http://web.mit.edu/biology/guarente/references/5.pdf

hSIR2SIRT1 Functions as an NAD-Dependent p53 Deacetylase

The spindle-assembly checkpoint is a safety device that monitors the attachment of spindle microtubules to kinetochores and ensures the fidelity of chromosome segregation in mitosis. Molecular studies are finally starting to reveal the mechanisms of checkpoint activation and inactivation.

The spindle-assembly checkpoint in space and time.

Recent studies have revealed extensive genetic diversity both between and within tumours. This heterogeneity affects key cancer pathways, driving phenotypic variation, and poses a significant challenge to personalized cancer medicine. A major cause of genetic heterogeneity in cancer is genomic instability. This instability leads to an increased mutation rate and can shape the evolution of the cancer genome through a plethora of mechanisms. By understanding these mechanisms we can gain insight into the common pathways of tumour evolution that could support the development of future therapeutic strategies.

The causes and consequences of genetic heterogeneity in cancer evolution.

Aging is accompanied by a decline in the healthy function of multiple organ systems, leading to increased incidence and mortality from diseases such as type II diabetes mellitus, neurodegenerative diseases, cancer, and cardiovascular disease. Historically, researchers have focused on investigating individual pathways in isolated organs as a strategy to identify the root cause of a disease, with hopes of designing better drugs. Studies of aging in yeast led to the discovery of a family of conserved enzymes known as the sirtuins, which affect multiple pathways that increase the life span and the overall health of organisms. Since the discovery of the first known mammalian sirtuin, SIRT1, 10 years ago, there have been major advances in our understanding of the enzymology of sirtuins, their regulation, and their ability to broadly improve mammalian physiology and health span. This review summarizes and discusses the advances of the past decade and the challenges that will confront the field in the coming years.

/pdf/mammalian-sirtuins-biological-insights-and-disease-relevance-195sikg6e7.pdf

Mammalian sirtuins: biological insights and disease relevance.

Sirtuins are a conserved family of proteins found in all domains of life. The first known sirtuin, Sir2 (silent information regulator 2) of Saccharomyces cerevisiae, from which the family derives its name, regulates ribosomal DNA recombination, gene silencing, DNA repair, chromosomal stability and longevity. Sir2 homologues also modulate lifespan in worms and flies, and may underlie the beneficial effects of caloric restriction, the only regimen that slows aging and extends lifespan of most classes of organism, including mammals. Sirtuins have gained considerable attention for their impact on mammalian physiology, since they may provide novel targets for treating diseases associated with aging and perhaps extend human lifespan. In this review we describe our current understanding of the biological function of the seven mammalian sirtuins, SIRT1-7, and we will also discuss their potential as mediators of caloric restriction and as pharmacological targets to delay and treat human age-related diseases.

/pdf/sirtuins-in-mammals-insights-into-their-biological-function-4n0wwh14ol.pdf

Sirtuins in mammals: insights into their biological function

Kinetochores are multiprotein complexes that assemble on centromeric DNA and attach chromosomes to spindle microtubules. Over the past six years, the number of proteins known to localize to the Saccharomyces cerevisiae kinetochore has increased from around 10 to over 60. However, relatively little is known about the protein-protein interactions that mediate kinetochore assembly or about the overall structure of microtubule-attachment sites. Here we used biophysical techniques, affinity purification, mass spectrometry, and in vivo assays to examine the state of association of 31 centromere-binding proteins, including six proteins newly identified as kinetochore subunits. We found that yeast kinetochores resemble transcriptional enhancers in being composed of at least 17 discrete subcomplexes that assemble on DNA to form a very large structure with a mass in excess of 5 MD. Critical to kinetochore assembly are proteins that bridge subunits in direct contact with DNA and subunits bound to microtubules. We show that two newly identified kinetochore complexes, COMA (Ctf19p-Okp1p-Mcm21p-Ame1p) and MIND (Mtw1p including Nnf1p-Nsl1p-Dsn1p) function as bridges. COMA, MIND, and the previously described Ndc80 complex constitute three independent and essential platforms onto which outer kinetochore proteins assemble. In addition, we propose that the three complexes have different functions with respect to force generation and MT attachment.

/pdf/hierarchical-assembly-of-the-budding-yeast-kinetochore-from-48sv166l6r.pdf

Hierarchical assembly of the budding yeast kinetochore from multiple subcomplexes

Background: Kinetochores are large multi-protein structures that assemble on centromeric DNA (CEN DNA) and mediate the binding of chromosomes to microtubules. Comprising 125 base-pairs of CEN DNA and 70 or more protein components, Saccharomyces cerevisiae kinetochores are among the best understood. In contrast, most fungal, plant and animal cells assemble kinetochores on CENs that are longer and more complex, raising the question of whether kinetochore architecture has been conserved through evolution, despite considerable divergence in CEN sequence. Results: Using computational approaches, ranging from sequence similarity searches to hidden Markov model-based modeling, we show that organisms with CENs resembling those in S. cerevisiae (point CENs) are very closely related and that all contain a set of 11 kinetochore proteins not found in organisms with complex CENs. Conversely, organisms with complex CENs (regional CENs) contain proteins seemingly absent from point-CEN organisms. However, at least three quarters of known kinetochore proteins are present in all fungi regardless of CEN organization. At least six of these proteins have previously unidentified human orthologs. When fungi and metazoa are compared, almost all have kinetochores constructed around Spc105 and three conserved multiprotein linker complexes (MIND, COMA, and the NDC80 complex). Conclusion: Our data suggest that critical structural features of kinetochores have been well conserved from yeast to man. Surprisingly, phylogenetic analysis reveals that human kinetochore proteins are as similar in sequence to their yeast counterparts as to presumptive Drosophila melanogaster or Caenorhabditis elegans orthologs. This finding is consistent with evidence that kinetochore proteins have evolved very rapidly relative to components of other complex cellular

/pdf/phylogenetic-and-structural-analysis-of-centromeric-dna-and-cjyokycyyj.pdf

Phylogenetic and structural analysis of centromeric DNA and kinetochore proteins.

…S. cerevisiae centromeres represent the ultimate stage of centromere optimization… ▪ Abstract       (Malik & Henikoff 2002) Kinetochores are multiprotein complexes that assemble on centromeric DNA and mediate the attachment and movement of chromosomes along the microtubules (MTs) of the mitotic spindle. This review focuses on the simplest eukaryotic centromeres and kinetochores, those found in the budding yeast Saccharomyces cerevisiae. Research on kinetochore function and chromosome segregation is focused on four questions of general significance: What specifies the location of centromeres? What are the protein components of kinetochores, and how do they assemble a MT attachment site? How do MT attachments generate force? How do cells sense the state of attachment via the spindle assembly checkpoint?

Structure, Function, and Regulation of Budding Yeast Kinetochores

Mitotic spindles are self-organizing protein machines that harness teams of multiple force generators to drive chromosome segregation. Kinesins are key members of these force-generating teams. Different kinesins walk directionally along dynamic microtubules, anchor, crosslink, align and sort microtubules into polarized bundles, and influence microtubule dynamics by interacting with microtubule tips. The mechanochemical mechanisms of these kinesins are specialized to enable each type to make a specific contribution to spindle self-organization and chromosome segregation.

/pdf/prime-movers-the-mechanochemistry-of-mitotic-kinesins-380m09mm28.pdf

Prime movers: the mechanochemistry of mitotic kinesins

There are two main pathways in eukaryotic cells for the repair of DNA double-strand breaks: homologous recombination and nonhomologous end joining. Because eukaryotic genomes are packaged in chromatin, these pathways are likely to require the modulation of chromatin structure. One way to achieve this is by the acetylation of lysine residues on the N-terminal tails of histones. Here we demonstrate that Sin3p and Rpd3p, components of one of the predominant histone deacetylase complexes of Saccharomyces cerevisiae, are required for efficient nonhomologous end joining. We also show that lysine 16 of histone H4 becomes deacetylated in the proximity of a chromosomal DNA double-strand break in a Sin3p-dependent manner. Taken together, these results define a role for the Sin3p/Rpd3p complex in the modulation of DNA repair.

https://www.pnas.org/content/pnas/101/6/1644.full.pdf

Andrew D. McAinsh

Papers

Hierarchical assembly of the budding yeast kinetochore from multiple subcomplexes

Phylogenetic and structural analysis of centromeric DNA and kinetochore proteins.

Structure, Function, and Regulation of Budding Yeast Kinetochores

Prime movers: the mechanochemistry of mitotic kinesins

Saccharomyces cerevisiae Sin3p facilitates DNA double-strand break repair