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Published May 28, 2010 | Published
Journal Article Open

Mouse Dfa Is a Repressor of TATA-box Promoters and Interacts with the Abt1 Activator of Basal Transcription


Our study of the mouse Ate1 arginyltransferase, a component of the N-end rule pathway, has shown that Ate1 pre-mRNA is produced from a bidirectional promoter that also expresses, in the opposite direction, a previously uncharacterized gene (Hu, R. G., Brower, C. S., Wang, H., Davydov, I. V., Sheng, J., Zhou, J., Kwon, Y. T., and Varshavsky, A. (2006) J. Biol. Chem. 281, 32559–32573). In this work, we began analyzing this gene, termed Dfa (divergent from Ate1). Mouse Dfa was found to be transcribed from both the bidirectional P_(Ate1/Dfa) promoter and other nearby promoters. The resulting transcripts are alternatively spliced, yielding a complex set of Dfa mRNAs that are present largely, although not exclusively, in the testis. A specific Dfa mRNA encodes, via its 3′-terminal exon, a 217-residue protein termed Dfa^A. Other Dfa mRNAs also contain this exon. DfaA is sequelogous (similar in sequence) to a region of the human/mouse HTEX4 protein, whose physiological function is unknown. We produced an affinity-purified antibody to recombinant mouse DfaA that detected a 35-kDa protein in the mouse testis and in several cell lines. Experiments in which RNA interference was used to down-regulate Dfa indicated that the 35-kDa protein was indeed Dfa^A. Furthermore, Dfa^A was present in the interchromatin granule clusters and was also found to bind to the Ggnbp1 gametogenetin-binding protein-1 and to the Abt1 activator of basal transcription that interacts with the TATA-binding protein. Given these results, RNA interference was used to probe the influence of Dfa levels in luciferase reporter assays. We found that Dfa^A acts as a repressor of TATA-box transcriptional promoters.

Additional Information

© 2010 by The American Society for Biochemistry and Molecular Biology, Inc. Received February 27, 2010. Revision received March 30, 2010. First Published on March 31, 2010. This work was supported, in whole or in part, by National Institutes of Health Grants GM31530 and DK39520 (to A. V.). This work was also supported by a grant from the March of Dimes Foundation. We are grateful to current and former members of the Varshavsky laboratory for advice and help. We thank K. I. Piatkov for helpful discussions, J. Zavzavadjian and E. Wall for advice with shRNA, and E. Udartseva for excellent technical assistance.

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