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M Mieszczak

Publications and source records attributed to M Mieszczak.

7 recordsLinked to original sources
PubMed1992

Multiple plant RNA binding proteins identified by PCR: expression of cDNAs encoding RNA binding proteins targeted to chloroplasts in Nicotiana plumbaginifolia.

Pre-mRNA processing in eukaryotic cells requires the participation of multiple protein factors and ribonucleoprotein particles. One class of proteins involved in this process are RNA-binding proteins, which contain a domain of ca. 90 amino acids with a characteristic ribonucleoprotein consensus sequence (RNP-CS). A PCR approach that is suitable for the characterization of RNP-CS-type proteins is described. Fifteen different RNA-binding domains were amplified from Nicotiana tabacum (tobacco) using oligonucleotide primers specific for the sequences (K/R)G(F/Y)(G/A)FVX(F/Y) and (L/I/V)(F/Y)(V/I)(G/K)(N/G)L, which are conserved in known RNP-CS proteins. Using the tobacco domains as probes, cDNAs encoding two RNA-binding proteins, each containing two RNP-CS-type domains, were characterized in N. plumbaginifolia. The proteins, designated CP-RBP30 and CP-RBP31, are targeted to chloroplasts as demonstrated by expression of epitope-tagged cDNAs in transfected protoplasts, followed by indirect immunofluorescence. High levels of mRNA for each protein were found in leaves but not in roots, and expression of the CP-RBP31 mRNA was strongly regulated by light. The N. plumbaginifolia proteins described in this work are distinct from chloroplast RNA-binding proteins characterized recently in tobacco and spinach.

Amino Acid Sequence↗
PubMed1988

Nuclear omnipotent suppressors of premature termination codons in mitochondrial genes affect the 37S mitoribosomal subunit.

nam3 and R705, yeast nuclear omnipotent suppressors of mitochondrial mit- mutations, reverse the superimposed spectrum of trans-recessive splicing defects by affecting the protein composition of the small mitoribosomal subunit. Analysis of the suppressor's interaction suggests that suppression results from mutations in the mitoribosomal polypeptides. These data indicate an obligatory connection between mitoribosome function and splicing of introns bI2, bI4 and aI1 in yeast mitochondria.

DNA, Mitochondrial↗
PubMed1988

Protein composition of Saccharomyces cerevisiae mitochondrial ribosomes.

Protein composition of mitochondrial ribosomes of the yeast Saccharomyces cerevisiae was analysed by two-dimensional electrophoresis. The small (37S) mitoribosomal subunit contains 36 different polypeptides with molecular weights ranging from 10,000 to 60,000. The large (50S) subunit is composed of 41 proteins with molecular weights from 10,000 to 43,000. The molecular weights of mitoribosomal small and large subunits are 1.85 MDa and 2.35 MDa, respectively. Proteins represent 60-62% and 42-45% of the total mass of 37S and 50S subunits respectively. On the basis of the protein content and molecular weights of individual proteins we conclude that all mitoribosomal proteins are present in the mitoribosome in equimolar proportions.

Adenosine Diphosphate↗
PubMed1987

Phenotypic suppression and nuclear accommodation of the mit- oxi1-V25 mutation in isolated yeast mitochondria.

Phenotypic suppression by the antibiotic, paromomycin, of the mitochondrial oxi1- -V25 mutation, a mutation which arrests by premature ochre codon the synthesis of the cox II subunit, was studied in isolated yeast mitochondria competent in translation. This antibiotic is known to suppress the mutation in vivo (Dujardin et al. 1984) and allowed in vitro, at concentrations of 20-1100 micrograms per ml. the synthesis of the cox II subunit. This strongly suggests that phenotypic suppression of mit- mutations is due to the direct action of paromomycin on mitochondrial ribosomes. The effect of paromomycin bears a resemblance to the function of the omnipotent nuclear suppressor mutation R705. The nuclear suppression was expressed in isolated mitochondria; suppressor mutation influenced the structure of the mitoribosome. Therefore, it appears that mitoribosomes are indeed the common target in the phenotypical and genetic nuclear suppression of the oxi1-V25 mutation.

Cell Nucleus↗
PubMed1987

Protein synthesis in mitochondria from yeast strains carrying nam and mim suppressor genes.

Yeast mitochondria isolated from two different wild type strains (gal+ and gal-), whether grown on galactose or glucose, synthesize all mitochondrial polypeptides with similar efficiencies and in proportions approximating those detected in vivo. Mitochondria isolated from mit- mutants synthesize in vitro a mutant pattern of mitochondrial proteins, indistinguishable from the in vivo products. The mutant pattern is restored to the wild type one in mitochondria isolated from pseudorevertant strains carrying an additional nuclear (nam3-1 and R705) or mitochondrial (mim3-1) informational suppressor gene. Suppression is expressed in isolated mitochondria without the obligatory presence of cytosol at the level of both respiratory control and specific polypeptide synthesis. Translation in isolated mitochondria is sensitive to paromomycin. The antibiotic differentiates between translation in mitochondria from wild type strains and that in nam-type gene carrying strains. This strongly suggests that nam-type mutations affect the mitoribosome, enhancing ambiguity of translation, thus allowing for the pseudoreversion of mit- phenotypes.

Adenosine Diphosphate↗
PubMed1987

mim3 and nam3 omnipotent suppressor genes similarly affect the polypeptide composition of yeast mitoribosomes.

Yeast informational suppressors of mit- mutations coded for by nuclear (nam3-1, nam3-2) or by mitochondrial DNA (mim3-1) affect the mitoribosome. Nuclear mutations result in the appearance of an additional polypeptide called SI in the small mitoribosomal subunit. An identical polypeptide, not detected in the wild type 37S subunit, is present in crude preparations of mitoribosomes isolated from a mim3-1 suppressor carrying strain. Traces of the SI polypeptide may be found in highly purified small subunits from the mim3-1 strain. Therefore, mutations affecting either mitochondrial rRNA (mim3-1) or mitochondrial r-proteins (nam3-1, nam3-2) could be followed by similar changes in overall mitoribosome structure. This may explain the functional similarity of nuclear and mitochondrially coded suppressors.

Cell Nucleus↗