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P Ordentlich

Publications and source records attributed to P Ordentlich.

12 recordsLinked to original sources
PubMed2001

Sharp, an inducible cofactor that integrates nuclear receptor repression and activation.

A yeast two-hybrid screen using the conserved carboxyl terminus of the nuclear receptor corepressor SMRT as a bait led to the isolation of a novel human gene termed SHARP (SMRT/HDAC1 Associated Repressor Protein). SHARP is a potent transcriptional repressor whose repression domain (RD) interacts directly with SMRT and at least five members of the NuRD complex including HDAC1 and HDAC2. In addition, SHARP binds to the steroid receptor RNA coactivator SRA via an intrinsic RNA binding domain and suppresses SRA-potentiated steroid receptor transcription activity. Accordingly, SHARP has the capacity to modulate both liganded and nonliganded nuclear receptors. Surprisingly, the expression of SHARP is itself steroid inducible, suggesting a simple feedback mechanism for attenuation of the hormonal response.

Amino Acid Motifs↗
PubMed2000

Identification of a nuclear domain with deacetylase activity.

Here, we describe the identification and characterization of a nuclear body (matrix-associated deacetylase body) whose formation and integrity depend on deacetylase activity. Typically, there are 20-40 0.5-microM bodies per nucleus, although the size and number can vary substantially. The structure appears to contain both class I and the recently described class II histone deacetylases (HDAC)5 and 7 along with the nuclear receptor corepressors SMRT (silencing mediator for retinoid and thyroid receptor) and N-CoR (nuclear receptor corepressor). Addition of the deacetylase inhibitors trichostatin A and sodium butyrate completely disrupt these nuclear bodies, providing a demonstration that the integrity of a nuclear body is enzyme dependent. We demonstrate that HDAC5 and 7 can associate with at least 12 distinct proteins, including several members of the NuRD and Sin3A repression complexes, and appear to define a new but related complex.

Amino Acid Sequence↗
PubMed2000

Isolation of a novel histone deacetylase reveals that class I and class II deacetylases promote SMRT-mediated repression.

The transcriptional corepressor SMRT functions by mediating the repressive effect of transcription factors involved in diverse signaling pathways. The mechanism by which SMRT represses basal transcription has been proposed to involve the indirect recruitment of histone deacetylase HDAC1 via the adaptor mSin3A. In contrast to this model, a two-hybrid screen on SMRT-interacting proteins resulted in the isolation of the recently described HDAC5 and a new family member termed HDAC7. Molecular and biochemical results indicate that this interaction is direct and in vivo evidence colocalizes SMRT, mHDAC5, and mHDAC7 to a distinct nuclear compartment. Surprisingly, HDAC7 can interact with mSin3A in yeast and in mammalian cells, suggesting association of multiple repression complexes. Taken together, our results provide the first evidence that SMRT-mediated repression is promoted by class I and class II histone deacetylases and that SMRT can recruit class II histone deacetylases in a mSin3A-independent fashion.

Amino Acid Sequence↗
PubMed1999

Unique forms of human and mouse nuclear receptor corepressor SMRT.

Nuclear hormone receptors have been shown to repress transcription in the absence of ligand. This repression is mediated by a corepressor complex that contains the Sin3A protein and histone deacetylases (HDAC1 and 2). Studies by several groups demonstrate that this complex is recruited to nuclear receptors through the highly related corepressors SMRT (silencing mediator of retinoid acid and thyroid hormone receptor) and N-CoR (nuclear receptor corepressor). We describe here the cloning, characterization, and chromosomal mapping of forms of human and mouse SMRT that includes a 1,000-aa extension, which reveals striking homology to the amino terminus of N-CoR. Structure and function studies of wild-type and natural splicing variants suggest the presence of 3-4 amino terminal domains that repress in a cooperative as well as mechanistically distinct fashion.

Amino Acid Sequence↗
PubMed1998

A histone deacetylase corepressor complex regulates the Notch signal transduction pathway.

The Delta-Notch signal transduction pathway has widespread roles in animal development in which it appears to control cell fate. CBF1/RBP-Jkappa, the mammalian homolog of Drosophila Suppressor of Hairless [Su(H)], switches from a transcriptional repressor to an activator upon Notch activation. The mechanism whereby Notch regulates this switch is not clear. In this report we show that prior to induction CBF1/RBP-Jkappa interacts with a corepressor complex containing SMRT (silencing mediator of retinoid and thyroid hormone receptors) and the histone deacetylase HDAC-1. This complex binds via the CBF1 repression domain, and mutants defective in repression fail to interact with the complex. Activation by Notch disrupts the formation of the repressor complex, thus establishing a molecular basis for the Notch switch. Finally, ESR-1, a Xenopus gene activated by Notch and X-Su(H), is induced in animal caps treated with TSA, an inhibitor of HDAC-1. The functional role for the SMRT/HDAC-1 complex in CBF1/RBP-Jkappa regulation reveals a novel genetic switch in which extracellular ligands control the status of critical nuclear cofactor complexes.

3T3 Cells↗
PubMed1998

Human deltex is a conserved regulator of Notch signalling.

A fundamental cell-fate control mechanism regulating multicellular development is defined by the Notch-signalling pathway. Developmental and genetic studies of wild type and activated Notch-receptor expression in diverse organisms suggest that Notch plays a general role in development by governing the ability of undifferentiated precursor cells to respond to specific signals. Notch signalling has been conserved throughout evolution and controls the differentiation of a broad spectrum of cell types during development. Genetic studies in Drosophila have led to the identification of several components of the Notch pathway. Two of the positive regulators of the pathway are encoded by the suppressor of hairless [Su(H)] and deltex (dx) genes. Drosophila dx encodes a ubiquitous, novel cytoplasmic protein of unknown biochemical function. We have cloned a human deltex homologue and characterized it in parallel with its Drosophila counterpart in biochemical assays to assess deltex function. Both human and Drosophila deltex bind to Notch across species and carry putative SH3-binding domains. Using the yeast interaction trap system, we find that Drosophila and human deltex bind to the human SH3-domain containing protein Grb2 (ref. 10). Results from two different reporter assays allow us for the first time to associate deltex with Notch-dependent transcriptional events. We present evidence linking deltex to the modulation of basic helix-loop-helix (bHLH) transcription factor activity.

Amino Acid Sequence↗
PubMed1998

Notch inhibition of E47 supports the existence of a novel signaling pathway.

E47 is a widely expressed transcription factor that activates B-cell-specific immunoglobulin gene transcription and is required for early B-cell development. In an effort to identify processes that regulate E47, and potentially B-cell development, we found that activated Notch1 and Notch2 effectively inhibit E47 activity. Only the intact E47 protein was inhibited by Notch-fusion proteins containing isolated DNA binding and activation domains were unaffected-suggesting that Notch targets an atypical E47 cofactor. Although overexpression of the coactivator p300 partially reversed E47 inhibition, results of several assays indicated that p300/CBP is not a general target of Notch. Notch inhibition of E47 did not correlate with its ability to activate CBF1/RBP-Jkappa, the mammalian homolog of Suppressor of Hairless, a protein that associates physically with Notch and defines the only known Notch signaling pathway in drosophila. Importantly, E47 was inhibited independently of CBF1/RPB-Jkappa by Deltex, a second Notch-interacting protein. We provide evidence that Notch and Deltex may act on E47 by inhibiting signaling through Ras because (i) full E47 activity was found to be dependent on Ras and (ii) both Notch and Deltex inhibited GAL4-Jun, a hybrid transcription factor whose activity is dependent on signaling from Ras to SAPK/JNK.

3T3 Cells↗
PubMed1997

Selective utilization of basic helix-loop-helix-leucine zipper proteins at the immunoglobulin heavy-chain enhancer.

The microE3 E box within the immunoglobulin heavy-chain (IgH) enhancer binds several proteins of the basic helix-loop-helix-leucine zipper (bHLHzip) class, including TFE3, USF1, and Max. Both TFE3 and USF have been described as transcriptional activators, and so we investigated their possible roles in activating the IgH enhancer in vivo. Although TFE3 activated various enhancer-based reporters, both USF1 and Max effectively inhibited transcription. Inhibition by USF correlated with the lack of a strong activation domain and was the result of the protein neutralizing the microE3 site. The effects of dominant-negative derivatives of TFE3 and USF1 confirmed that TFE3, or a TFE3-like protein, is the primary cellular bHLHzip protein that activates the IgH enhancer. In addition to providing a physiological role for TFE3, our results call into question the traditional view of USF1 as an obligate transcriptional activator.

3T3 Cells↗
PubMed1996

HMG box-activating factors 1 and 2, two HMG box transcription factors that bind the human Ig heavy chain enhancer.

We present the isolation of two cDNAs that encode distinct, yet related, proteins that bind the HE2 region of the human Ig heavy chain (IgH) enhancer. Designated HMG box-activating factors (HAF) 1 and 2, the two proteins are new members of the HMG box family of DNA binding proteins. Both are potent transcription activators when expressed 1) as GAL4 fusions targeted to promoters containing GAL4 operators, or 2) as intact proteins targeted to minimal promoters containing binding sites derived from the IgH enhancer. HAF-1 and HAF-2 mRNAs are apparently expressed in both B cells and non-B cells. However, activity generated by the isolated HE2 region in B cells is dependent on both an intact HAF-1/HAF-2 binding site and at least one additional site that has been reported previously to bind a B cell-restricted protein. Our results suggest a collaborative role for either or both HAF-1 and HAF-2 in establishing the B cell activity of the human IgH enhancer.

Amino Acid Sequence↗
PubMed1991

Secretion of lysosomal enzymes by drug-sensitive and multiple drug-resistant cells.

The multiple drug-resistant human lymphoblastic leukemic cell, CEM/VLB100, in which P-glycoprotein (P-170) is overexpressed, has a lowered content of lysosomal enzymes, such as N-acetylglucosaminidase and beta-galactosidase, and the relative rates of secretion of these enzymes are significantly greater than those of its drug-sensitive counterpart, CEM. The ability of CEM/VLB100 cells to accumulate [3H]vinblastine ([3H]-VLB) is also greatly reduced. Multiple drug-resistant cells whose mode of resistance is not associated with P-170 do not have reduced enzyme content, and their rate of secretion is the same as that of their drug-sensitive parents. Linkage of drug and enzyme elimination is suggested by the observation that verapamil inhibits both the efflux of [3H]VLB and the secretion of lysosomal enzymes in CEM/VLB100 cells; the content of both [3H]VLB and enzyme increases in these cells when chronically exposed to verapamil. Further, both secretion of N-acetylglucosaminidase and efflux of [3H]VLB by CEM/VLB100 cells are enhanced by the addition of NaCl to the suspending, sucrose-containing medium. When cells have taken up [3H]VLB and are then fractionated by means of a Percoll centrifugation gradient, the distribution of drug among the various populations of vesicles is similar to that of N-acetylglucosaminidase. Losses of both enzyme and drug take place from these vesicular populations to varying degrees, when CEM/VLB100 cells are induced to secrete. It is proposed that, in a multiple drug-resistant cell such as CEM/VLB100, the presence of P-170 in the plasma membrane may, in some indirect manner, lead to increased exocytosis of lysosomal enzyme, ultimately resulting in a significant depletion of enzyme. Further, a toxic, cationic drug such as vinblastine, accumulating in lysosomes and acidic vesicles, is also eliminated from the cell by exocytosis. This pathway may supplement the known, major mode of efflux directly involving P-170.

ATP Binding Cassette Transporter, Subfamily B, Mem↗