qpcrtag-analysis-high-throughput-real-time-pcr-assay-for-sc20

qPCRTag Analysis - A High Throughput, Real Time PCR Assay for Sc2.0 Genotyping

The Synthetic Yeast Genome Project (Sc2.0) aims to build 16 designer yeast chromosomes and combine them into a single yeast cell. To date one synthetic ...

Roche Life Science, USA

A genetic predisposition for progressive enlargement of thoracic aortic aneurysms leading to type A dissection (TAAD) is inherited in an autosomal-dominant manner in up to 19% of patients, and a number of chromosomal loci have been identified for the condition. Having mapped a TAAD locus to 3p24-25, we sequenced the gene for transforming growth factor-beta receptor type II (TGFBR2) to determine whether mutations in this gene resulted in familial TAAD.

Mutations in transforming growth factor-beta receptor type II cause familial thoracic aortic aneurysms and dissections.

Thoracic aortic aneurysms leading to type A dissections (TAAD) can be inherited in isolation or in association with genetic syndromes, such as Marfan syndrome and Loeys-Dietz syndrome. When TAAD occurs in the absence of syndromic features, it is inherited in an autosomal dominant manner with decreased penetrance and variable expression, the disease is referred to as familial TAAD. Familial TAAD exhibits significant clinical and genetic heterogeneity. The first genes identified to cause TAAD were FBN1, TGFBR2, and TGFBR1. The identification and characterization of these genes suggested that increased TGF-beta signaling plays a role in pathogenesis. The recent discovery that mutations in the vascular smooth muscle cell (SMC)-specific beta-myosin (MYH11) and alpha-actin (ACTA2) can also cause this disorder has focused attention on the importance of the maintenance of SMC contractile function in preserving aortic structure and preventing TAAD.

Genetic basis of thoracic aortic aneurysms and dissections: focus on smooth muscle cell contractile dysfunction.

Transforming growth factor-β (TGF-β) signaling is critical for the differentiation of smooth muscle cells (SMCs) into quiescent cells expressing a full repertoire of contractile proteins. Heterozygous mutations in TGF-β receptor type II (TGFBR2) disrupt TGF-β signaling and lead to genetic conditions that predispose to thoracic aortic aneurysms and dissections (TAADs). The aim of this study is to determine the molecular mechanism by which TGFBR2 mutations cause TAADs.

TGFBR2 mutations alter smooth muscle cell phenotype and predispose to thoracic aortic aneurysms and dissections.

Sister chromatids are held together, from the time they are made during S phase until they are pulled apart just before cell division, by a protein complex called cohesin. The mechanistic details by which sister chromatid cohesion is established and maintained have remained elusive, particularly in vertebrate systems. Sororin, a protein that interacts with the cohesin complex, is essential for cohesion in vertebrates, but how it participates in the process is unknown. Here we demonstrate that sororin recruitment depends on active DNA replication and that sororin loading onto chromosomes depends upon another essential cohesion factor, the acetyltransferase Eco2. We find that Eco2, like sororin, is a substrate of the anaphase-promoting complex (APC), which ensures that protein levels remain low before S phase. These findings demonstrate that sororin and Eco2 work together to form a unique regulatory module that limits cohesion to cells with replicated chromatin and support a model in which cohesion in vertebrates is not fully established until the G2 phase of the cell cycle.

Sororin cooperates with the acetyltransferase Eco2 to ensure DNA replication-dependent sister chromatid cohesion.

Acetylation of the Smc3 subunit of cohesin is essential to establish functional cohesion between sister chromatids. Smc3 acetylation is catalyzed by members of the Eco family of acetyltransferases, although the mechanism by which acetylation is regulated and how it promotes cohesion are largely unknown. In vertebrates, the cohesin complex binds to chromatin during mitotic exit and is converted to a functional form during or shortly after DNA replication. The conserved proliferating cell nuclear antigen-interacting protein box motif in yeast Eco1 is required for function, and cohesin is acetylated during the S phase. This has led to the notion that acetylation of cohesin is stimulated by interaction of Eco1 with the replication machinery. Here we show that in vertebrates Smc3 acetylation occurs independently of DNA replication. Smc3 is readily acetylated before replication is initiated and after DNA replication is complete. However, we also show that functional acetylation occurs only in association with the replication machinery: disruption of the interaction between XEco2 and proliferating cell nuclear antigen prevents cohesion establishment while having little impact on the overall levels of Smc3 acetylation. These results demonstrate that Smc3 acetylation can occur throughout interphase but that only acetylation in association with the replication fork promotes sister chromatid cohesion. These data reveal how the generation of cohesion is limited to the appropriate time and place during the cell cycle and provide insight into the mechanism by which acetylation ensures cohesion.

Article	Year
Mutations in transforming growth factor-beta receptor type II cause familial thoracic aortic aneurysms and dissections. Circulation\| PubMed ID: 16027248	2005
Genetic basis of thoracic aortic aneurysms and dissections: focus on smooth muscle cell contractile dysfunction. Annual review of genomics and human genetics\| PubMed ID: 18544034	2008
TGFBR2 mutations alter smooth muscle cell phenotype and predispose to thoracic aortic aneurysms and dissections. Cardiovascular research\| PubMed ID: 20628007	2010
Sororin cooperates with the acetyltransferase Eco2 to ensure DNA replication-dependent sister chromatid cohesion. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 21059905	2010
Cohesin acetylation promotes sister chromatid cohesion only in association with the replication machinery. The Journal of biological chemistry\| PubMed ID: 22896698	2012

Andrea Lafont

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