The C-Terminal Domain of CENP-C Displays Multiple and Critical Functions for Mammalian Centromere Formation

CENP-C is a fundamental component of functional centromeres. The elucidation of its structure-function relationship with centromeric DNA and other kinetochore proteins is critical to the understanding of centromere assembly. CENP-C carries two regions, the central and the C-terminal domains, both of which are important for the ability of CENP-C to associate with the centromeric DNA. However, while the central region is largely divergent in CENP-C homologues, the C-terminal moiety contains two regions that are highly conserved from yeast to humans, named Mif2p homology domains (blocks II and III). The activity of these two domains in human CENP-C is not well defined. In this study we performed a functional dissection of C-terminal CENP-C region analyzing the role of single Mif2p homology domains through in vivo and in vitro assays. By immunofluorescence and Chromatin immunoprecipitation assay (ChIP) we were able to elucidate the ability of the Mif2p homology domain II to target centromere and contact alpha satellite DNA. We also investigate the interactions with other conserved inner kinetochore proteins by means of coimmunoprecipitation and bimolecular fluorescence complementation on cell nuclei. We found that the C-terminal region of CENP-C (Mif2p homology domain III) displays multiple activities ranging from the ability to form higher order structures like homo-dimers and homo-oligomers, to mediate interaction with CENP-A and histone H3. Overall, our findings support a model in which the Mif2p homology domains of CENP-C, by virtue of their ability to establish multiple contacts with DNA and centromere proteins, play a critical role in the structuring of kinethocore chromatin

Comparative Analysis of Tandem Repeats from Hundreds of Species Reveals Unique Insights into Centromere Evolution

Centromeres are essential for chromosome segregation, yet their DNA sequences evolve rapidly. In most animals and plants that have been studied, centromeres contain megabase-scale arrays of tandem repeats. Despite their importance, very little is known about the degree to which centromere tandem repeats share common properties between different species across different phyla. We used bioinformatic methods to identify high-copy tandem repeats from 282 species using publicly available genomic sequence and our own data. The assumption that the most abundant tandem repeat is the centromere DNA was true for most species whose centromeres have been previously characterized, suggesting this is a general property of genomes. Our methods are compatible with all current sequencing technologies. Long Pacific Biosciences sequence reads allowed us to find tandem repeat monomers up to 1,419 bp. High-copy centromere tandem repeats were found in almost all animal and plant genomes, but repeat monomers were highly variable in sequence composition and in length. Furthermore, phylogenetic analysis of sequence homology showed little evidence of sequence conservation beyond ~50 million years of divergence. We find that despite an overall lack of sequence conservation, centromere tandem repeats from diverse species showed similar modes of evolution, including the appearance of higher order repeat structures in which several polymorphic monomers make up a larger repeating unit. While centromere position in most eukaryotes is epigenetically determined, our results indicate that tandem repeats are highly prevalent at centromeres of both animals and plants. This suggests a functional role for such repeats, perhaps in promoting concerted evolution of centromere DNA across chromosomes

Assessment of a pragmatic strategy to improve health of kacang goats in subsistence agricultural communities in Indonesian Borneo

Poverty limits options available to smallholder, subsistence farmers to prevent or reverse livestock malnutrition and endoparasitism, two of the global drivers of goat morbidity and mortality in resource-constrained, tropical, agricultural communities. Our first study objectives describe changes observed in body condition and anaemia after implementation of three feasible and simple husbandry changes to improve health of smallholder herds of kacang goats in rural, Indonesian Borneo. These changes included routine hoof trimming and increased access to food and fresh water. We observed an impressive six-fold decrease in emaciated animals from 26 % to 4 % and an almost doubling of goats in ideal body condition from 29 % to 54 % after fourteen months of improved hoof care and nutrition. The second study objective described herd health changes observed fourteen months after adding a targeted, selective, refugia deworming regimen to the enhanced husbandry program. We observed a significant decrease in proportion of anaemic goats from 88 % to 74 % fourteen months after initiating the targeted selective herd anthelmintic treatment. Impoverished, smallholder subsistence agricultural communities with limited resources should first initiate feasible husbandry enhancements to begin improving overall herd health especially when anthelmintic expense or availability delays establishing an ideal program which includes a deworming component

By the rivers of Babylon: patterns of heterarchy, sustainable wetland agroecology, and urban dynamics in old Babylonian Mashkan-shapir

Archaeological investigations of the largest urban centers in southern Mesopotamia have excluded collection and detailed interpretation of faunal remains. This exclusion has resulted in a biased interpretation of urban dynamics based largely on architecture, site planning, artifact distribution, and textual evidence. The samples that do exist from these sites are often incomplete. Additionally, textual evidence pertaining to animal exploitation is essentially silent when it comes to pig husbandry and offers little information on the exploitation of fish and other wild resources. While addressing these biases with the analysis of faunal material from the late second millennium (BCE) urban site of Mashkan-shapir, this study also aims to shed light on the complex interplay between urban life and the natural diversity of the southern Mesopotamian wetlands. The site is presented as a model for heterarchical sociopolitical organization and sustainable agroecological approach to subsistence. A fundamental link is made between sustainability and heterarchical organization and consensus. Results of the analysis of over 7000 specimens from excavation, as well as nearly 2900 specimens from systematic flotation, indicate that wetland resources were an integral part of the site economy. The data suggest pigs were a major dietary component, and suggest low intensity cultivation of free roaming "street pigs" as the likely pig rearing strategy. Ovicaprid remains indicate a strong bias towards sheep with the primary goal of wool production. The study attempts to describe and quantify the role of wetlands as a sustainable resource adding to the vitality and success of Mashkan-shapir. The data suggest an urban setting intimately linked to wetland ecosystems. This model of wetland exploitation is compared to both ancient and modern data including modern models of mixed species sustainable agroecosystems to illustrate the efficiency and sustainability of the proposed Mashkan-shapir model. The data from Mashkan-shapir suggest that a heretofore unexamined or hidden portion of the economy based on fishing, hunting, household level pig husbandry, and wetland resource exploitation, played a crucial role in the lives of Mesopotamian urbanites

Histone lysine methylation in the context of nuclear architecture

The impact of histone lysine methylation as an essential epigenetic mechanismn for gene regulation has been demonstrated by numerous studies where it was functionally and structurally linked to euchromatin and heterochromatin.In this work the 3D architecture and spatial interrealtionships of different histone lysine methylation sites was investigated in various human cell types

Role of extracellular matrix in muscle repair and regeneration

Extracellular matrices (ECMs) play important structural and mechanical roles in muscle tissue. They are also critical for normal muscle homeostasis, but it is poorly understood how individual matrix proteins, or the mix of ECM proteins found in vivo affect myoblast behaviour. Aspects of this question have been examined in this thesis using both matrices from skeletal muscle and artificial scaffolds prepared from silk proteins. The ability of these surfaces to support myoblast proliferation and differentiation was examined using a murine myoblast cell line and primary human skeletal muscle myoblasts