Here we report the isolation of a cDNA encoding a new p53‐associating protein. This new protein has been called MDMX on the basis of its structural similarity to MDM2, which is especially notable in the p53‐binding domain. In addition, the putative metal binding domains in the C‐terminal part of MDM2 are completely conserved in MDMX. The middle part of the MDMX and MDM2 proteins shows a low degree of conservation. We can show by co‐immunoprecipitation that the MDMX protein interacts specifically with p53 in vivo. This interaction probably occurs with the N‐terminal part of p53, because the activity of the transcription activation domain of p53 was inhibited by co‐transfection of MDMX. Northern blotting showed that MDMX, like MDM2, is expressed in all tissues tested, and that several mRNAs for MDMX can be detected. Interestingly, the level of MDMX mRNA is unchanged after UV irradiation, in contrast to MDM2 transcription. This observation suggests that MDMX may be a differently regulated modifier of p53 activity in comparison with MDM2. Our study indicates that at least one additional member of the MDM protein family exists which can modulate p53 function.

Plants were grown in the greenhouse or in the field under standard conditions. We used the B73 wild-type line. For cytokinin induction experiments, 2-week-old seedlings were excised at the shoot-root junction, and stood in water supplemented with different concentrations of cytokinin. After treatments, the seedlings were dissected into a shoot fraction (apical and axillary shoot meristems, stem and 4-5 young leaf primordia) or a leaf fraction (expanding and mature leaves). For embryo culture experiments, pollinated ears at either 13 days after pollination (first leaf stage) or 16 days after pollination (second to third leaf stage) were sterilized in 30% commercial bleach solution for 20 min then rinsed five times with sterile water. The embryos were dissected out and cultured on maize embryo culture medium (1 £ Murashige and Skoog salts, 1 £ Gamborg's vitamins, 2% sucrose, 0.7% agar, pH 5.7) in some cases with the addition of cytokinin (kinetin, 10 25

In the present study, the human TEB4 is identified as a novel ER (endoplasmic reticulum)-resident ubiquitin ligase. TEB4 has homologues in many species and has a number of remarkable properties. TEB4 contains a conserved RING (really interesting new gene) finger and 13 predicted transmembrane domains. The RING finger of TEB4 and its homologues is situated at the N-terminus and has the unconventional C4HC3 configuration. The N-terminus of TEB4 is located in the cytosol. We show that the isolated TEB4 RING domain catalyses ubiquitin ligation in vitro in a reaction that is ubiquitin Lys48-specific and involves UBC7 (ubiquitin-conjugating enzyme 7). These properties are reminiscent of E3 enzymes, which are involved in ER-associated protein degradation. TEB4 is an ER degradation substrate itself, promoting its own degradation in a RING finger- and proteasome-dependent manner

The distribution and stability of the cellular tumor antigen p53 were studied in baby rat kidney cells transformed by region El sequences of nononcogenic adenovirus (Ad) type 5 (Ad5) or oncogenic type 12 (Adl2). In transformed cells expressing the large ElB T antigen of AdS, p53 was associated with this T antigen. The complexed proteins were concentrated in a cytoplasmic body, which has been shown to consist of a cluster of 8-nm filaments (A. Zantema et al., Virology 142:44-58, 1985). In transformed cells expressing the EIB region of Adl2, however, no association between the viral large T antigen and p53 was detectable. In the latter case, both proteins were found almost exclusively in the nucleus. The stability of p53 in both Ad5-and Adl2-transformed cells was increased relative to that in primary cells or cells immortalized by the ElA region only. Thus, the increased stability of p53 in Ad-transformed cells is not caused by association with a viral T antigen, but it correlates with expression of E1B and with morphological transformation.

SummaryRNA viruses pose a threat to public health that is exacerbated by the dearth of antiviral therapeutics. The RNA-dependent RNA polymerase (RdRp) holds promise as a broad-spectrum, therapeutic target because of the conserved nature of the nucleotide-substrate-binding and catalytic sites. Conventional, quantitative, kinetic analysis of antiviral ribonucleotides monitors one or a few incorporation events. Here, we use a high-throughput magnetic tweezers platform to monitor the elongation dynamics of a prototypical RdRp over thousands of nucleotide-addition cycles in the absence and presence of a suite of nucleotide analog inhibitors. We observe multiple RdRp-RNA elongation complexes; only a subset of which are competent for analog utilization. Incorporation of a pyrazine-carboxamide nucleotide analog, T-1106, leads to RdRp backtracking. This analysis reveals a mechanism of action for this antiviral ribonucleotide that is corroborated by cellular studies. We propose that induced backtracking represents a distinct mechanistic class of antiviral ribonucleotides.