Avian retroviral mRNAs contain a long 5' untranslated leader of approximately 380 nucleotides. The leader includes sequences required for viral replication and three AUG codons which precede the AUG codon used for translational initiation of the gag and env genes. We have used sensitive, quantitative assays of viral gene transcription and translation to analyze the role of this mRNA leader in viral gene expression. By substituting segments from related viruses, we had previously shown that the endogenous avian provirus ev-1 contained a defective leader segment (B. R. Cullen, A. M. Skalka, and G. Ju, Proc. Natl. Acad. Sci. USA 80:2946-2950, 1983). The sequence analysis presented here, followed by comparison with the nondefective ev-2 endogenous provirus segment, identified the critical changes at nucleotides 4 and 7 upstream of the initiator AUG. These differences do not alter the most conserved nucleotides within the consensus sequence which precedes eucaryotic initiation codons, but lie within a nine-nucleotide region that is otherwise highly conserved among avian retrovirus strains. Analysis of a series of deletion mutants indicated that other sequences within the leader are also required for efficient expression. Characterization of the altered transcripts demonstrated that the presence of the defective ev-1 segment or the deletion of a ca. 200-nucleotide leader segment did not affect the steady-state level or splicing efficiency of these mRNAs. Thus, we conclude that the reduced expression of these mRNAs is due to a translational deficiency. These results indicate that specific leader sequences, other than the previously identified consensus nucleotides which precede eucaryotic AUG initiator codons, can influence eucaryotic gene translation.
Role of arginine 292 in the catalytic activity of chondroitin AC lyase from Flavobacterium heparinum
