|Function and cDNA Resources Section
DOE Human Genome Program Contractor-Grantee
113. Diversity in the Proteome: Homologous DNA Replicase Genes Use Alternatives of Transcriptional Slippage or Translational Frameshifting for Gene Expression
Norma M. Wills, Bente Larsen, Chad Nelson, John F. Atkins, and Raymond F. Gesteland
Department of Human Genetics, University of Utah, 15 N. 2030 East Room 7410, Salt Lake City, UT 84112-5330
A newly discovered contributor to the complexity of the proteome stems from generation of multiple RNAs from a single gene by transcriptional slippage. In the Thermus thermophilus dnaX gene, transcriptional slippage on a run of nine T residues results in a mixture of mRNAs differing in the number of A residues. Standard translation of a subpopulation of mRNAs yields the full-length tau protein while another subpopulation produces the shortened gamma protein. Transcriptional slippage was implicated by determining the masses of PCR products spanning the run of A residues using mass spectrometry. With genomic DNA as the PCR template, the predominant signals correspond to molecules containing nine A/Ts. The pattern is strikingly different using reverse-transcribed mRNA as template. There are multiple signals corresponding to molecules containing 8-18 A/Ts showing heterogeneity in the mRNA population transcribed from the single dnaX gene.
This method of dnaX gene expression in Thermus thermophilus differs markedly from dnaX expression in E. coli where two analogous proteins are produced from a single dnaX gene by ribosomal frameshifting. Standard translation of the homogeneous mRNA population produces the full-length tau protein. Approximately 50% of the time, ribosomes shift to the -1 reading frame at a specific sequence, A AAA AAG, stimulated by signals in the mRNA and produce the shortened gamma protein. It is surprising that two rather similar dnaX sequences lead to very different modes of expression in the two organisms. The global importance of these and other alternative mechanisms of gene expression will be revealed by proteome analysis now underway.
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