Biomedical subjects
J A Atwater
Publications and source records attributed to J A Atwater.
Removal of an mRNA destabilizing element correlates with the increased oncogenicity of proto-oncogene fos.
AU-sequence motifs present in the 3' untranslated region (UTR) of many rapidly inducible messenger RNAs have been proposed to mediate their selective degradation. We have analyzed by quantitative nuclease S1 analysis the mRNA decay-rates of viral (v)/c-fos deletion mutants following transfection in a transient assay system. A 67 nucleotide mRNA destabilizing element, encompassing three conserved AUUUA motifs, was identified in the c-fos 3' UTR. The transforming ability of several v/c-fos recombinants correlates with the removal of the AU-rich sequence. Insertion of a c-fos DNA fragment containing the destabilizing element into the alpha-globin 3' UTR confers high instability to the otherwise stable alpha-globin mRNA. We conclude that a major parameter of oncogenicity by the c-fos gene is predicated by the presence or absence of AT-rich sequences located in the 3' non-coding region.
Biosynthesis of reovirus-specified polypeptides. Molecular cDNA cloning and nucleotide sequence of the reovirus serotype 1 Lang strain s2 mRNA which encodes the virion core polypeptide sigma 2.
Human reovirus serotype 1 Lang strain s2 mRNA, which encodes the virion inner capsid core polypeptide sigma 2, was cloned as a cDNA:mRNA heteroduplex in Escherichia coli using phage M13. A complete consensus nucleotide sequence was determined. The Lang strain s2 mRNA is 1331 nucleotides in length and possesses an open reading frame with a coding capacity of 335 amino acids, sufficient to account for a sigma 2 polypeptide of 37,682 daltons. Comparison of the serotype 1 Lang s2 sequence derived from cDNA clones of s2 mRNA with the serotype 3 Dearing S2 sequence derived from cDNA clones of the S2 dsRNA genome segment reveals 86 percent homology at the nucleotide level. The predicted sigma 2 polypeptides of the Lang and Dearing strains display 98 percent homology at the amino acid level. Of 147 silent nt differences in the translated region, 136 were in the third base position of codons.
Biosynthesis of reovirus-specified polypeptides. Efficiency of expression of cDNAs of the reovirus S1 and S4 genes in transfected animal cells differs at the level of translation.
Full-length cDNAs of the reovirus serotype 1 Lang strain S1 and S4 genes were cloned in Escherichia coli using bacteriophage M13 and expressed in monkey COS cells under the control of the SV40 late promoter using the eukaryotic expression vector pJC119. The s1-encoded sigma 1 and s4-encoded sigma 3 gene products were expressed in transfected COS cells and were indistinguishable from the authentic sigma 1 and sigma 3 polypeptides synthesized in reovirion-infected COS cells. The relative translational efficiencies of the s1 and s4 mRNAs in transfected COS cells were similar to the efficiencies observed in virion-infected cells; the s4 mRNA was translated approximately five times more efficiently than the s1 mRNA. Our results suggest that the differential translation of the reovirus s1 and s4 mRNAs in vivo may be attributed to intrinsic structural properties of the individual mRNAs and is independent of competition with other viral mRNAs.
Biosynthesis of reovirus-specified polypeptides. Molecular cDNA cloning and nucleotide sequence of the reovirus serotype 1 Lang strain bicistronic s1 mRNA which encodes the minor capsid polypeptide sigma 1a and the nonstructural polypeptide sigma 1bNS.
Human reovirus serotype 1 Lang strain s1 mRNA, which encodes the minor capsid cell attachment protein sigma 1a and the nonstructural protein sigma 1bNS, was cloned as a cDNA:mRNA heteroduplex in Escherichia coli using phage M13. The Lang strain s1 mRNA is 1462 nucleotides in length and possesses two open reading frames. The first begins at nt 14 and has a coding capacity of 418 amino acids, sufficient to account for sigma 1a; the second begins at nt 75 and has a coding capacity of 119 amino acids, sufficient to account for sigma 1bNS. Comparison of the Lang serotype s1 sequence derived from cDNA clones of s1 mRNA with the Lang S1 sequence derived from cDNA clones of the S1 dsRNA genome segment definitively establishes that reovirus plus-strand mRNA is structurally equivalent to the plus-strand of the dsRNA genome segment.
Biosynthesis of reovirus-specified polypeptides. Molecular cDNA cloning and nucleotide sequence of the reovirus serotype 1 Lang strain s3 mRNA which encodes the nonstructural RNA-binding protein sigma NS.
Human reovirus serotype 1 Lang strain s3 mRNA, which encodes the nonstructural RNA-binding polypeptide sigma NS, was cloned as a cDNA:mRNA heteroduplex in Escherichia coli using phage M13. A complete consensus nucleotide sequence was determined. The Lang strain s3 mRNA is 1198 nucleotides in length and possesses an open reading frame with a coding capacity of 366 amino acids, sufficient to account for a sigma NS polypeptide of 41,179 daltons. Comparison of the serotype 1 (Lang) s3 sequence with the serotype 3 (Dearing) s3 sequence reveals 86.8 percent homology at the nucleotide level. The predicted sigma NS polypeptides of the Lang and Dearing strains display 97 percent homology at the amino acid level.
Biosynthesis of reovirus-specified polypeptides. Molecular cDNA cloning and nucleotide sequence of the reovirus serotype 1 Lang strain s4 mRNA which encodes the major capsid surface polypeptide sigma 3.
Serotype 1 Lang strain s4 mRNA, which encodes the major capsid surface polypeptide sigma 3 of reovirions, was cloned as a cDNA:mRNA heteroduplex in Escherichia coli using phage M13. A complete consensus nucleotide sequence for s4 mRNA has been determined from cDNA clones. The Lang strain s4 mRNA is 1196 nucleotides in length and possesses an open reading frame with a coding capacity of 365 amino acids, sufficient to account for a sigma 3 polypeptide of 41,212 daltons. Comparison of the serotype 1 (Lang) s4 sequence with the serotype 3 (Dearing) s4 sequence reveals 94% homology at the nucleotide level; the predicted sigma 3 polypeptides of the Lang and Dearing strains display 96% homology at the amino acid level. Two third base C codons (leu:CUC and ser:AGC) are used about one-tenth as frequently in the reovirus s4 mRNAs as compared to mammalian cellular mRNAs.
Purification of double-stranded RNA-dependent protein kinase from mouse fibroblasts.
Explore the source record for details and available documents.
Biosynthesis of reovirus-specified polypeptides. The s1 mRNA synthesized in vivo is structurally and functionally indistinguishable from in vitro-synthesized s1 mRNA and encodes two polypeptides, sigma 1a and sigma 1bNS.
The structural and functional properties of the reovirus serotype 1 (Lang strain) s1 mRNA were examined. Reovirus s-class mRNAs, synthesized either in vivo within infected mouse L cells or in vitro by chymotrypsin-derived cores of purified virions, were purified by filter-hybridization using cDNA clones of the S-class genome segments. S1 cDNA-selected mRNA encoded the synthesis of the Mr approximately 12,000 nonstructural polypeptide designated sigma 1bNS in addition to the well-established structural polypeptide sigma 1, now designated sigma 1a. The coding properties of in vivo- and in vitro-synthesized s1 mRNA were equivalent: both encoded sigma 1a and sigma 1bNS. Primer extension analysis of s1 mRNA revealed a single major 5' terminus for both in vivo- and in vitro-synthesized s1 mRNA. These results suggest that there is a single transcript of the reovirus S1 genome segment which is functionally dicistronic, and likely encodes both sigma 1a and sigma 1bNS.