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G V Shpakovskiĭ

Publications and source records attributed to G V Shpakovskiĭ.

At least 19 recordsLinked to original sources
PubMed2000

[Chromosomal localization of rpb9+ and tfa1+ genes, coding for components of the mRNA synthesis apparatus of Schizosaccharomyces pombe].

Using DNA hybridization on cosmid filters of high density, we established chromosomal localization of the rpb9+ gene encoding one of the specific subunits of RNA polymerase II of Schizosaccharomyces pombe and thus filled in the last gap in the mapping of the genes encoding components of RNA polymerase II of the fission yeast. The primary structure of three extended regions of the Sz. pombe chromosome I was elucidated and, as a result, genes neighboring on rpb9+ were identified. One of them proved to be the tfa1+ gene, encoding the large (alpha) subunit of the general factor of transcription initiation TFIIE.

Amino Acid Sequence↗
PubMed1999

[Molecular cloning of some components of the translation apparatus of fission yeast Schizosaccharomyces pombe and a list of its cytoplasm ic proteins genes].

Full-length cDNAs of four new genes encoding cytoplasmic ribosomal proteins L14 and L20 (large ribosomal subunit) and S1 and S27 (small ribosomal subunit) were isolated and sequenced during the analysis of the fission yeast Schizosaccharomyces pombe genome. One of the Sz. pombe genes encoding translation elongation factor EF-2 was also cloned and its precise position on chromosome I established. A unified nomenclature was proposed, and the list of all known genetic determinants encoding cytoplasmic ribosomal proteins of Sz. pombe was compiled. By now, 76 genes/cDNAs encoding different ribosomal proteins have been identified in the fission yeast genome. Among them, 35 genes are duplicated and three homologous genes are identified for each of the ribosomal proteins L2, L16, P1, and P2.

Amino Acid Sequence↗
PubMed1999

[Characteristics of the cDNA of the Schizosaccharomyces pombe rpa43+ gene: structural similarity of the Rpa43 subunit of RNA-polymerase I with the Rpc25 subunit of RNA-polymerase III].

We isolated and characterized full-length cDNA of the rpa43+ gene encoding one of subunits of the nuclear RNA polymerase I of Schizosaccharomyces pombe. The gene contains two introns and is located on chromosome II. Comparison of the primary structure of the subunit Rpa43 of Sz. pombe (173 aa; M 19,385 Da; pl 5.36), deduced from the cDNA obtained, with the amino acid sequences of subunits A43 from Saccharomyces cerevisiae and Drosophila melanogaster demonstrates a high divergence of this protein in evolution. A comparison of the Rpa43 with other proteins from the SwissProt database revealed a similarity of this subunit to subunit Rpc25 of RNA polymerase III, which, as was shown previously, is structurally similar to subunit Rpb7 of RNA polymerase II. Thus, including the Rpa43<-->Rpc25<-->Rpb7 family, nuclear RNA polymerases I-III contain at least 11 identical and/or similar subunits. This fact illustrates a pronounced resemblance of the organization of all three enzymes of the eukaryotic transcription apparatus. Moreover, at least ten out of these eleven families of eukaryotic RNA polymerase subunits have homologues in the 13-subunit archaeal RNA polymerase.

Amino Acid Sequence↗
PubMed1999

[Molecular evolution and structure of eukaryotic nuclear RNA polymerase subunits in light of the exon-intron organization of their genes].

Analysis of literary data (for Saccharomyces cerevisiae, Caenorhabditis elegans, Arabidopsis thaliana, Homo sapiens, and some other Eucarya) and our data (for Schizosaccharomyces pombe) on the exon-intron organization of the genes encoding subunits of nuclear RNA polymerases showed that introns in the orthologous genes from different organisms are arranged nonrandomly, namely, their positions, if projected on the map of the comparison of the amino acid sequences of the orthologous subunits, not infrequently coincide in evolutionarily distant species. As a rule, intron positions correspond to the boundaries of the structurally conserved regions (domains) or to the sites of possible turns of the polypeptide chain. For example, introns flank the secondary structure elements in the Rpb8 subunit with the known three-dimensional structure or the structure-function modules in subunits Rpb10 and Rpc10. These facts are in agreement with the idea of the ancient origin of introns, and with the notion of evolution of ancient protein sequences through the assembly of their genes from short protoexons selected by the nature as far back as the RNA world times. Comparative analysis of the primary structures of the subunits of eukaryotic RNA polymerases allowed us to reveal a nuclear localization signal in subunit Rpb10 and some hypothetical archaeal homologues of subunit Rpc10.

Amino Acid Sequence↗
PubMed1998

[Molecular cloning and characteristics of rpc19+ and rpc40+ Schizosaccharomyces pombe genes, coding for common subunits of nuclear RNA polymerase I and III].

Full-length copies of cDNAs of the rpc19+ and rpc40+ genes encoding the common subunits of nuclear RNA polymerases I and III and the corresponding fragments of chromosomes were isolated from genomic and cDNA libraries of Schizosaccharomyces pombe and characterized. It was established that the cloned genes are located on chromosomes III and II of the fission yeast, respectively. The rpc40+ gene lacks introns, and the rpc19+ gene contains two intervening sequences. The comparison of subunits Rpc19 (125 aa; M 13 722 Da; pI 4.51) and Rpc40 (348 aa; M 39 141 Da; pI 5.40) of Sz. pombe, whose characteristics were deduced from the sequences of their cDNAs, with the orthologous components of other eukaryotes allowed the most conserved structure-functional domains of these proteins to be identified.

Amino Acid Sequence↗
PubMed1998

[Structural-functional characteristics of the Schizosaccharomyces pombe rpb8+ gene, coding the subunit of RNA polymerase I-III, specific only for eukaryotes].

A full-length cDNA of the rpb8+ gene encoding a common subunit Rpb8 of nuclear RNA polymerases I-III only specific for Eucarya was isolated from an expression library of the fission yeast Schizosaccharomyces pombe. The primary structure of the corresponding fragment of the Sz. pombe genome was also established. The rpb8+ gene contains two short introns, 59 and 48 bp long. Only short segments of homology were found upon comparing the Rpb8 subunit homologs from various eukaryotic species, and substantial differences exist between the corresponding proteins of unicellular and multicellular organisms. Subunit Rpb8 of Sz. pombe proved to be the smallest one among the known related proteins: it lacks the 21-aa fragment corresponding to amino acids residues 68-88 of the central part of the homologous subunit ABC14.5 of Saccharomyces cerevisiae. Accordingly, subunit Rpb8 of the fission yeast was not capable of substituting in vivo subunit ABC14.5 in nuclear RNA polymerases of the baker's yeast.

Amino Acid Sequence↗
PubMed1998

[Exon-intron structure of the fet5+ gene of Schizosaccharomyces pombe and physical mapping of genome encompassing regions].

Plasmid pYUK3 bearing the fet5+ gene of Schizosaccharomyces pombe was isolated from a genomic library of the fission yeast, and a detailed physical map of the whole genomic insert (ca. 9.6 Kbp) was constructed. The primary structure of the fet5+ gene and its flanking regions is established. The gene contains a single 45-bp intron in its distal part. A typical TATA-box (TATAAG) was found in the 5'-noncoding region ca. 50 bp upstream of the putative start of transcription, and the 3'-noncoding region contains AT-rich palindromes, which are probably involved in termination of the fet5+ transcription. A previously unidentified gene of Sz. pombe encoding a protein with some similarity to one of the transcriptional activators from the TBP (TATA-binding protein) group of SPT factors of transcription was found in the vicinity of the fet5+ gene. Taking into account that cDNA of the fet5(+)-gene was isolated as a suppressor of the genetic-defect of nuclear RNA polymerases I-III (Bioorg. Khim., 1997, vol. 23, No 3, pp. 234-237), this vicinity may be the first evidence of possible clustering, in the genome of the fission yeast, of genes participating in transcription regulation.

Alternative Splicing↗
PubMed1997

[Cloning of cDNA for RNA polymerase subunit from the fission yeast Schizosaccharomyces pombe by heterospecific complementation in Saccharomyces cerevisiae].

The rpb10 cDNA of the fission yeast Schizosaccharomyces pombe, encoding one of the five small subunits common to all three nuclear DNA-dependent RNA polymerases, was isolated from an expression cDNA library by two independent approaches: PCR-based screening and direct suppression by means of heterospecific complementation of a temperature-sensitive mutant defective in the corresponding gene of Saccharomyces cerevisiae. The cloned Sz. pombe cDNA encodes a protein Rpb10 of 71 amino acids with an M of 8,275 Da, sharing 51 amino acids (71% identity) with the subunit ABC10 beta of RNA polymerases I-III from S. cerevisiae. All eukaryotic members of this protein family have the same general organization featuring two highly conserved motifs (RCFT/SCGK and RYCCRRM) around an atypical zinc finger and an additional invariant HVDLIEK motif toward the C-terminal end. The last motif is only characteristics for homologs from eukaryotes. In keeping with this remarkable structural conservation, the Sz. pombe cDNA also fully complemented a S. cerevisiae deletion mutant lacking subunit ABC10 beta (null allele rpb10-delta 1::HIS3).

Amino Acid Sequence↗
PubMed1997

[The first member of a novel family of eukaryotic transcription factors detected by the heterospecific complementation].

The cDNA of a previously uncharacterized gene fet5 (factor of eukaryotic transcription, clone no. 5) of the fission yeast Schizosaccharomyces pombe was cloned by the heterospecific complementation of a conditional mutant of Saccharomyces cerevisiae defective in the function of the RNA polymerases I-III common subunit ABC10 beta. The gene encodes a new factor of eukaryotic transcription, Fet5, the first member of a superfamily of proteins for which the area of functioning is determined. The Fet5-superfamily consists of three distinct families of proteins highly evolutionarily conserved and widely spread among eukaryotes. Features of the Fet5 amino acid sequence suggest that it belongs to ATP/GTP-binding proteins.

Amino Acid Sequence↗
PubMed1997

[Molecular cloning and characterization of cDNA of the rpc10+ gene encoding the smallest subunit of nuclear RNA polymerases of Schizosaccharomyces pombe].

The full-length cDNA of the rpc10+ gene encoding mini-subunit Rpc10, which is common for all three nuclear RNA polymerases of the fission yeast Schizosaccharomyces pombe, was cloned and sequenced. The Rpc10 subunit of Sz. pombe and its homologs from S. cerevisiae and H. sapiens are positively charged proteins with a highly conserved C-terminal region and an invariant zinc-binding domain (Zn-finger) of a typical amino acid composition: YxCx2Cx12RCx2CGxR. Functional tests of heterospecific complementation, using tetrad analysis or plasmid shuffling, showed that the Rpc10 subunit of Sz. pombe can successfully replace the homologous ABC10 alpha subunit in nuclear RNA polymerases I-III of S. cerevisiae.

Amino Acid Sequence↗
PubMed1996

[Three regions of Rpb10 mini-subunit of nuclear RNA polymerases are strictly conserved in all eukaryotes].

The rpb10+ cDNA from the fission yeast Schizosaccharomyces pombe was cloned using two independent approaches (PCR and genetic suppression). The cloned cDNA encoded the Rpb10 subunit common for all three RNA polymerases. Comparison of the deduced amino acid sequence of the Sz. pombe Rbp10 subunit (71 amino acid residues) with those of the homologous subunits of RNA polymerases I, II, and III from Saccharomyces cerevisiae and Home sapiens revealed that heptapeptides RCFT/SCGK (residues 6-12), RYCCRRM (residues 43-49), and HVDLIEK (residues 53-59) were evolutionarily the most conserved structural motifs of these subunits. It is shown that the Rbp10 subunit from Sz. pombe can substitute its homolog (ABC10 beta) in the baker's yeast S. cerevisiae.

Amino Acid Sequence↗
PubMed1989

[Structure of the recombination zone during abnormal excision of the transducing bacteriophage lambda plac9].

Investigating molecular mechanism of illegitimate recombinations in prokaryote we study transducing bacteriophages of the lambda lac series. We have carried out physical mapping of bacteriophage lambda plac9 DNA and, by comparing the obtained results with the data on the structure of lambda DNA and lac operon of E. coli, located the phage-bacterial junction corresponding to the lambda-lac9 abnormal excision and elucidated the nucleotide sequence around the junction. It led to the primary structure of phage and bacterial segments in the lysogenic bacterium which took part in the recombinational act leading to the abnormal excision and lambda lac9 formation. Structural homology of the partners in the lambda plac9 excision proved to be lower than in case of the earlier studied lambda plac5 and lambda plac10 whose excision proceeded regioselectively. Various aspects of the crossover area, including the crossover point's probable position and enzymic systems participating in the abnormal excision, are discussed.

Bacteriophage lambda↗
PubMed1986

[Complete primary structure of DNA from the transducing bacteriophage lambda plac5].

In studying molecular mechanisms of specialized transduction, primary structure of the junction between the E. coli gene lacI and the lambda phage locus Ea47 in transducing bacteriophage lambda plac5 has been established. Along with the lambda DNA and E. coli lac operon structures as well as with our earlier data on another phage-bacterial junction in lambda plac5, it lead to the complete sequence of lambda plac5 DNA, including the lac5 substitution, a wellknown segment of lambdoid cloning vehicles. The lambda plac5 DNA is shown to consist of 48645 b.p. distributed as follows: 19368 (lambda left arm) + 3924 (lac5 substitution) + 25353 (lambda right arm). The presence of the phage pbL promoter near to the right end of the lac5 insert is shown. The lacI gene distal end in lambda plac5 proved to be considerably more long-stretched than it used to be believed, coding for 224 C-terminal amino-acid residues of lac repressor. The recombination studied in this paper, similarly to the abnormal prophage excision, occurred near to a Chi-like structure, which is partly homologous to the chi+lacZ site present in lambda plac5. On the basis of the data obtained, a key role of the E. coli RecBC system and Chi sites in the formation of long-stretched deletions in the bacterial cell has been suggested.

Bacteriophage lambda↗
PubMed1983

[Structure of a recombination site in the transducing bacteriophage lambda plac5 DNA].

A recombination site in the transducing bacteriophage lambda plac5 DNA has been structurally elucidated. Comparison of primary structures of E. coli lac-operon (distal end of lacZ gene, Z-Y spacer, and proximal end of lacY gene) described earlier with corresponding segments of bacteriophages lambda CI857 and lambda plac 5-2 DNAs sequenced in this paper showed that the bacterial DNA insert ends immediately after Z-Y spacer, just before the initiating triplet ATG of lacY gene. It thus follows that in contrast to the earlier conception, the insert does not seem to include any part of lacY gene. The recombination sites in both phage and bacterial DNA contain structurally homological segments about 20 b. p. long (crossover region), with two extra basepairs in the bacterial DNA (AT in the sense-strand). We suppose that the very dinucleotide plays a substantial role in initiation of recombinational event: causing formation of a nonperfect heteroduplex structure, it determines the T-A internucleotide bond to be endonucleolytically cut (crossover point) followed by exonucleolytic elimination of the extra links (AT) and reciprocal strand exchange. The second recombination site in lambda plac5 DNA has been localized by us within lacI gene as being close to the HindII site (nucleotides 854 to 859 of the gene). The structures of the two regions of site-specific recombination may shed light upon mechanisms of the phage abnormal excision leading to formation of transducing phages.

Bacteriophage lambda↗
PubMedDate not supplied

[New genes on human chromosome 7: bioinformatic analysis of a gene cluster from the POLR2J family].

Four independent genes encoding various variants of the hRPB11 subunit of Homo sapiens RNA polymerase II were revealed in human chromosome 7. Three genes (POLR2J1, POLR2J2, and POLR2J3) form a cluster of total length 214530 bp in the genetic locus 7q22.1 on the long arm of chromosome 7 (contig NT_007933). The fourth gene (POLR2J4, 31040 bp) was localized in the cytogenetic locus 7p13 of the short arm of chromosome 7 (contig NT_007819). An analysis enabled us to refine dissimilar experimental data on the mapping of the hRPB11 subunit gene on chromosome 7. In particular, the presence of three sites of its localization according to data on hybridization with fluorescent-labeled probes (the FISH method) was explained. It was established that, upon the expression of the four human POLR2J genes, at least 14 types of mature mRNAs encoding somewhat differing hRPB11 isoforms can be synthesized. Eleven of these mRNAs were revealed (as full-length copies or clearly identifiable fragments) in the available databases of expressed sequence tags and cDNAs. The most probable scheme of origination of the multiple genes of the POLR2J family, as a result of three consecutive segmented duplications increasing in size, was proposed and substantiated. On the basis of the scheme, some assumptions on the pathways of evolution of separate human genes and the mechanisms of generation of protein diversity in higher eukaryotes were made. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2004, vol. 30, no. 6; see also http://www.maik.ru.

Chromosome Mapping↗