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Biomedical subjects

T D Mashkova

Publications and source records attributed to T D Mashkova.

At least 19 recordsLinked to original sources

Type D retrovirus specific sequences in lymphocytes of the children with Burkitt-type lymphoma and their parents.

Type D retroviruses cause immunodeficiency in monkey. Earlier we have revealed genetical and serological markers of type D retroviruses in children with Burkitt-type lymphoma. Using PCR/Southern blotting assay we have found sequences related to MPMV in PBMC's DNA from children with Burkitt-type lymphoma and from their parents. Moreover, the data on sequencing of virus specific sequences from one ill child and from his mother have been presented.

Blotting, Southern↗

Structural rearrangements and insertions of dispersed elements in pericentromeric alpha satellites occur preferably at kinkable DNA sites.

Centromeric region of human chromosome 21 comprises two long alphoid DNA arrays: the well homogenized and CENP-B box-rich alpha21-I and the alpha21-II, containing a set of less homogenized and CENP-B box-poor subfamilies located closer to the short arm of the chromosome. Continuous alphoid fragment of 100 monomers bordering the non-satellite sequences in human chromosome 21 was mapped to the pericentromeric short arm region by fluorescence in situ hybridization (alpha21-II locus). The alphoid sequence contained several rearrangements including five large deletions within monomers and insertions of three truncated L1 elements. No binding sites for centromeric protein CENP-B were found. We analyzed sequences with alphoid/non-alphoid junctions selectively screened from current databases and revealed various rearrangements disrupting the regular tandem alphoid structure, namely, deletions, duplications, inversions, expansions of short oligonucleotide motifs and insertions of different dispersed elements. The detailed analysis of more than 1100 alphoid monomers from junction regions showed that the vast majority of structural alterations and joinings with non-alphoid DNAs occur in alpha satellite families lacking CENP-B boxes. Most analyzed events were found in sequences located toward the edges of the centromeric alphoid arrays. Different dispersed elements were inserted into alphoid DNA at kinkable dinucleotides (TG, CA or TA) situated between pyrimidine/purine tracks. DNA rearrangements resulting from different processes such as recombination and replication occur at kinkable DNA sites alike insertions but irrespectively of the occurrence of pyrimidine/purine tracks. It seems that kinkable dinucleotides TG, CA and TA are part of recognition signals for many proteins involved in recombination, replication, and insertional events. Alphoid DNA is a good model for studying these processes.

Alu Elements↗

Evidence for selection in evolution of alpha satellite DNA: the central role of CENP-B/pJ alpha binding region.

Conservation of DNA segments performing sequence-related functions is a landmark of selection and functional significance. Phylogenetic variability of alpha satellite and apparent absence of conserved regions calls its functional significance into question, even though sequence-specific alpha satellite-binding proteins pJ alpha and CENP-B have been discovered. Moreover, the function of pJ alpha is obscure and CENP-B binding satellite DNA, which is thought to participate in centromere formation, is found only in few species and not necessarily in all chromosomes. Analysis of alpha satellite evolution allows us to recognize the order in this variability. Here we report a new alpha satellite suprachromosomal family, which together with the four defined earlier, covers all known alpha satellite sequences. Although each family has its characteristic types of monomers, they all descend from two prototypes, A and B. We show that most differences between prototypes are concentrated in a short region (positions 35 to 51), which exists in two alternative states: it matches a binding site for pJ alpha in type A and the one for CENP-B in type B. Lower primates have only type A monomers whereas great apes have both A and B. The new family is formed by monomeric types almost identical to A and B prototypes, thus representing a living relic of alpha satellite. Analysis of these data shows that selection-driven evolution, rather than random fixation of mutations, formed the distinction between A and B types. To our knowledge, this is the first evidence for selection in any of the known satellite DNAs.

Animals↗

Cloning of a cDNA encoding a human protein which binds a sequence in the c-myc gene similar to the interferon-stimulated response element.

A human cDNA clone encoding a c-myc promoter-binding protein (IRLB) was selected by screening a human fibroblast lambda gt11 phage library with the hexamer oligodeoxyribonucleotide (oligo) 5'-GGCGGGAAAAAGAACGGA, corresponding to the protein-binding element of human c-myc similar to the interferon-stimulated response element (ISRE). The lambda gt11 phage clone, encoding a fusion protein which bound the probe oligo, was used to create an strain of Escherichia coli. The deduced amino-acid sequence of the cloned protein contains a putative alpha-helix which is expected to act as the DNA-binding domain. DNase footprinting analysis and oligo-binding specificity assays showed that the cloned factor recognizes the ISRE-like element of the P2 promoter region of human c-myc.

Amino Acid Sequence↗

Genomic organization, sequence and polymorphism of the human chromosome 4-specific alpha-satellite DNA.

Two alpha-satellite fragments specific for human chromosome 4 have been cloned and characterized. Under stringent annealing conditions, they hybridized in situ only to the pericentromeric region of chromosome 4, but under non-stringent conditions they hybridized to all chromosomes containing the sequences of alpha-satellite suprachromosomal family 2 (viz., chromosomes 2, 4, 8, 9, 13, 14, 15, 18, 20, 21 and 22). Southern blot analysis reveals the 3.2-kb higher-order repeated unit which exists in two forms: as a single MspI fragment or a combination of the 2.6-kb and 0.6-kb MspI fragments. The two chromosome-4-specific cloned sequences appear to be different parts of this repeated unit. Taken together they constitute about 60% of its length. The primary structure of the higher-order repeated unit is characterized by a dimeric periodicity of the D1-D2 type which is usual to suprachromosomal family 2. At least in one site this regularity is disrupted by monomer deletion leading to the D2-D2 monomeric order. The most likely mechanism of this monomer excision is homologous unequal crossing-over. These sequences may serve as both cytogenetic and restriction-fragment length polymorphism (RFLP) markers for the pericentromeric region of chromosome 4.

Base Sequence↗

Segment substitutions in alpha satellite DNA. Unusual structure of human chromosome 3-specific alpha satellite repeat unit.

We have sequenced the full-length copy of the alpha satellite higher-order repeated unit characteristic of human chromosome 3. Its internal structure, the regular alterations of J1 and J2 type monomers, is typical of the alphoid suprachromosomal family 1. This dimeric order is disrupted by the substitution of one J1 unit by an alien dimer which is not clearly related to any of the established monomeric types. We have also observed some other similar cases of segment substitutions in alpha satellite DNA. They probably represent a special type of molecular event which could be generated by gene conversion. Segment substitutions may be one of the important factors responsible for the extreme variability of localization patterns and actual sequences of alpha satellite DNA that should be taken into account in reconstructions of alpha satellite evolution.

Biological Evolution↗

Definition of a new alpha satellite suprachromosomal family characterized by monomeric organization.

We have analyzed more than 500 alphoid monomers either sequenced in our laboratory or available in the literature. Most of them belonged to the well studied suprachromosomal families 1, 2 and 3 characterized by dimeric (1 and 2) and pentameric (3) ancestral periodicities. The sequences that did not belong to the previously known families were subjected to further analysis. About a half of them formed a relatively homogenous family. Its members were on average 80.5% identical and 89.5% homologous to the M1 consensus sequence derived from this group (39 monomers). In the genome they do not form any ancestral periodicities other than a monomeric one, and are found at least in chromosomes 13, 14, 15, 21, 22 and Y. The newly defined family was termed suprachromosomal family 4. Comparison of all 10 alphoid monomeric groups identified so far showed that the M1 sequence is closely related to the J1-D2-W4-W5 homology grouping. Notably the African Green Monkey alpha satellite, also characterized by monomeric construction, appears to be a member of the same group.

Animals↗

Chromosome-specific alpha satellites: two distinct families on human chromosome 18.

Two types of human chromosome 18-specific alpha satellite fragments have been cloned and sequenced. They represent closely related but distinct alphoid families formed by two different types of the higher-order repeated units (1360-bp EcoRI and 1700-bp HindIII fragments) that do not alternate in the genome. The individual repeats within each family are 99% identical and interfamily homology is about 78%. Sequence analysis shows that both repeats belong to alphoid suprachromosomal family 2, but their homology is not higher than that of family members located on different chromosomes. Therefore, the two repeats shared a common origin in the recent past, although they are not the direct offspring of one ancestral sequence. Our data indicate that these two 18-specific domains have appeared as a result of two separate amplification events. Despite the high degree of homology, they are not undergoing intrachromosomal homogenization, although some variation of this process might take place within each domain.

Base Sequence↗

The primary structure of lupin seed 5.8 S ribosomal RNA.

The lack of colinearity between nucleotide sequence of the lupin 5.8 S rDNA gene (Rafalski, A.J., Wiewiórowski, M. and Soll, D. (1983) FEBS Lett. 152, 241-246) and 5.8 S rRNA of other plants (Erdmann, V.A. and Wolters, J. (1986) Nucleic Acids Res. 14, r1-r59.) prompted us to clarify this point by sequencing the native lupin 5.8 S rRNA. The sequence analysis was carried out using enzymatic and chemical methods. Lupin seed 5.8 S rRNA contains 164 nucleotides, including four modified ones: two residues of 2'-O-methylguanosine, one pseudouridine and one 2'-O-methyladenosine. The nucleotide sequence homology with the other plant 5.8 S rRNAs is approx. 88-96%.

Base Sequence↗

Molecular evolution of plants as deduced from changes in free energy of 5S ribosomal RNAs.

The nucleotide sequence of Pinus silvestyris 5S rRNA was determined using two independent methods and compared with other plant 5S rRNAs. It shows more than 90% sequence homology with gymnosperm 5S RNAs. The free energy (delta G) analysis of 5S rRNAs from gymnosperms, angiosperms and the other higher plants revealed that the free energy of this ribosomal RNA decreases with evolution.

Base Sequence↗

The calculation of plant 5S rRNAs secondary structure.

Using commercially available computer software package for ribonucleic acid (RNA) secondary structure analysis we calculated the free energy (delta G) of all higher plant 5S rRNA species. To gain insight into the relation between structure (nucleotide sequence) and free energy we generated point mutants of plant 5S rRNA and calculated their secondary structure. This analysis permitted to identify single sites which affect the stability and conformation of RNA molecule. Furthermore, the calculated data were compared with the electrophoretic mobility of 5S rRNA on polyacrylamide gels.

Base Sequence↗

The secondary structure of oocyte and somatic 5S ribosomal RNAs of the fish Misgurnus fossilis L. from nuclease hydrolyses and chemical modification data.

We have studied the accessibility of 5'- 32P labeled oocyte and somatic 5S rRNAs from the fish Misgurnus fossilis L. to S1, T1 and cobra venom nucleases and have found that the cleavage sites of 5S rRNAs closely related in primary structures differ in these molecules. The data of nuclease hydrolyses revealed the existence of two conformers corresponding to renatured and partially denatured somatic 5S rRNA and capable of mutual interconversions. The exposed cytosine residues were located in oocyte and somatic 5S rRNAs converted into uridine ones by sodium bisulfite treatment. The data have been used to construct the secondary structure models of somatic and oocyte 5S rRNAs by means of specially devised computer program. These models differ in their 5'-halves which contain all the nucleotide substitutions in the primary structure, all differences in location of the exposed cytosine residues, and finally, in the cleavage pattern by the nucleases used.

Animals↗

The primary structure of oocyte and somatic 5S rRNAs from the loach Misgurnus fossilis.

Somatic and oocyte 5S rRNAs from the liver and unfertilized eggs of the loach (Misgurnus fossilis have been sequenced and found to differ in six nucleotides. All the substitutions are confined to the 5'-half of the molecules; 4 of them are pyrimidine-pyrimidine substitutions, and 2 are purine-pyrimidine ones. Considerable differences, both in the position and the character of substitutions, have been established when these 5S rRNAs were compared with somatic and oocyte 5S rRNAs from Xenopus borealis and Xenopus laevis. Among the known primary structures, somatic 5S rRNA of M. fossilis is most similar to trout 5S rRNA.

Animals↗