In order to characterize Sterling's triiodothyronine (T3) mitochondrial receptor using photoaffinity labeling, we observed two specific T3-binding proteins in the inner membrane (28 kDa) and in the matrix (43 kDa) of rat liver mitochondria. Western blots and immunoprecipitation using antibodies raised against the T3-binding domain of the T3 nuclear receptor c-Erb A alpha 1 indicated that at least the 43-kDa protein was c-Erb A alpha 1-related. In addition, gel mobility shift assays demonstrated the occurrence of a c-Erb A alpha 1-related mitochondrial protein that specifically binds to a natural or a palindromic thyroid-responsive element. Moreover, this protein specifically binds to a direct repeat 2 sequence located in the D-loop of the mitochondrial genome. Furthermore, electron microscopy studies allowed the direct observation of a c-Erb A-related protein in mitochondria. Lastly, the relative amounts of the 43-kDa protein related to c-Erb A alpha 1 were in good correlation with the known mitochondrial mass in three typical tissues. Interestingly, expression of a truncated form of the c-Erb A alpha 1 nuclear receptor in CV1 cells was associated with a mitochondrial localization and a stimulation of mitochondrial activity. These results supply evidence of the localization of a member of the nuclear receptor superfamily in the mitochondrial matrix involved in the regulation of mitochondrial activity that could act as a mitochondrial T3-dependent transcription factor.

The signal transducer and activator of transcription STAT5 plays a major role in the cellular response to cytokines, but the mechanism by which it activates transcription remains poorly understood. We show here that deacetylase inhibitors (trichostatin A, suberoylanilide hydroxamic acid, and sodium butyrate) prevent induction of endogenous STAT5 target genes, implying that a deacetylase activity is required for that process. Microarray analyses revealed that this requirement is common to all STAT5 target genes. Using chromatin immunoprecipitation, we show that, following STAT5 DNA binding, deacetylase inhibitors block transcription initiation by preventing recruitment of the basal transcription machinery. This inhibition is not due to effects on histone H3 and H4 acetylation or chromatin remodeling within the promoter region. This novel mechanism of transactivation by STAT5 provides a rationale for the use of deacetylase inhibitors for therapeutic intervention in STAT5-associated cancers.Transcription in eukaryotes is a multistep process that requires distinct multiprotein complexes. Histone acetyltransferases and histone deacetylases (HDACs) are chromatinmodifying enzymes that tightly cooperate with chromatin-remodeling enzymes to regulate accessibility of the template to DNA binding factors and RNA polymerase II (43). Beside histone acetylation, a variety of histone and nonhistone protein modifications (acetylation, phosphorylation, and methylation), as well as modification of DNA itself by methylation, regulate transcription initiation (reviewed in references 3, 7, 17, 26, 46, 52, 57, and 59). Orchestration of the events required for transcriptional activation is promoter specific. These events (recruitment of the transcription factor, chromatin modification and remodeling, and assembly of the preinitiation complex) do not follow an obligate order and are coordinated so that each step facilitates the next, eventually resulting in efficient transcription initiation (1,10,11,58).Transcriptional activation is generally correlated with histone acetylation by histone acetyltransferase complexes, and repression is correlated with deacetylation by HDAC complexes (22,23,34,54,60). However, the analysis of a variety of Saccharomyces cerevisiae promoters has recently revealed that transcriptional activation is not necessarily associated with increased histone acetylation (13). This is consistent with the observation that expression of a small subset of genes (2%) is affected in response to histone hyperacetylation induced by the deacetylase inhibitor trichostatin A (TSA) (61). In addition, genome-wide genetic studies with yeast clearly demonstrated that HDACs are required in both transcriptional activation and repression (4,44,63,64,69).The signal transducer and activator of transcription STAT5functions as an important downstream effector of cytokine signaling. It plays key roles in regulating immune responses, cell proliferation, differentiation, survival, and oncogenesis. STAT5 proteins are present as inactive monome...

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Previous studies have identified the DUB family of cytokine-regulated murine deubiquitinating enzymes, which play a role in the control of cell proliferation and survival. Through data base analyses and cloning, we have identified a human cDNA (DUB-3) that shows significant homology to the known murine DUB family members. Northern blotting has shown expression of this gene in a number of tissues including brain, liver, and muscle, with two transcripts being apparent (1.6 and 1.7 kb). In addition, expression was observed in cell lines including those derived from a number of hematopoietic tumors such as the Burkitt's lymphoma cell line RAJI. We have also demonstrated that DUB-3, which was shown to be an active deubiquitinating enzyme, is induced in response to interleukin-4 and interleukin-6 stimulation. Finally, we have demonstrated that constitutive expression of DUB-3 blocks proliferation and can initiate apoptosis in both IL-3-dependent Ba/F3 cells and NIH3T3 fibroblasts. These findings suggest that human DUB-3, like the murine DUB family members, is transiently induced in response to cytokines and can, when constitutively expressed, block growth factor-dependent proliferation.