The family Potyviridae includes >30% of known plant virus species, many of which are of great agricultural significance. These viruses have a positive sense RNA genome that is ≈10 kb long and contains a single long ORF. The ORF is translated into a large polyprotein, which is cleaved into ≈10 mature proteins. We report the discovery of a short ORF embedded within the P3 cistron of the polyprotein but translated in the +2 reading-frame. The ORF, termed pipo , is conserved and has a strong bioinformatic coding signature throughout the large and diverse Potyviridae family. Mutations that knock out expression of the PIPO protein in Turnip mosaic potyvirus but leave the polyprotein amino acid sequence unaltered are lethal to the virus. Immunoblotting with antisera raised against two nonoverlapping 14-aa antigens, derived from the PIPO amino acid sequence, reveals the expression of an ≈25-kDa PIPO fusion product in planta . This is consistent with expression of PIPO as a P3-PIPO fusion product via ribosomal frameshifting or transcriptional slippage at a highly conserved G _1-2 A _6-7 motif at the 5′ end of pipo . This discovery suggests that other short overlapping genes may remain hidden even in well studied virus genomes (as well as cellular organisms) and demonstrates the utility of the software package MLOGD as a tool for identifying such genes.

Influenza's Cryptic Constraint: Because of the well-known pandemic potential of influenza viruses, it is important to understand the range of molecular interactions between the virus and its host. Despite years of intensive research on the virus, Jagger et al. (p. 199 , published online 28 June; see the Perspective by Yewdell and Ince ) have found that the influenza A virus has been hiding a gene in its small negative-sense RNA genome. An overlapping open reading frame was found contained in the PA viral RNA polymerase gene, which is accessed by ribosomal frameshifting to produce a fusion protein containing the N-terminal messenger RNA (mRNA) endonuclease domain of PA and an alternative C-terminal X domain. The resulting polypeptide, PA-X, selectively degrades host mRNAs and, in a mouse model of infection, modulated cellular immune responses, thus limiting viral pathogenesis.

Selenoprotein P (Se-P) contains most of the selenium in plasma. Its function is not known. Mice with the Se-P gene deleted (Sepp(-/-)) were generated. Two phenotypes were observed: 1) Sepp(-/-) mice lost weight and developed poor motor coordination when fed diets with selenium below 0.1 mg/kg, and 2) male Sepp(-/-) mice had sharply reduced fertility. Weanling male Sepp(+/+), Sepp(+/-), and Sepp(-/-) mice were fed diets for 8 weeks containing <0.02-2 mg selenium/kg. Sepp(+/+) and Sepp(+/-) mice had similar selenium concentrations in all tissues except plasma where a gene-dose effect on Se-P was observed. Liver selenium was unaffected by Se-P deletion except that it increased when dietary selenium was below 0.1 mg/kg. Selenium in other tissues exhibited a continuum of responses to Se-P deletion. Testis selenium was depressed to 19% in mice fed an 0.1 mg selenium/kg diet and did not rise to Sepp(+/+) levels even with a dietary selenium of 2 mg/kg. Brain selenium was depressed to 43%, but feeding 2 mg selenium/kg diet raised it to Sepp(+/+) levels. Kidney was depressed to 76% and reached Sepp(+/+) levels on an 0.25 mg selenium/kg diet. Heart selenium was not affected. These results suggest that the Sepp(-/-) phenotypes were caused by low selenium in testis and brain. They strongly suggest that Se-P from liver provides selenium to several tissues, especially testis and brain. Further, they indicate that transport forms of selenium other than Se-P exist because selenium levels of all tissues except testis responded to increases of dietary selenium in Sepp(-/-) mice.

Shifting frames to make more proteins: Severe acute respiratory syndrome coronavirus 2 critically depends on the ribosomal frameshifting that occurs between two large open reading frames in its genomic RNA for expression of viral replicase. Programmed frameshifting occurs during translation, when the ribosome encounters a stimulatory pseudoknot RNA fold. Using a combination of cryo–electron microscopy and biochemistry, Bhatt et al. revealed that the pseudoknot resists unfolding as it lodges at the entry of the ribosomal messenger RNA channel. This causes back slippage of the viral RNA, resulting in a minus-1 shift of the reading frame of translation. A partially folded nascent viral polyprotein forms specific interactions inside the ribosomal tunnel that can influence the efficiency of frameshifting. Science , abf3546, this issue p. 1306

Genetic decoding is not 'frozen' as was earlier thought, but dynamic. One facet of this is frameshifting that often results in synthesis of a C-terminal region encoded by a new frame. Ribosomal frameshifting is utilized for the synthesis of additional products, for regulatory purposes and for translational 'correction' of problem or 'savior' indels. Utilization for synthesis of additional products occurs prominently in the decoding of mobile chromosomal element and viral genomes. One class of regulatory frameshifting of stable chromosomal genes governs cellular polyamine levels from yeasts to humans. In many cases of productively utilized frameshifting, the proportion of ribosomes that frameshift at a shift-prone site is enhanced by specific nascent peptide or mRNA context features. Such mRNA signals, which can be 5' or 3' of the shift site or both, can act by pairing with ribosomal RNA or as stem loops or pseudoknots even with one component being 4 kb 3' from the shift site. Transcriptional realignment at slippage-prone sequences also generates productively utilized products encoded trans-frame with respect to the genomic sequence. This too can be enhanced by nucleic acid structure. Together with dynamic codon redefinition, frameshifting is one of the forms of recoding that enriches gene expression.