of p53 by mot-2 was supported by the down-regulation of p53-responsive genes p21 WAF-1 and mdm-2 in mot-2-transfected cells only. Furthermore, NIH 3T3 cells transfected with expression plasmid encoding green fluorescent proteintagged mot-2 but not mot-1 showed an abrogation of nuclear translocation of wild-type p53. These results demonstrate a novel mechanism of p53 inactivation by mot-2 protein.Evidence has been accumulating that inactivation of p53, a tumor suppressor and cellular transcription factor (1), is involved in cellular transformation and immortalization (2-5). Extensive analyses of p53 have defined at least four functional domains, including an amino terminus transactivation domain (amino acids 1-44), a sequence-specific DNA-binding domain (amino acids 100 -300), a carboxyl terminus oligomerization domain, and a regulatory domain (amino acids Ref. 6), and shown that the conformation of p53 and its interactions with other proteins have key roles in its various cellular activities (7,8). Several cellular proteins, including some of the hsp70 family members, have been shown to interact with p53 (9 -12). Although mutational or mdm-2-mediated inactivation of p53 is a common event involved in cellular transformation (1), p53 is inactivated in a considerable number of tumors and transformed cells by an unknown mechanism(s).We initially cloned mortalins mot-1 and mot-2, which code for pancytosolically and perinuclearly distributed members of the hsp70 family of proteins, from normal and immortal murine cells, respectively (13,14). The open reading frames of the two types of murine mortalins differ in two nucleotides, encode proteins differing in two amino acids, arise from distinct genes, and have contrasting biological activities (13-16). RNA in situ hybridization and immunohistochemical studies on mortalin in normal murine tissues showed a higher level of expression in nondividing cell populations than in dividing cells. However, tumor tissues were seen to have a high intensity of mortalin staining by an antibody that reacts with both the mot-1 and mot-2 proteins (17, 18). Mortalin was also identified as PBP-74, mtHSP70, and Grp75 and has been assigned roles in antigen processing, in vivo nephrotoxicity, and radioresistance in independent studies from other groups (19,20)...

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