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M B Mathews

Publications and source records attributed to M B Mathews.

At least 109 records · Page 6Linked to original sources
PubMed1987

Cyclic AMP inhibits increased collagen production by cyclically stretched smooth muscle cells.

Rabbit arterial smooth muscle cells, grown on elastin membranes which were cyclically elongated and relaxed, responded by increasing their rates of synthesis of protein and, in particular, of collagen, compared to stationary controls. Raising intracellular cyclic AMP (cAMP) levels by adding theophylline or dibutyryl cAMP to the culture medium prevented the synthetic response to cyclic stretching, but did not alter the rates of protein or collagen synthesis by stationary controls. Both synthesis and degradation of collagen by cyclically stretched cells increased in parallel such that the proportion of synthesized collagen that was degraded was similar to that found in the stationary cultures. Collagen degradation was not affected by theophylline administration to stationary cell cultures but the drug increased degradation of collagen by cyclically stretched cells. We conclude that the net production of protein, and in particular of a structural protein, collagen, by arterial smooth muscle cells subjected to the mechanical force of stretching was inhibited when intracellular levels of cAMP were raised. The results suggest that cAMP may play a role in the modulation of structural protein content of artery walls in response to changes in tensile stress.

Animals↗
PubMed1987

Two human tRNA(Ala) families are recognized by autoantibodies in polymyositis sera.

Autoantibodies to aminoacyl-tRNA synthetases are common in myositis. Sera of one particular class, the PL-12 specificity, contain separate antibodies reacting with alanyl-tRNA synthetase and tRNA(Ala). We show here that the anti-RNA antibodies recognize at least six distinguishable human tRNA(Ala) species, grouped in two sequence families. We have elucidated the complete nucleotide sequence of two tRNA(Ala) species from HeLa cells that are closely related to silkworm moth tRNA(Ala), as well as the partial sequence of a third species. All three contain the anticodon IGC. No tRNAs with pyrimidine in the "wobble" position were found in the immunoprecipitate, and such species may fail to interact with the antibody.

Alanine↗
PubMed1986

Autoantibodies against alanyl-tRNA synthetase and tRNAAla coexist and are associated with myositis.

The sera of six patients with autoimmune disease, predominantly myositis with pulmonary fibrosis, contain antibodies of the PL-12 specificity. These autoantibodies react with both protein and RNA components of human cells. The protein has a subunit molecular mass of 110 kD, and the RNA comprises a group of bands in the tRNA size class. Aminoacylation experiments identify the antigens as alanyl-tRNA synthetase and its corresponding tRNAs, tRNAAla. Anti-tRNA antibody can be absorbed out without depleting antisynthetase activity, showing that the antigens are recognized independently by separable antibodies that coexist in these sera. The concurrence of separate antibodies to the two components suggests that the autoimmune response may be mounted against the charging enzyme-tRNA complex. However, the antisynthetase antibody fails to coprecipitate tRNA with the enzyme, suggesting that the antibody reacts with its target only when it is not complexed with tRNA.

Adult↗
PubMed1986

A mechanism for the control of protein synthesis by adenovirus VA RNAI.

In the absence of VA RNAI, protein synthesis in adenovirus-infected HeLa cells fails because of defective initiation. Earlier work showed that the defect results from phosphorylation of the initiation factor elF-2 on its alpha subunit. We have identified the protein kinase responsible as the dsRNA-activated inhibitor of protein synthesis (DAI). DAI is present in uninfected state. It is activated in cells infected with the adenovirus mutant Ad5 dl331, which produces no VA RNAI, but not in cells infected with wild-type virus. Activation occurs during the late phase of infection with the mutant virus, and the activator appears to be dsRNA produced by symmetrical transcription of the viral genome. VA RNAI antagonizes the activation of DAI by dsRNA, but it cannot inhibit the activity of DAI once activated. We propose a mechanism for VA RNAI action based on its partially double-stranded nature.

Adenoviruses, Human↗
PubMed1986

Immunochemical and biochemical analysis of the proliferating cell nuclear antigen (PCNA) in HeLa cells.

PCNA, also known a cyclin, a protein of molecular weight (MW) 35 000, accumulates in the nuclei of dividing and transformed cells and reacts with autoantibodies from certain lupus patients. Using an indirect immunofluorescence technique, we show that lupus sera containing anti-PCNA antibodies reveal a heterogeneous nuclear fluorescence pattern upon reaction with asynchronous HeLa cells, whereas with synchronized cells a sequence of distinct patterns is disclosed on progression through the cell cycle. Cell-free translation of HeLa cell mRNA followed by immunoprecipitation with anti-PCNA sera shows a single protein with the same apparent MW as PCNA labelled in vivo, suggesting that PCNA is not derived from a larger precursor protein and not grossly modified by post-translational events. However, a group of at least nine nuclear polypeptides ranging in MW from about 12 000 to 110 000 are recognized by immunoblotting with anti-PCNA sera, indicating either that additional antigenic sites are produced on denaturation of native proteins or that additional autoantibodies are present in these sera. We also show that PCNA and several of these polypeptides are associated with nuclear structures containing chromatin.

Antigens↗
PubMed1986

Lytic and transforming functions of individual products of the adenovirus E1A gene.

To distinguish the individual roles of the 13S, 12S, and 9S adenovirus E1A gene products, we isolated the corresponding cDNA clones and recombined them into both plasmids and viruses. Only the expected E1A mRNA products were made from the corresponding 12S and 13S viruses. The 9S mRNA was detected when the 9S virus was coinfected with the 13S virus but not when either virus was infected alone. The 13S virus formed plaques equally well in 293 cells, HeLa cells, and A549 cells, a human lung oat cell carcinoma line. Plaque titers of the 12S virus were much reduced in HeLa and A549 cells compared with 293 cells, although the 12S virus is multiplicity-dependent leaky in both HeLa and A549 cells. A549 cells were significantly more permissive than HeLa cells for growth of the 12S virus. In A549 cells even at low multiplicities of infection the final yield of 12S virus eventually approached the maximum yield from 293 cells. Expression from the adenovirus early region 2 and early region 3 promoters in HeLa cells was activated in the presence of a 13S cDNA E1A region but not in the presence of a 12S E1A cDNA region. Although defective for lytic growth in HeLa cells, the 12S virus immortalized BRK cells at very high efficiency, whereas infection of these cells with 13S virus, as with wild-type E1A virus, resulted mainly in cell death. The 13S product does have an immortalization function, however, revealed in the absence of adenovirus lytic functions when a plasmid containing the E1A 13S cDNA region was transfected into BRK cells. The 9S virus failed to immortalize infected BRK cells or to interfere with focus formation when coinfected with the 12S virus.

Adenovirus Early Proteins↗
PubMed1986

Identification of separate domains in the adenovirus E1A gene for immortalization activity and the activation of virus early genes.

The transformation and early adenovirus gene transactivation functions of the E1A region were analyzed with deletion and point mutations. Deletion of amino acids from position 86 through 120 had little effect on the lytic or transforming functions of the E1A products, while deletion of amino acids from position 121 through 150 significantly impaired both functions. The sensitivity of the transformation function to alterations in the region from amino acid position 121 to 150 was further indicated by the impairment of transforming activity resulting from single amino acid substitutions at positions 124 and 135. Interestingly, conversion of a cysteine residue at position 124 to glycine severely impaired the transformation function without affecting the early adenovirus gene activating functions. Single amino acid substitutions in a different region of the E1A gene had the converse effect. All the mutants produced polypeptides of sufficient stability to be detected by Western immunoblot analysis. The single amino acid substitutions at positions 124 and 135, although impairing the transformation functions, did not detectably alter the formation of the higher-apparent-molecular-weight forms of the E1A products.

Adenovirus Early Proteins↗
PubMed1986

The SL autoantibody-antigen system: clinical and biochemical studies.

A recently described autoantibody, SL, was found in serum from 27 patients with autoimmune disease, including 20 with systemic lupus erythematosus (SLE) where the frequently was 7%. Analysis of clinical, serological, and HLA data from 119 SLE patients showed no positive associations with anti-SL antibody apart from a higher frequency of non-infective fever. Most SL positive sera contained other precipitins, notably antibodies to Ro(SS-A) and the proliferating cell nuclear antigen, PCNA. Anti-SL IgG recognised a protein of 32 000 daltons without associated RNA. This polypeptide was distinguished from a similarly sized component of the Sm and RNP ribonucleoprotein particles by demonstrating different products of partial proteolysis. Although anti-SL antibody is of limited clinical importance, it occurs with twice the frequency of anti-SM antibody in white patients with SLE. Preliminary studies indicate that SL and the Japanese Ki system are identical.

Antibodies, Antinuclear↗
PubMed1985

Translational control by adenovirus: lack of virus-associated RNAI during adenovirus infection results in phosphorylation of initiation factor eIF-2 and inhibition of protein synthesis.

The dl331 mutant of adenovirus serotype 5 fails to produce virus-associated (VA) RNAI, and cells infected with this mutant do not synthesize proteins efficiently at late times in infection. The translational defect occurs at the level of polypeptide chain initiation, and cell-free extracts prepared from dl331-infected cells exhibit the defect observed in vivo. Addition of either eukaryotic initiation factor 2 (eIF-2) or guanine nucleotide exchange factor (GEF) to these cell-free extracts restores translational activity, with GEF functioning more efficiently in this regard. These results suggest that cells infected with the dl331 mutant develop a translational block at the level of GEF-catalyzed guanine nucleotide exchange and that this block is most likely established through phosphorylation of the alpha subunit of eIF-2. In the present investigation we show that endogenous HeLa cell GEF activity is significantly reduced in cells infected with the dl331 mutant. Further, in contrast to cells infected with wild-type serotype 2 adenovirus, dl331-infected cells contain increased eIF-2 alpha kinase activity. These results indicate that VA RNAI plays a role in suppressing eIF-2 alpha kinase activity during adenovirus infection of HeLa cells.

Adenoviridae↗
PubMed1985

Characterization and purification of lupus antigen La, and RNA-binding protein.

HeLa cell La antigen, an RNA-binding protein, was characterized by using two-dimensional gel electrophoresis. Eight isoelectric forms (pI 6 to 7) were observed, many containing phosphate. An in vitro translation product similar in size and antigenicity was identified. The HeLa cell protein purified by using an assay based on ribonucleoprotein reconstitution with adenovirus VA RNAI also comprised several isoelectric forms.

Antigens↗
PubMed1984

Anti-threonyl-tRNA synthetase, a second myositis-related autoantibody.

An autoantibody known as PL-7 was found in the serum of four patients with myositis and one with a systemic lupus erythematosus-like syndrome. The PL-7 antigen is an 80,000 dalton polypeptide that coprecipitates with transfer RNA. In aminoacylation reactions, PL-7 IgG inhibited the charging of tRNA with threonine but had little or no effect on charging with other amino acids. Experimental antibodies raised against purified threonyl-tRNA synthetase recognized the same 80,000 dalton polypeptide, but tRNA was not coprecipitated. We conclude that PL-7 antibody is directed at threonyl-tRNA synthetase, and that different antigenic sites are recognized by the human and experimental autoantibodies. Our findings emphasize the link between myositis and autoimmunity to tRNA-related structures.

Adult↗
PubMed1984

Anti-Jo-1 antibody: a marker for myositis with interstitial lung disease.

An autoantibody known as anti-Jo-1 antibody is found in 25% of patients with myositis. Its prevalence in patients with both myositis and cryptogenic fibrosing alveolitis was 68% (13 out of 19 patients), compared with 7.5% in patients with myositis alone (four of 53) and 3% in patients with cryptogenic fibrosing alveolitis alone (two of 62). Anti-Jo-1 antibody may be useful in indicating patients with myositis and cryptogenic fibrosing alveolitis. Raynaud's phenomenon, the sicca syndrome, and mild arthritis are also often part of the syndrome.

Autoantibodies↗
PubMed1984

Alterations of transcription and translation in HeLa cells exposed to amino acid analogs.

Amino acid analogs, like other effectors of the stress response, induce in mammalian cells the same gene products that are induced upon heat shock; incorporation of the analog into protein is required for induction. We show here that induction by analogs involves controls operating at the levels of both transcription and translation. The electrophoretic patterns of newly made mRNAs simplify with time such that the putative stress protein mRNAs are the only species transported from the nucleus. Concomitantly, the patterns of protein synthesis simplify such that the stress proteins become nearly exclusive polypeptide products. Although the normal mRNAs are either not used or used with greatly reduced efficiency, they are not degraded and retain translatability when transferred to cell-free systems. Soon after the stress response has been induced, there follows a defect in the initiation of polypeptide chains, as evidenced by examination of polysome profiles. Upon prolonged exposure, polysomes are recovered, and although they give rise to stress proteins almost exclusively, the normal mRNAs are still present in these structures. Thus, in addition to the initiation defect, a lesion in elongation may also be involved. The extreme sensitivity of protein synthesis to the inhibition of RNA synthesis, together with the parallel simplifications in the patterns of newly made mRNAs and polypeptides, may imply that only newly made mRNAs are efficiently translated in analog-treated cells.

Azetidinecarboxylic Acid↗
PubMed1984

La antigen recognizes and binds to the 3'-oligouridylate tail of a small RNA.

The La antigen is a cellular protein which interacts with many RNA species that are products of RNA polymerase III, including the adenovirus virus-associated (VA) RNAs. We demonstrate that the efficiency of antigen binding in vitro is determined by the number of U residues at the RNA 3' terminus. Forms of VA RNAI with more than two terminal U residues are fully bound, forms with two U residues are partially bound, and forms with fewer than two U residues are not bound at all. The antigen can be covalently linked to VA RNA by UV irradiation, and the site of cross-linking is shown to contain the 3' terminus of the RNA. We conclude that the antigen recognizes the U-rich 3' tail of VA RNA, and presumably that of other polymerase III products, and that it binds at or close to this site.

Adenine Nucleotides↗
PubMed1984

Cellular protein and RNA antigens in autoimmune disease.

Antibodies directed against soluble cellular antigens are a distinctive feature of systemic autoimmune disease. We have examined 22 autoantibodies in sera from 1111 patients and present the disease associations together with a biochemical analysis of the antigens. The data emphasize the clinical specificity of the antibodies and the restricted number of cellular components that commonly elicit an immune response. In several instances, serological relationships between antibodies mirror biochemical relationships between the corresponding antigens. The antigens are mainly proteins and are often present in complexes with additional protein or nucleic acid molecules. In myositis the antibodies react chiefly with cytoplasmic antigens such as aminoacyl-tRNA synthetases, in contrast to the mainly antinuclear response in SLE. It is argued that both environmental stimuli and genetic factors govern autoantibody specificity, and that molecular characterization of the cellular antigens may yield clues to the aetiology of the disease and of the concomitant, specific autoimmune response.

Antibody Specificity↗
PubMed1983

Structural characterization of the proteins encoded by adenovirus early region 2A.

Proteins encoded by adenovirus type 2 and type 5 early region 2A isolated from infected HeLa cells were compared to translation products of E2A-specific messenger RNA in a reticulocyte cell-free system and in Xenopus oocytes. The main cell-free translation product is a 72,000 Mr polypeptide which in HeLa cells as well as in Xenopus oocytes is converted into a 75,000 Mr phosphoprotein capable of binding to single-stranded DNA. Some minor proteins are proteolytic cleavage products of the major protein. In the cell-free system, three E2A polypeptides, 32,000, 37,000 and 44,000 Mr, are translated from minor polyadenylated mRNA species that can be separated from the major mRNA. Synthesis of all E2A polypeptides in vitro is inhibited by cap-analogs. The 44,000 Mr protein is also synthesized in Xenopus oocytes. Tryptic peptide maps of [35S]methionine-labeled E2A proteins were constructed using high pressure liquid chromatography and the position of the methionyl residues within each peptide was determined by amino acid sequencing procedures. This information and the DNA sequence of the adenovirus 5 E2A gene published by Kruijer et al. (1981) were used to align the peptides and to construct a map of the E2A proteins. Our data demonstrate that the major 75,000 Mr protein is coded for by a leftward reading frame of 529 amino acid residues located between 62 and 66 map units. The data also map six sites as targets for proteolytic enzymes. The minor E2A translation products have the same carboxy terminus as the major protein. The initiation codons of the 44,000, 37,000 and 32,000 Mr polypeptides probably correspond to amino acids 170, 243 or 244 and 290 of the major protein. Some functional properties of the major E2A protein are shared by the minor proteins and thus could be mapped. Major sites of phosphorylation, the region involved in binding to single-stranded DNA and the antigenic regions recognized by immune sera are located between amino acid residues 50 to 120, 170 to 470 and 170 to 240, respectively.

Adenoviruses, Human↗