Evolution and ecology of influenza A viruses.

The small (40S) subunit of eukaryotic ribosomes is believed to bind initially at the capped 5'-end of messenger RNA and then migrate, stopping at the first AUG codon in a favorable context for initiating translation. The first-AUG rule is not absolute, but there are rules for breaking the rule. Some anomalous observations that seemed to contradict the scanning mechanism now appear to be artifacts. A few genuine anomalies remain unexplained.

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The scanning model for translation: an update.

5-Noncoding sequences have been tabulated for 211 messenger RNAs from higher eukaryotic cells. The 5'-proximal AUG triplet serves as the initiator codon in 95% of the mRNAs examined. The most conspicuous conserved feature is the presence of a purine (most often A) three nucleotides upstream from the AUG initiator codon; only 6 of the mRNAs in the survey have a pyrimidine in that position. There is a predominance of C in positions -1, -2, -4 and -5, just upstream from the initiator codon. The sequence CCAGCCAUG (G) thus emerges as a consensus sequence for eukaryotic initiation sites. The extent to which the ribosome binding site in a given mRNA matches the -1 to -5 consensus sequence varies: more than half of the mRNAs in the tabulation have 3 or 4 nucleotides in common with the CCACC consensus, but only ten mRNAs conform perfectly.

Compilation and analysis of sequences upstream from the translational start site in eukaryotic mRNAs

The influenza virus neuraminidase glycoprotein is a tetramer with a box-shaped head, 100 X 100 X 60 A, attached to a slender stalk. The three-dimensional structure of neuraminidase heads shows that each monomer is composed of six topologically identical beta-sheets arranged in a propeller formation. The tetrameric enzyme has circular 4-fold symmetry stabilized in part by metal ions bound on the symmetry axis. Sugar residues are attached to four of the five potential glycosylation sequences, and in one case contribute to the interaction between subunits in the tetramer.

Structure of the influenza virus glycoprotein antigen neuraminidase at 2.9 Å resolution

The catalytic sites of influenza virus neuraminidase are located on the upper corners of the box-shaped tetramer that forms the head of the molecule. Antigenic determinants form a nearly-continuous surface across the top of the monomer encircling the catalytic site. Approximately the same number of amino acid sequence changes occurred in these determinants between the years 1968 and 1975 as occurred in the antigenic sites of influenza virus haemagglutinin in the same period.

Structure of the catalytic and antigenic sites in influenza virus neuraminidase

RNA segment 6 of the influenza B virus genome codes for a previously unidentified polypeptide designated NB. The reading frame for this polypeptide begins with the first AUG codon on the mRNA and overlaps the reading frame for the viral neuraminidase by 292 nucleotides. The amino acid sequence of polypeptide NB deduced from the nucleotide sequence of the B/Lee/40 strain consists of 100 amino acids with a molecular weight of 11,242. The sequence contains four potential glycosylation sites, and the protein has been found to be glycosylated in infected cells. NB has not been found in virions. Sucrose gradient sedimentation and analysis of the structure of the mRNA by nuclease S1 mapping and sequence analysis by the primer extension method indicated that polypeptide NB and the neuraminidase are translated from a single bicistronic mRNA. A protein analogous to NB has not been found with influenza A virus, and this represents a major difference between the two virus types.

A previously unrecognized influenza B virus glycoprotein from a bicistronic mRNA that also encodes the viral neuraminidase

The complete nucleotide sequence of the neuraminidase gene of influenza virus B/Lee/40 was derived from a cloned cDNA copy of virion RNA segment 6 and its corresponding mRNA. The RNA segment contains 1,557 virus-specific nucleotides, and the protein encoded by the longest open reading frame has a total of 466 amino acids with a molecular weight of 51,721. As is the case with the influenza A virus neuraminidases, the deduced amino acid sequence of the influenza B protein includes a single hydrophobic region near the amino terminus which would be capable of spanning the lipid bilayer of the viral or cell membrane. There are four potential glycosylation sites in the protein, two of which are near the amino-terminal hydrophobic region. Comparisons of the nucleotide and amino acid sequences with those of influenza A virus neuraminidases revealed seven regions of extensive homology within the central portion of the molecules, including 12 conserved cysteine residues. Five other cysteine residues in the terminal portions were also conserved.

Complete nucleotide sequence of the neuraminidase gene of influenza B virus

The nucleotide sequences of the 3' noncoding regions of all eight segments of influenza B virus RNA and the sequences of the 5' noncoding regions of segments 4-8 were determined in virus strains isolated over a period of 40 years. Nearly complete conservation of the noncoding sequences was found. Nine nucleotides at the 3' termini and 11 nucleotides at the 5' termini were common to all segments examined. In the region immediately adjacent to the common 3' terminal region, the nucleotides were specific for each segment and these segment-specific sequences were conserved in all strains examined. In each of the five segments in which both termini were examined, the segment-specific 3' sequences exhibited perfect inverted complementarity to a segment-specific sequence adjacent to the common 5' terminus. In addition, in the 3' noncoding region of RNA segments 1-3, which encode proteins involved in RNA synthesis, a single nucleotide substitution at position 10 was found that distinguishes these segments from segments 4-8. Comparison of these data with published reports has revealed that some of the features found in the noncoding regions of influenza B virus are also present in influenza A and C virus RNAs. In the RNAs of all three virus types, there is a segment-specific sequence of nucleotides near the 3' terminus that shows inverted complementarity to a sequence near the 5' terminus. This segment-specific sequence may play a role in the transcription of individual segments or in sorting of segments during virion assembly.

https://www.pnas.org/content/pnas/84/9/2703.full.pdf

Michael W. Shaw

Papers

A previously unrecognized influenza B virus glycoprotein from a bicistronic mRNA that also encodes the viral neuraminidase

Complete nucleotide sequence of the neuraminidase gene of influenza B virus

Segment-specific and common nucleotide sequences in the noncoding regions of influenza B virus genome RNAs.

A specific sub-set of host-cell mRNAs prime influenza virus mRNA synthesis

Studies on the synthesis of the influenza b virus nb glycoprotein