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S Venditti

Publications and source records attributed to S Venditti.

14 recordsLinked to original sources
PubMed2000

Telomere-based neo-Darwinian selection of yeast clonal subpopulations.

In Saccharomyces cerevisiae, imbalance of the genes coding for the heterochromatin components Sir3p and histone H4 (namely, overdosage of SIR3 and lack of one of the two genes coding for H4) causes modifications in telomere length and telomere sequence organization, favoring the insertion of Y' elements into a stably shortened (C1-3A)n repeat tract. We report here that the newly inserted Y' elements are unstable and are lost with high frequency, generating clonal subpopulations with short telomeres, as revealed by the analysis of a specific telomere (LIII) and of the overall population of telomeres. Moreover, the growth rates of the subpopulations with and without Y' elements on LIII are different, the Y'-less individuals reproducing 20% more slowly than individuals bearing Y' elements. When grown together with Y'-bearing individuals, the subpopulations with the normal LIII telomere (which are viable and genetically stable if grown alone) are rapidly competed out. Hence, genetic imbalance for the structural components of heterochromatin results in a complex and rapidly changing mixture of subpopulations in such cultures. Thus, in situations where subpopulations are allowed to compete, heterochromatin-based differential growth rates result in neo-Darwinian clonal selection.

Genes, Fungal↗
PubMed1999

Heterochromatin organization of a natural yeast telomere. Recruitment of Sir3p through interaction with histone H4 N terminus is required for the establishment of repressive structures.

The chromatin organization of eukaryotic telomeres is essential for telomeric function and is currently receiving great attention. In yeast, the structural organization of telomeres involves a complex interplay of telomeric proteins that results in the formation of heterochromatin. This telomeric heterochromatin involves homotypic and heterotypic protein interactions that have been summarized in a general model. Recent analyses have focused on the study of the structural complexity at yeast telomeres to the level of specific nucleosomes and of the distribution of protein complexes in a natural telomeric region (LIII). In this report, we further analyze the structural complexity of LIII and the implication of this structure on telomeric silencing. It is shown that the establishment of repressive heterochromatin structures at LIII requires the recruitment of Sir3p through interaction with the N terminus of histone H4. The establishment of such structures does not require acetylation of any of four lysines located in the H4 N terminus (lysines 5, 8, 12, and 16).

Acetylation↗
PubMed1999

Imbalance in dosage of the genes for the heterochromatin components Sir3p and histone H4 results in changes in the length and sequence organization of yeast telomeres.

Telomeric heterochromatin plays an essential role in telomere function, including the regulation of telomere length. We observe that in Saccharomyces cerevisiae an imbalance in the dosage of genes for two protein components of heterochromatin (namely Sir3p and histone H4) causes modifications in telomere length and telomere sequence organization. The effects of Sir3p/H4 imbalance were analyzed in yeast strains in which the wild-type SIR3 gene (normally a single-copy gene) was either absent or present in 20-30 copies, and both histone H4 genes (HHF1 and HHF2) were present or HHF1 was deleted, thus covering a wide range of viable gene-dosage combinations. Modifications of telomeres and of subtelomeric regions were identified by analyzing both the overall telomere population and by focusing on two single telomeric regions: the left telomere of chromosome III (LIII) and the right telomere of chromosome XI (RXI). The modifications induced by alteration of the Sir3p/H4 ratio consist of a reduction in the length and an increase in the instability of the terminal block of (C(1-3)A)n repeats and in susceptibility to insertion of Y' elements into this repeat element. Restoration of the wild-type gene ratio (by removal of the extra copies of SIR3 or by complementation with the missing second copy of HHF) restored the original telomere organization, both with respect to the length of the (C(1-3)A)n repeat stretch and the absence of Y' elements. This behavior shows that the stability of the wild-type sequence organization requires maintenance of the normal structure of telomeric heterochromatin.

Chromosomes, Fungal↗
PubMed1998

Heterochromatin organization of a natural yeast telomere. Changes of nucleosome distribution driven by the absence of Sir3p.

We have defined the in vivo heterochromatin structure of the left telomere of Saccharomyces cerevisiae chromosome III (LIII). Analysis of heterochromatin of a single telomere was so far lacking, due to the difficulties intrinsic to the highly repetitive nature of telomeric sequences. In LIII, the terminal (C1-3A)n repetitive sequences are followed by a complete X element and by the single copy Ty5-1 retrotransposon. Both the telosome and the X element exhibit overall resistance to micrococcal nuclease digestion reflecting their tight chromatin structure organization. The X element contains protein complexes and irregularly distributed but well localized nucleosomes. In contrast, a regular array of phased nucleosomes is associated with the promoter region of Ty5-1 and with the more centromere-proximal sequences. The lack of a structural component of yeast telomeres, the SIR3 protein, does not alter the overall tight organization of the X element but causes a nucleosome rearrangement within the promoter region of Ty5-1 and releases Ty5-1 silencing. Thus, Sir3p links the modification of the heterochromatin structure with loss of transcriptional silencing.

Chromosomes, Fungal↗
PubMed1997

Chromatin structure of the Saccharomyces cerevisiae DNA topoisomerase I promoter in different growth phases.

We have determined the chromatin organization of the Saccharomyces cerevisiae DNA topoisomerase I promoter. Three nucleosomal core particles have been mapped at nucleotide level over the promoter region, encompassing the presumptive TATA sequence and the two RNA initiation sites; the most upstream nucleosome particle forms on to a 29 bp-long poly(dA-dT) element. This simple organization remains constant throughout both the logarithmic and the linear phase of growth, with the exception of an increased accessibility to micrococcal nuclease of the nucleosome covering the TATA box and the RNA initiation sites during the diauxic shift (the switching from the fermentative to the respiratory metabolism) in parallel with an increase of the DNA topoisomerase I mRNA. In addition, a strong disorganization of the bulk chromatin structure in the late stationary phase is also reported.

Chromatin↗
PubMed1994

Conformational information in DNA: its role in the interaction with DNA topoisomerase I and nucleosomes.

Information in DNA is not limited to sequence information. Both local and global conformational parameters are pivotal to the interaction with a number of relevant proteins. The function of the major components of the transcription machinery (RNA polymerase II, DNA topoisomerase I, nucleosomes, the TATA-binding factor) is dependent on the topological status of the substrate DNA molecule. The topological requirements and the conformational consensus that dictate the rules for localization of nucleosomes and define the active sites for DNA topoisomerase I have been established; the reaction of DNA topoisomerase I is regulated by a topological feedback mechanism. The integrating function of the free energy of supercoiling in the transcription process and the regulatory role of DNA topoisomerase I are discussed.

Animals↗
PubMed1994

In vivo analysis of chromatin following nystatin-mediated import of active enzymes into Saccharomyces cerevisiae.

In vivo DNA-protein interactions are usually studied at the molecular level using DNA-degrading agents of low molecular weight. In order to be useful, macromolecular probes of chromatin structure, such as enzymes must first cross the cell membrane. In this paper we describe the introduction and evaluation of macromolecules with enzymatic activity into yeast spheroplasts treated with the polyene antibiotic nystatin. We report the low resolution analysis of chromatin structure in the promoter region of the Saccharomyces cerevisiae gene encoding DNA topoisomerase I by this technique using micrococcal nuclease and restriction enzymes.

Base Sequence↗
PubMed1994

Effects of DNA topology in the interaction with histone octamers and DNA topoisomerase I.

Several simple proteins and complex protein systems exist which do not recognize a defined sequence but--rather--a specific DNA conformation. We describe experiments and principles for two of these systems: nucleosomes and eukaryotic DNA topoisomerase I. Evidences are summarized that describe the effects of negative DNA supercoiling on nucleosome formation and the influence of DNA intrinsic curvature on their localization. The function of the DNA rotational information in nucleosome positioning and in the selection of multiple alternative positions on the same helical phase are described. This function suggests a novel genetic regulatory mechanism, based on nucleosome mobility and on the correlation between in vitro and in vivo positions. We observe that the same rules that determine the in vitro localization apply to the in vivo nucleosome positioning, as determined by a technique that relies on the use of nystatin and on the import of active enzymes in living yeast cells. The sensitivity of DNA topoisomerase I to the topological condition of the DNA substrate is reviewed and discussed taking into account recent experiments that describe the effect of the DNA tridimensional context on the reaction. These topics are discussed in the following order: (i) Proteins that look for a consensus DNA conformation; (ii) Nucleosomes; (iii) Negative supercoiling and nucleosomes; (iv) DNA curvature/bending and nucleosomes; (v) Multiple positioning; (vi) Multiple nucleosomes offer a contribution to the solution of the linking number paradox; (vii) Rotational versus translational information; (viii) A regulatory mechanism; (ix) DNA topoisomerase I; (x) DNA topoisomerase I and DNA supercoiling: a regulation by topological feedback; (xi) DNA topoisomerase I and DNA curvature; (xii) The in-and-out problem in the accessibility of DNA information; (xiii) The integrating function of the free energy of supercoiling.

DNA Topoisomerases, Type I↗
PubMed1991

A DNA conformational alteration induced by a neighboring oligopurine tract on GAATTA enables nicking by EcoRI.

The pseudo EcoRI site GAATTA in the U3 region of the long terminal repeat of human immunodeficiency virus, which is flanked by a 26-base pair oligopurine tract, is readily nicked by either EcoRI or RsrI. The strand-specific nick occurs predominantly between the G and A residues and is independent of negative supercoiling. Other GAATTA sites surrounded by random (non-oligopurine) sequences are not nicked by these restriction endonucleases. However, other types and lengths of oligopurine tracts are effective in inducing the nicking in neighboring GAATTA sites. Hence, we propose that the flanking oligopurine tracts induce an altered DNA conformation on the GAATTA target site which may be similar to the transition state induced by EcoRI when binding to its canonical recognition site. Gel retardation analyses on restriction fragments containing the oligopurine-GAATTA-oligopurine sequences suggest the presence of helical axis distortions which are consistent with this interpretation.

Base Sequence↗
PubMed1989

Linkage reduction allows reconstitution of nucleosomes on DNA microdomains.

We have established an experimental system for reconstitution of an individual nucleosome on a closed DNA microdomain (operationally defined as a DNA domain of a size so small as to be unable to establish titratable superhelical turns). The microdomain (185 base-pairs (bp), composed of 128 bp encompassing the central part of the Saccharomyces cerevisiae ADH II promoter plus 57 bp of a polylinker) was obtained by ligation under conditions that produced three circularized forms characterized by different linkage numbers. These linkomers were tested for nucleosome reconstitution with S. cerevisiae histones. It was observed that only microcircles with linkage reduction (delta Lk = 1 or 2) could form a nucleosome, as defined by protection of a 145(+/- 2) bp DNA fragment from micrococcal nuclease, relaxed forms (open or closed circles) could not.

Base Composition↗
PubMed1988

DNA conformational variations in the in vitro torsionally strained Ig kappa light chain gene localize on consensus sequences.

We have analyzed the localization and the dependence upon superhelical density of the DNA sites which modify their conformation under torsional strain in a mouse Ig L kappa gene. The conformational variations occur on DNA sites which have been defined as protein interaction sites and consensus sequence motifs: the 5'-upstream regulatory decanucleotides, the TATA sequence, the consensus heptanucleotides of the J recombinational sequences.

Animals↗
PubMed1988

The intrinsic topological information of the wild-type and of up-promoter mutations of the Saccharomyces cerevisiae alcohol dehydrogenase II regulatory region.

A 569-base pair fragment encompassing the upstream regulatory region, the RNA initiation sites, and the initial part of the coding region of the Saccharomyces cerevisiae alcohol dehydrogenase II gene has been analyzed for the presence of sites which undergo conformational modification under torsional stress. Fine mapping of P1 and S1 endonuclease-sensitive sites was obtained on single topoisomers produced by in vitro ligation. It was shown that the upstream activator sequence, the TATA sequence, a region directly upstream to the RNA initiation sites, and several positions in the first segment of the transcribed region change conformation as a function of the applied torsional stress in a precisely coordinate fashion. The superhelical density optima for this coordinate modifications have been determined. Analysis of the conformational changes of the promoter sequence in several naturally occurring (Young, E. T., Williamson, V. M., Taguchi, A., Smith, M., Sledziewski, L., Russel, D., Osterman, J., Denis, C., Cox, D., and Beier, D., (1982) in Genetic Engineering of Microorganisms for Chemicals (Hollander, A., De Moss, R. D., Kaplan, S., Konisky, J., Savage, D., and Wolle, R. S., eds) pp. 335-361, Plenum Publishing Corp., New York) up-promoter constitutive mutants was performed. This analysis has shown that the conformation of functionally relevant sites changes as a function of sequence mutations that have taken place elsewhere; this shows that the conformational behavior of the whole promoter region is linked and suggests transmission in cis of topological effects in RNA polymerase II promoters.

Alcohol Dehydrogenase↗