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Biomedical subjects

F Azorín

Publications and source records attributed to F Azorín.

11 recordsLinked to original sources
PubMed1991

SV40 recombinants carrying a d(CT.GA)22 sequence show increased genomic instability.

Repetitive d(CT.GA)n sequences are commonly found in eukaryotic genomic DNA. They are frequently located in sites involved in genetic recombination or in promoter regions. To test for their possible biological function, a d(CT.GA)22 synthetic sequence was introduced into the genome of SV40, since it constitutes an appropriate model system for eukaryotic chromatin. When SV40 infects permissive cells, it proliferates in the form of a minichromosome. The simple repetitive sequence indicated above was inserted at the unique HpaII site of SV40 (at nt 346), and the genomic stability of SV40 recombinants carrying the d(CT.GA)22 sequence (SV/CT22 viruses) was analyzed. Upon serial passage through permissive CV1 cells, SV/CT22 recombinants show an increased production of defective viruses. Generation of SV/CT22 variants is likely to take place via recombination between and within viral molecules. The enhancement of the rate of recombination induced by the repetitive sequence is likely to be related to its known propensity to form triple-stranded structures. Many different variants coexist in the same viral population indicating that the mechanism by which they are produced is not unique. One variant (SV/X), showing a replicative advantage, was characterized in detail. Variant SV/X accounts for a large proportion of the total viral population. Its genomic organization corresponds to a tandem duplication of an early SV40 DNA fragment spanning from approx. nt 3200-nt 160. Variant SV/X contains a duplicated SV40 ori.

Base Sequence↗
PubMed1991

Identification of sequence elements contributing to the intrinsic curvature of the mouse satellite DNA repeat.

In this paper, the contribution of different sequence elements to the intrisic curvature of the mouse satellite DNA repeat was investigated. This DNA fragment contains nineteen groups of three or more consecutive adenines which are only poorly phased with respect to the helical repeat. The mouse satellite DNA repeat shows a sinusoidal pattern of cleavage by the hydroxyl radical; the waves of reactivity are phased with respect to the A-tracts. Some interesting observations arise from a detailed analysis of these cleavage patterns: a) the maxima of hydroxyl radical cleavage are more periodically spaced along the DNA sequence than the A-tracts themselves. As a consequence, the position of each maximum with respect to the A-tract is variable; b) the sequence 5' TGGAATATG/AA 3' shows a sinusoidal pattern of hydroxyl radical cleavage. This sequence shows a retarded migration in polyacrylamide gels indicating that it is actually intrinsically curved. These results are discussed in view of the current models for DNA curvature.

Animals↗
PubMed1991

The effect of the simple repeating d(CG.GC)n, d(CA.GT)n, and d(A.T)n DNA sequences on the nucleosomal organization of SV40 minichromosomes.

The effect of several simple repeating DNA sequences--d(CG.GC)5, d(CA.GT)30, and d(A.T)60--on the nucleosomal organization of the SV40 minichromosome is analyzed. These three different sequences were cloned at the Hpa II site of SV40 (position 346) which occurs at the 3' border of the nucleosome-free SV40 control region. Our results show that neither the d(A.T)60 sequence nor the d(CG.GC)5 sequence appear to have any relevant effect on the nucleosomal organization of the region of the minichromosome surrounding the inserted repeated sequence. Both sequences are hypersensitive to micrococcal nuclease cleavage in the minichromosome, indicating that they are not organized into nucleosomes. On the other hand, the d(CA.GT)30 sequence is found organized as nucleosomes and causes the delocation of nucleosomes in the minichromosomal region close to the inserted repeated sequence.

Chromosomes↗
PubMed1990

DNA-sequence and metal-ion specificity of the formation of *H-DNA.

The homopyrimidine-homopurine sequence d(CT/GA)22 undergoes, in the presence of zinc ions, transition to an altered DNA conformation (*H-DNA) which is neither H-DNA nor B-DNA. *H-DNA is characterized by a peculiar chemical reactivity pattern in which most of the polypyrimidine strand is hyperreactive to osmium tetroxide and the central part of the polypurine strand is sensitive to diethylpyrocarbonate. Formation of *H-DNA is specific of metal-ion. *H-DNA is detected in the presence of Zn++, Cd++ and Mn++. The efficiency on promoting the transition is in the order of Zn++ greater than Cd++ much greater than Mn++. Formation of *H-DNA is also specific of nucleotide sequence. From all the different homopolymeric sequences tested only the d(CT/GA)22 sequence showed the zinc-induced transition to *H-DNA. These results suggest that stabilization of *H-DNA involves the formation of a specific complex between the metal-ion and the nucleotide sequence. The biological relevance of these results is discussed in view of the important role that zinc ions play on many nucleic acids processes.

Base Sequence↗
PubMed1990

Determination of the DNA conformation of the simian virus 40 (SV40) enhancer in SV40 minichromosomes.

The simian virus 40 (SV40) enhancer contains three 8-bp purine-pyrimidine alternating sequences which are known to adopt the left-handed Z-DNA conformation in vitro. In this paper, we have undertaken the determination of the DNA conformation adopted by these Z-motifs in the SV40 minichromosome. We have analyzed the presence of Z-DNA through the change in linkage which should accompany formation of this left-handed conformation. Our results indicate that, regardless of the precise moment of the viral lytic cycle at which minichromosomes are harvested and the condition of the transfected DNA, either relaxed or negatively supercoiled, none of the three Z motifs of the SV40 enhancer exist to a significant extent as Z-DNA in SV40 minichromosomes. The SV40 enhancer adopts predominantly a right-handed B-DNA conformation in vivo.

Animals↗
PubMed1990

Satellite DNAs contain sequences that induced curvature.

The repeating units of mouse, rat, and alpha-monkey satellites have been cloned. All three show properties that are characteristic of curved DNA: (i) their migration in polyacrylamide gels is slower than predicted from their sequences, and (ii) they appear as curved molecules when visualized by electron microscopy. All three satellite repeats contain runs of d(A.T)n greater than or equal to 3 residues that are likely to be responsible for their curvature. From analysis of 20 different satellite DNA sequences, we conclude that, in satellite DNA, adenine residues show a high tendency to cluster in groups of three or more.

Animals↗
PubMed1989

Detection and molecular cloning of highly repeated DNA in the sea cucumber sperm.

A highly reiterated sequence in the sperm DNA of the echinoderm Holothuria tubulosa has been isolated by digestion with EcoRI, and cloned in the phagemid Bluescript. The monomeric unit has a repeat length of 391 bp and is arranged in tandem. The uncloned genomic monomer as well as two independent cloned fragments have been sequenced. The repeated element constitutes about 1.8% of total Holothuria DNA which corresponds to a repetition frequency of about 1.4 x 10(5) copies per haploid complement. The repetitive sequence has a high A + T content (66.8%) characterized by scattered tracts of A and T residues with no apparent internal sub-repeats, although several inverted and direct repeats are present. Heterogeneity between monomers derived from individual clones is low, whereas sequence similarity to known repetitive elements appears to be negligible.

Animals↗
PubMed1989

Structural polymorphism of homopurine--homopyrimidine sequences: the secondary DNA structure adopted by a d(GA.CT)22 sequence in the presence of zinc ions.

In this paper, we have analysed the conformational behaviour shown by the homopurine--homopyrimidine alternating d(GA.CT)22 sequence cloned into SV40. Our results show that, in the presence of zinc ions, the d(GA.CT)22 sequence adopts an altered secondary DNA structure (*H-DNA) which differs from either B-DNA or H-DNA. Formation of *H-DNA is facilitated by negative supercoiling and does not appear to require base protonation, since it is induced at neutral pH by approximately 0.4 mM ZnCl2. The patterns of OsO4 and DEPC modification obtained in the presence of zinc are compatible with a homopurine--homopurine--homopyridimine triplex, though other structural models for *H-DNA are also possible. The hypersensitivity to S1-cleavage of the d(GA.CT)22 sequence is reinterpreted in terms of the equilibria between the B-, H- and *H-forms of the sequence. These results reveal the high degree of structural polymorphism shown by homopurine-homopyrimidine sequences. Its biological relevance is discussed.

Animals↗
PubMed1987

Supercoiled induced transition to the Z-DNA conformation affects the ability of a d(CG/GC)12 sequence to be organized into nucleosome-cores.

Nucleosome-cores were reconstituted by the salt-dialysis method onto closed circular pDHg16 DNA which contains a d(CG/GC)12 sequence. Alternating d(CG/GC)n sequences form left-handed Z-DNA readily when contained in negatively supercoiled DNA. We have investigated the ability of the d(CG/GC)12 sequence to be organized into nucleosome-cores when stabilized as Z-DNA through negative supercoiling. We have found that nucleosome assembly at the d(CG/GC)12 insert is prevented when the sequence is stable in the Z-conformation but it is not affected at all when the sequence adopts the right-handed B-form.

Animals↗
PubMed1985

Condensation of DNA by the C-terminal domain of histone H1. A circular dichroism study.

The condensation of DNA by the C-terminal domain of histone H1 has been studied by circular dichroism in physiological salt concentration (0.14 M NaF). As the intact H1 molecule, its C-terminal domain induces the so-called psi state of DNA that is characterized by a nonconservative circular dichroism spectrum which is currently attributed to ordered aggregation of the DNA molecules. On a molar basis, intact H1 and its C-terminal domain give spectra of similar intensity. Neither the globular domain of H1 nor an N-terminal fragment, that includes both the globular and N-terminal domains, has any effect on the conservative circular dichroism of DNA. From these results it is concluded that the condensation of DNA mediated by histone H1 is mainly due to its C-terminal domain. The effect of the salt concentration and the size of DNA molecules on the circular dichroism of the complexes are also examined.

Animals↗