Identification of functional p53-binding motifs in the mouse wig-1 promoter

AbstractWe previously identified wig-1 as a p53-induced mouse gene that encodes a nuclear zinc finger protein with unknown function. To investigate whether wig-1 is a direct target of p53-dependent transactivation, a DNA fragment corresponding to the promoter region was cloned and sequenced. Three regions containing consensus p53-binding sites were identified. Two p53-binding motifs formed DNA–protein complexes with p53 and were able to drive p53-dependent transcription in a luciferase reporter assay. Our results demonstrate that wig-1 is a direct target of p53-mediated transcriptional transactivation

The p53 target protein Wig-1 binds hnRNP A2/B1 and RNA Helicase A via RNA

AbstractThe p53-induced Wig-1 gene encodes a double stranded RNA-binding zinc finger protein. We generated Saos-2 osteosarcoma cells expressing tetracycline-inducible Flag-tagged human Wig-1. Induction of Wig-1 expression by doxycycline inhibited cell growth in a long-term assay but did not cause any changes in cell cycle distribution nor increased fraction of apoptotic cells. Using co-immunoprecipitation and mass spectrometry, we identified two Wig-1-binding proteins, hnRNP A2/B1 and RNA Helicase A, both of which are involved in RNA processing. The binding was dependent on the presence of RNA. Our results establish a link between the p53 tumor suppressor and RNA processing via hnRNPA2/B1 and RNA Helicase A.Structured summaryMINT-6542926, MINT-6542899:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with hnRNP A2/B1 (uniprotkb:P22626) by anti bait coimmunoprecipitation (MI:0006)MINT-6542945:RHA (uniprotkb:Q08211) physically interacts (MI:0218) with hnRNP A2/B1 (uniprotkb:P22626) by anti bait coimmunoprecipitation (MI:0006)MINT-6542918, MINT-6542891:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with RHA (uniprotkb:Q08211) by anti bait coimmunoprecipitation (MI:0006)MINT-6542867:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with RHA (uniprotkb:Q08211) by anti tag coimmunoprecipitation (MI:0007)MINT-6542879:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with hnRNP A2/B1(uniprotkb:P22626) by anti tag coimmunoprecipitation (MI:0007

Activation of a Translocated Human c-\u3ci\u3emyc\u3c/i\u3e Gene by an Enhancer in the Immunoglobulin Heavy-Chain Locus

A tissue-specific transcriptional enhancer element that is associated with the human immunoglobulin heavy-chain locus is defined. In a non-Hodgkin’s lymphoma that contains a translocated c-myc gene this enhancer is retained on the 14q+ chromosome and occurs within sequences shown to activate previously cryptic promoters of the c-myc gene

PRIMA-1MET induces nucleolar translocation of Epstein-Barr virus-encoded EBNA-5 protein

The low molecular weight compound, PRIMA-1MET restores the transcriptional transactivation function of certain p53 mutants in tumor cells. We have previously shown that PRIMA-1MET induces nucleolar translocation of p53, PML, CBP and Hsp70. The Epstein-Barr virus encoded, latency associated antigen EBNA-5 (also known as EBNA-LP) is required for the efficient transformation of human B lymphocytes by EBV. EBNA-5 associates with p53-hMDM2-p14ARF complexes. EBNA-5 is a nuclear protein that translocates to the nucleolus upon heat shock or inhibition of proteasomes along with p53, hMDM2, Hsp70, PML and proteasome subunits. Here we show that PRIMA-1MET induces the nucleolar translocation of EBNA-5 in EBV transformed B lymphoblasts and in transfected tumor cells. The PRIMA-1MET induced translocation of EBNA-5 is not dependent on the presence of mutant p53. It also occurs in p53 null cells or in cells that express wild type p53. Both the native and the EGFP or DSRed conjugated EBNA-5 respond to PRIMA-1MET treatment in the same way. Image analysis of DSRed-EBNA-5 expressing cells, using confocal fluorescence time-lapse microscopy showed that the nucleolar translocation requires several hours to complete. FRAP (fluorescence recovery after photobleaching) and FLIP (fluorescence loss in photobleaching) measurements on live cells showed that the nucleolar translocation was accompanied by the formation of EBNA-5 aggregates. The process is reversible since the aggregates are dissolved upon removal of PRIMA-1MET. Our results suggest that mutant p53 is not the sole target of PRIMA-1MET. We propose that PRIMA-1MET may reversibly inhibit cellular chaperons that prevent the aggregation of misfolded proteins, and that EBNA-5 may serve as a surrogate drug target for elucidating the precise molecular action of PRIMA-1MET

Targeting of MCL-1 kills MYC-driven mouse and human lymphomas even when they bear mutations in p53

The transcriptional regulator c-MYC is abnormally overexpressed in many human cancers. Evasion from apoptosis is critical for cancer development, particularly c-MYC-driven cancers. We explored which anti-apoptotic BCL-2 family member (expressed under endogenous regulation) is essential to sustain c-MYC-driven lymphoma growth to reveal which should be targeted for cancer therapy. Remarkably, inducible Cre-mediated deletion of even a single Mcl-1 allele substantially impaired the growth of c-MYC-driven mouse lymphomas. Mutations in p53 could diminish but not obviate the dependency of c-MYC-driven mouse lymphomas on MCL-1. Importantly, targeting of MCL-1 killed c-MYC-driven human Burkitt lymphoma cells, even those bearing mutations in p53. Given that loss of one allele of Mcl-1 is well tolerated in healthy tissues, our results suggest that therapeutic targeting of MCL-1 would be an attractive therapeutic strategy for MYC-driven cancers

WRAP53 Is Essential for Cajal Body Formation and for Targeting the Survival of Motor Neuron Complex to Cajal Bodies

The WRAP53 protein regulates the formation and maintenance of Cajal bodies (nuclear sub-organelles), as well as directs the recruitment of nuclear factors to Cajal bodies

p53 in the Clinics

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