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M Barciszewska

Publications and source records attributed to M Barciszewska.

At least 19 recordsLinked to original sources
PubMed1999

Preparation of HIV TAR RNA with RNA scissors.

Two hammerhead ribozymes derived from plant pathogenic RNAs were used to cut off the HIV TAR RNA from the T7 RNA transcript through a cis cleavage reaction. Stem I of the (+)vLTSV ribozyme comprises 8 nucleotides of the 5' terminus of TAR RNA, but stem III of the (+)sTRSV ribozyme consists of 8 nucleotides of the 3' end of TAR RNA. The construct containing two GUC hammerhead ribozyme target sequences identified the cleavage sites to cut off a required RNA molecule. This method was applied for preparation of 35 nt long TAR RNA. Its activity was proved by the complex formation with the Tat protein. It seems that this approach based on RNA scissors can also be used for the generation of required RNA molecules, RNA decoys or RNA aptamers in vivo.

Bacteriophage T7↗
PubMed1997

Purification and characterization of transcription factor IIIA from higher plants.

Transcription factor IIIA (TF IIIA) binds and specifically activates transcription of eukaryotic 5S rRNA genes. It also forms a 7S ribonucleoprotein complex with mature 5S rRNA. Here, we describe the purification and properties of pTF IIIA from higher plants. The purified protein from tulip (Tulipa whittalii) has a molecular mass of about 40 kDa and also binds 5S rRNA and 5S rRNA genes. pTF IIIA also facilitates the transcription of a 5S rRNA gene in a HeLa cell extract.

DNA, Plant↗
PubMed1995

Glycine codon discrimination and the nucleotide in position 32 of the anticodon loop.

Using an in vitro protein-synthesizing system that allowed us to monitor separately the reading of each glycine codon, we have previously shown, that in constructs based on glycine tRNA1 from Escherichia coli the nature of the nucleotide in position 32 determines the ability of the anticodon UCC to discriminate between the glycine codons. Thus, with a U in position 32 the anticodon UCC discriminated according to the wobble rules, but with a C in this position it had lost its ability to discriminate. In the present paper we show that the same is true also for constructs based on mycoplasma glycine tRNA. When C32 in the wild type was changed to U32, the anticodon UCC discriminated between the glycine codons, while in wild type mycoplasma glycine tRNA it did not. Furthermore, when U32 was changed to C32 in glycine tRNA1(CCC), the anticodon CCC loses its ability to discriminate. We therefore conclude that the nature of the nucleotide in position 32 determines the discriminatory ability of both anticodons UCC and CCC in the glycine tRNA1 structural background, and that the same is true for the anticodon UCC in the mycoplasma glycine tRNA background.

Anticodon↗
PubMed1995

Are 5S ribosomal RNA primary transcripts edited?

Comparative analysis of the nucleotide sequences of plant 5S rRNA genes shows numerous discrepancies with the mature 5S rRNAs. Detailed inspection of these differences revealed that they are mainly due to single nucleotide substitutions, insertions and deletions. In several cases also longer insertions were found, two of which are particularly interesting. Their location in loops resembles tRNA introns. One of these insertions is 79 nucleotides long and can potentially form a tRNA-like structure, with 50% homology to plant tRNAGly. This observation led us to propose that, in the maturation of some 5S rDNA transcripts, editing and splicing mechanisms may be involved.

Base Sequence↗
PubMed1995

Purification and properties of the first specific 5S rRNA binding protein from plants which shows transcription factor IIIA activity.

Eukaryotic transcription factor IIIA (TF IIIA) is know to activate specifically transcription of 5S rRNA gene. It also interacts with mature 5S rRNA. The best known TF IIIA has been purified from Xenopus laevis. Up to know this protein has not been isolated from plants. In this paper we show for the first time purification and properties of TF IIIA like protein from tulip (Tulipa whittalli). It shows 5S rRNA and 5S rRNA gene binding activity.

Animals↗
PubMed1993

The nucleotide in position 32 of the tRNA anticodon loop determines ability of anticodon UCC to discriminate among glycine codons.

We have investigated the influence of structures in the tRNA anticodon loop and stem on the ability of the anticodon to discriminate among codons. We had previously shown that anticodon UCC, when placed in the structural context of tRNA(Gly1) from Escherichia coli, discriminated efficiently between the glycine codons, as required by the wobble rules. Thus, this anticodon read GGA and GGG but did not read GGU and GGC, whereas in mycoplasma tRNA(Gly), the same anticodon did not discriminate among the glycine codons. We have now determined the reading properties of three constructions based on tRNA(Gly1) containing the anticodon UCC in different structural contexts. In one of these constructs, tRNA(Gly1-ASL), the anticodon loop and stem are the same as in mycoplasma tRNA(Gly). The second construct, tRNA(Gly1-AS), has an anticodon stem identical with the mycoplasma tRNA(Gly), whereas in the last construct, tRNA(Gly1-C32), the only difference from tRNA(Gly1)(UCC) is that the uridine in position 32 of the anticodon loop has been replaced by cytidine. These constructs were tested for ability to read glycine codons in an in vitro protein-synthesizing system that allowed us to monitor separately the reading of each codon. We found that the anticodon UCC, when present in tRNA(Gly1-AS), discriminated among the glycine codons, whereas in the constructs tRNA(Gly1-ASL) and tRNA(Gly1-C32), the same anticodon had lost its ability to discriminate--i.e., it behaved as in mycoplasma tRNA(Gly). These results strongly suggest that nt 32 of the anticodon loop of tRNA(Gly1)(UCC) decisively influences the reading properties of the anticodon UCC.

Anticodon↗
PubMed1993

Undiscriminating codon reading with adenosine in the wobble position.

To investigate the reading properties of adenosine in the wobble position we have used site-directed mutagenesis of the Escherichia coli glycine tRNA1(CCC) gene to substitute the nucleotide A in the wobble position of the corresponding tRNA. The effect of this change on the ability of the tRNA to discriminate between the nucleotides in the third position of the glycine codons has been investigated. We have compared the ability of the mutant glycine tRNA1(UCC) and glycine tRNA1(ACC) as well as the mycoplasma glycine tRNA(UCC) to read the glycine codons. The results showed that glycine tRNA1(ACC) unlike glycine tRNA1(UCC) did not fully discriminate between the glycine codons. These experiments were carried out using a new in vitro protein synthesizing system that allows us to monitor the reading of all four glycine codons. In the present paper we give a detailed description of this new in vitro system.

Adenosine↗
PubMed1990

The primary structure of six leucine isoacceptor tRNAs of yellow lupin seeds. The structural requirements for amber tRNA suppressor activity.

Six tRNA(Leu) isoacceptors from yellow lupin seeds were purified, sequenced, and their readthrough properties over the UAG stop codon were tested using TMV RNA as a messenger. The tested tRNAs(Leu) did not show amber suppressor activity. The partial structure of tRNA(Gln), a minor species in yellow lupin, was also determined. Comparison of the nucleotide sequence of all known isoacceptors of tRNA(Tyr), tRNA(Gln) and tRNA(Leu) from plants, mammals and ciliates enabled us to find general structural requirements for tRNA to be a UAG suppressor. From the partial sequence of lupin tRNA(Gln) we suggest that it will have readthrough properties.

Autoradiography↗
PubMed1990

Higher plant 5S rRNAs share common secondary and tertiary structure. A new three domains model.

A new model of secondary and tertiary structure of higher plant 5S RNA is proposed. It consists of three helical domains: domain alpha includes stem I; domain beta contains stems II and III and loops B and C; domain gamma consists of stems IV and V and loops D and E. Except for, presumably, a canonical RNA-A like domain alpha, the two remaining domains apparently adopt a perturbed RNA-A structure due to irregularities within internal loops B and E and three bulges occurring in the model. Bending of RNA could bring loops B and E and/or C and D closer making tertiary interactions likely. The model differs from that suggested for eukaryotic 5S rRNA, by organization of domain gamma. Our model is based on the results of partial digestion obtained with single- and double-strand RNA specific nucleases. The proposed secondary structure is strongly supported by the observation that crude plant 5S rRNA contains abundant RNA, identified as domain gamma of 5S rRNA. Presumably it is excised from the 5S rRNA molecule by a specific nuclease present in lupin seeds. Experimental results were confirmed by computer-aided secondary structure prediction analysis of all higher plant 5S rRNAs. Differences observed between earlier proposed models and our proposition are discussed.

Base Sequence↗
PubMed1989

The nucleotide sequence of ribosomal 5S RNA from Rhizobium meliloti: comparison with 5S rRNA of Agrobacterium.

The complete nucleotide sequence of R. meliloti 5S ribosomal RNA has been determined and compared with the already known sequence of A. tumefaciens 5S rRNA (Vandenberghe et al., 1985, Eur. J. Biochem., 149, 537-542) and of other 5S rRNAs from Rodobacteria Alpha-2 (Wolters et al., 1988, Nucleic Acids Res., 16, rl-r70). The differences found at eight positions (23, 73, 83, 72 in helical fragments; 16, 40, 88 in loops; 54 in bulge), which might affect secondary structures of 5S rRNA, are small. Moreover, the sequence analysis specifies both variable and common positions in 5S rRNA secondary structure of Rodobacteria Alpha-2.

Base Sequence↗
PubMed1988

Yellow lupin cytoplasmic tRNAGlu is not a cofactor in chlorophyll biosynthesis.

Yellow lupin seeds (Lupinus luteus) cytoplasmic tRNAGlu was isolated and the primary structure was determined to be: (sequence in text) AGU CCCGGCGACGGAACCAOH. It is 76 nucleotides long and contains 8 modified nucleosides: 2 residues of pseudouridine, ribothymidine, 3 dihydrouridines, 5-methylcytosine and 1-methyladenosine. This tRNAGlu assayed in delta-aminolevulinic acid synthesis was shown to be inactive. Its structural features are discussed.

Base Sequence↗
PubMed1984

UAG readthrough during TMV RNA translation: isolation and sequence of two tRNAs with suppressor activity from tobacco plants.

The hypothetical replicase or replicase subunit cistron in the 5'-proximal part of tobacco mosaic virus (TMV) RNA yields a major 126-K protein and a minor 183-K ;readthrough' protein in vivo and in vitro. Two natural suppressor tRNAs were purified from uninfected tobacco plants on the basis of their ability to promote readthrough over the corresponding UAG termination codon in vitro. In a reticulocyte lysate the yield of 183-K readthrough protein increases from 10% in the absence of added tobacco plant tRNA up to 35% in the case of pure tRNA added. Their amino acid acceptance and anticodon sequence (GpsiA) identifies the two natural suppressor tRNAs as the two normal major cytoplasmic tyrosine-specific tRNAs. tRNA(1) has an A:U pair at the base of the TpsiC stem and an unmodified G(10), whereas tRNA(2) contains a G:C pair in the corresponding location and mG in position 10. This is the first case that, in a higher eukaryote, the complete structure is known of both the natural suppressor tRNAs and the corresponding viral RNA on which they exert their function. The corresponding codon-anticodon interaction, which is not in accordance with the wobble hypothesis, and the possible biological significance of the readthrough phenomenon is discussed.

Journal Article↗
PubMed1984

The molecular basis for the differential translation of TMV RNA in tobacco protoplasts and wheat germ extracts.

Translation of tobacco mosaic virus (TMV) RNA in tobacco protoplasts yields the 17.5-K coat protein, a 126-K protein and a 183-K protein which is generated by an efficient readthrough over the UAG termination codon at the end of the 126-K cistron. In wheat germ extracts, however, only the 5'-proximal 126-K cistron is translated whereas the 183-K readthrough protein is not synthesized. Purification and sequence analysis of the endogenous tyrosine tRNAs revealed that the uninfected tobacco plant contains two tRNAs, both with GPsiA anticodons which stimulate the UAG readthrough in vitro and presumably in vivo. In contrast, 85% of the tRNA from wheat germ contains a QPsiA anticodon and 15% has a GPsiA anticodon. Otherwise the sequences of tRNAs from wheat germ and tobacco are identical. UAG readthrough and hence synthesis of the 183-K protein is only stimulated by tRNA(GPsiA) and not at all by tRNA(QPsiA). The tRNAs from wheat leaves were also sequenced. This revealed that adult wheat contains tRNA(GPsiA) only. This is very much in contrast to the situation in animals, where Q-containing tRNAs are characteristic for adult tissues whereas Q deficiency is typical for the neoplastic and embryonic state.

Journal Article↗