Biomedical subjects

Arthur J Lustig

Publications and source records attributed to Arthur J Lustig.

8 recordsLinked to original sources

PubMed2006

Sir3 C-terminal domain involvement in the initiation and spreading of heterochromatin.

Heterochromatin is nucleated at a specific site and subsequently spreads into distal sequences through multiple interactions between modified histones and nonhistone proteins. In the yeast Saccharomyces cerevisiae, these nonhistone proteins include Sir2, Sir3, and Sir4. We have previously shown that loss of the C-terminal Rap1 domain containing Sir3 and Sir4 association sites can be overcome by tethering a 144-amino-acid C-terminal domain (CTD) of Sir3 adjacent to the telomere. Here, we explore the substructure and functions of the CTD. We demonstrate that the CTD is the minimum domain for Sir3 homodimerization, a function that is conserved in related yeasts. However, CTD heterodimers associate at only low efficiencies and correspondingly have low levels of tethered silencing, consistent with an essential role for dimerization in tethered silencing. Six missense alleles were generated, with ctd-Y964A producing the most extreme phenotypes when tethered to the LexA binding sites. Although ctd-Y964A is capable of dimerization, telomere silencing is abrogated, indicating that the CTD serves a second essential function in silencing. Chromatin immunoprecipitation analyses of wild-type and ctd-Y964A mutant cells indicate an association of the CTD with the deacetylated histone tails of H3 and H4 that is necessary for the recruitment of Sir3. The efficiency of spreading depends upon the apparent stoichiometry and stability during the initiation event. The predicted Cdc6 domain III winged-helix structure may well be responsible for dimerization.

Binding Sites↗

PubMed2006

Telomere dynamics in genome stability.

The past several years have seen an increasing interest in telomere recombinational interactions that provide many functions in telomere capping, in telomere size homeostasis and in overcoming the catastrophic effects of telomerase deficiency. Several key recombination mechanisms have emerged from recent investigations. In the yeasts, these mechanisms include exchange between subtelomeric regions and telomere sequences, rapid telomere expansion and telomere deletion. These processes proceed by pathways that use both the cellular recombination machinery and novel mechanisms such as rolling circle replication. The insights gained from recent studies extend our understanding of similar processes in higher eukaryotes and suggest that the recombinational dynamics of telomeres have additional roles that contribute to genomic stability and instability.

Animals↗

PubMed2005

Mre 11 p nuclease activity is dispensable for telomeric rapid deletion.

Telomeric rapid deletion (TRD) is an intrachromatid recombination process that truncates over-elongated telomeres to the genetically determined average telomere length. We have proposed that TRD is initiated by invasion of the 3' G-rich overhang into centromere-proximal telomere sequence, forming an intermediate that leads to excision of the distal telomere tract. TRD efficiency is dependent on Mre 11p and Rad50p, two members of the widely conserved Mre 11p/Rad50p/Xrs2p (MRX) complex. To investigate the role of Mre 11p in TRD, we conducted a structure/function analysis by testing the TRD rate and precision of mutations within known functional domains. We analyzed 12 alleles that disrupt different Mre 11p activities. Surprisingly, mutations in essential residues of the nuclease domain do not inhibit TRD, effectively ruling out nuclease activity as the source of the Mre 11p requirement. Interestingly, loss of Exo1p alone or loss of Exo1p in an Mre 11 nuclease deficient background does not eliminate TRD, suggesting the presence of an additional nuclease. Second, deletion of DNA binding sites A (residues 410--420) and B (residues 644--692) actually enhances the TRD rate. Even deletion of both DNA binding domains does not abrogate TRD, although its kinetics and precision are variable. This suggests altered DNA binding or a conformational defect in the MRX complex may affect the rate of TRD product formation and indicates that the DNA binding sites formally act as repressors of TRD. Remarkably, the H213Y allele (nuclease motif IV) confers an extraordinarily rapid kinetics, with the vast majority of elongated telomeres deleted imprecisely in a single round of subculturing. In striking contrast, the P162S allele that confers dissolution of the complex also exhibits the null phenotype. These data suggest that Mre 11p can act as a positive and negative regulator of TRD in context of the MRX complex that is essential for TRD.

Cell Cycle Proteins↗

PubMed2005

Ndj1p-dependent epigenetic resetting of telomere size in yeast meiosis.

Telomeres are essential for the protection of chromosomes against nucleases and recombinases and for the addition of G+T-rich simple sequence by the ribonucleoprotein reverse transcriptase telomerase . Telomere size instability and loss of telomerase activity in somatic cells is strongly associated with both oncogenesis and aging . Yet, an understanding of the mechanisms that maintain telomere size and structure during meiosis is still in its infancy . We have investigated the stability of single elongated telomeres during yeast meiosis. We find that elongated telomeres undergo high rates of precise deletion to wild-type telomere size via an intrachromatid pathway that shares properties with mitotic telomere rapid deletion (TRD). Loss of Ndj1p, a telomeric protein necessary for meiotic bouquet structure formation , confers a severe reduction in deletion rates. Return-to-growth (RTG) experiments suggest that deletion occurs at or near the period of meiotic recombination in NDJ1/NDJ1, but not in ndj1Delta/ndj1Delta diploids . We propose that Ndj1p facilitates deletion by promoting telomeric interactions during meiosis, resulting in an effective increase in the concentration of limiting factors for deletion.

Base Sequence↗

PubMed2004

Telomerase RNA: a flexible RNA scaffold for telomerase biosynthesis.

Determination of the structure of the yeast telomerase RNA component TLC1 has been hampered by its large size and high rate of evolutionary divergence. But detailed phylogenetic comparisons have now revealed the unusually flexible and modular architecture of this important RNA molecule.

DNA-Binding Proteins↗

PubMed2003

The paradoxical relationship between NHEJ and telomeric fusion.

Recent results shed new light on the origin of fusion products observed in the destabilized chromosomes of cancer and related diseases. These findings define an unusual relationship between nonhomologous end joining (NHEJ) and telomere "capping," with identical proteins playing opposing roles.

Cell Division↗

PubMed2003

Clues to catastrophic telomere loss in mammals from yeast telomere rapid deletion.

Catastrophic losses of telomeric sequences have recently been described during apoptosis, senescence and tumorigenesis in murine and human cells, in ataxia telangiectasia patients and in immortalized cells in which telomerase is inactive. A mechanism that underlies a single-step non-reciprocal telomere deletion called telomere rapid deletion in Saccharomyces cerevisiae might provide clues for future studies of catastrophic telomere loss in higher eukaryotes.

Animals↗

PubMed2003

Multiple roles for Saccharomyces cerevisiae histone H2A in telomere position effect, Spt phenotypes and double-strand-break repair.

Telomere position effects on transcription (TPE, or telomeric silencing) are nucleated by association of nonhistone silencing factors with the telomere and propagated in subtelomeric regions through association of silencing factors with the specifically modified histones H3 and H4. However, the function of histone H2A in TPE is unknown. We found that deletion of either the amino or the carboxyltails of H2A substantially reduces TPE. We identified four H2A modification sites necessary for wild-type efficiency of TPE. These "hta1tpe" alleles also act as suppressors of a delta insertion allele of LYS2, suggesting shared elements of chromatin structure at both loci. Interestingly, we observed combinatorial effects of allele pairs, suggesting both interdependent acetylation and deacetylation events in the amino-terminal tail and a regulatory circuit between multiple phosphorylated residues in the carboxyl-terminal tail. Decreases in silencing and viability are observed in most hta1tpe alleles after treatment with low and high concentrations, respectively, of bleomycin, which forms double-strand breaks (DSBs). In the absence of the DSB and telomere-binding protein yKu70, the bleomycin sensitivity of hta1tpe alleles is further enhanced. We also provide data suggesting the presence of a yKu-dependent histone H2A function in TPE. These data indicate that the amino- and carboxyl-terminal tails of H2A are essential for wild-type levels of yKu-mediated TPE and DSB repair.

Acetylation↗