Studies of factors affecting performance of growing-finishing lambs

This bulletin is a report on Department of Animal Husbandry Research Project 356, 'Lamb fattening'--P. [3].Digitized 2007 AES.Includes bibliographical references (pages 61-65)

Restricted Isometries for Partial Random Circulant Matrices

In the theory of compressed sensing, restricted isometry analysis has become a standard tool for studying how efficiently a measurement matrix acquires information about sparse and compressible signals. Many recovery algorithms are known to succeed when the restricted isometry constants of the sampling matrix are small. Many potential applications of compressed sensing involve a data-acquisition process that proceeds by convolution with a random pulse followed by (nonrandom) subsampling. At present, the theoretical analysis of this measurement technique is lacking. This paper demonstrates that the $s$th order restricted isometry constant is small when the number $m$ of samples satisfies $m \gtrsim (s \log n)^{3/2}$, where $n$ is the length of the pulse. This bound improves on previous estimates, which exhibit quadratic scaling

The brain as 'immunoprecipitator' of serum autoantibodies against N-Methyl-D-Aspartate receptor subunit NR1

Autoantibodies (AB) against N-methyl-D-aspartate receptor subunit NR1 (NMDAR1) are highly seroprevalent in health and disease. Symptomatic relevance may arise upon compromised blood-brain barrier (BBB). However, it remained unknown whether circulating NMDAR1 AB appear in the cerebrospinal fluid (CSF). Of n5271 subjects with CSF-serum pairs, 26 were NMDAR1 AB seropositive, but only 1 was CSF positive. Contrariwise, tetanus AB (non-brain-binding) were present in serum and CSF of all subjects, with CSF levels higher upon BBB dysfunction. Translational mouse experiments proved the hypothesis that the brain acts as an 'immunoprecipitator'; simultaneous injection of NMDAR1 AB and the non-brain-binding green fluorescent protein AB resulted in high detectability of the former in brain and the latter in CSF

Synthesis and characterization of hypoxia-mimicking bioactive glasses for skeletal regeneration

The cellular response to hypoxia (low oxygen pressure) is vital for skeletal tissue development and regeneration. Numerous processes, including progenitor cell recruitment, differentiation and angiogenesis, are activated via the hypoxia pathway. Novel materials-based strategies designed to activate the hypoxia pathway are therefore of great interest for orthopaedic tissue engineering. Resorbable bioactive glasses (BGs) were developed to activate the hypoxia pathway by the controlled release of cobalt ions (at physiological relevant concentrations) whilst controlling BG apatite-forming ability. Two series of soda-lime-phosphosilicate glasses were synthesised with increasing concentrations of cobalt. Compositions were calculated to maintain constant network connectivity (2.13) by considering that cobalt is taking part in the network in the first series, and is acting as a network modifier in the second series. Mg2+ and Zn2+ were added to one of the Co2+-containing glasses to inhibit HCA formation. The presence of HCA formation is undesirable for the use of BG in soft tissues e. g. cartilage. Cobalt was present in both the silicate and phosphate phases of the BG. In addition, evidence was found that it plays a dual role in the silicate phase, entering the network as well as disrupting it as a network modifying oxide. Consistent with this dual role, the presence of cobalt in the BG was shown to decrease ion release. HCA formation was delayed with cobalt addition as well as incorporation of Mg2+ and Zn2+ into the BGs. Importantly, cobalt release was found to be proportional to cobalt content of the BGs enabling the controlled delivery of cobalt in therapeutically active doses

Gelatin microparticles aggregates as three-dimensional scaffolding system in cartilage engineering

A three-dimensional (3D) scaffolding system for chondrocytes culture has been produced by agglomeration of cells and gelatin microparticles with a mild centrifuging process. The diameter of the microparticles, around 10 μ, was selected to be in the order of magnitude of the chondrocytes. No gel was used to stabilize the construct that maintained consistency just because of cell and extracellular matrix (ECM) adhesion to the substrate. In one series of samples the microparticles were charged with transforming growth factor, TGF-β1. The kinetics of growth factor delivery was assessed. The initial delivery was approximately 48 % of the total amount delivered up to day 14. Chondrocytes that had been previously expanded in monolayer culture, and thus dedifferentiated, adopted in this 3D environment a round morphology, both with presence or absence of growth factor delivery, with production of ECM that intermingles with gelatin particles. The pellet was stable from the first day of culture. Cell viability was assessed by MTS assay, showing higher absorption values in the cell/unloaded gelatin microparticle pellets than in cell pellets up to day 7. Nevertheless the absorption drops in the following culture times. On the contrary the cell viability of cell/TGF-β1 loaded gelatin microparticle pellets was constant during the 21 days of culture. The formation of actin stress fibres in the cytoskeleton and type I collagen expression was significantly reduced in both cell/gelatin microparticle pellets (with and without TGF-β1) with respect to cell pellet controls. Total type II collagen and sulphated glycosaminoglycans quantification show an enhancement of the production of ECM when TGF-β1 is delivered, as expected because this growth factor stimulate the chondrocyte proliferation and improve the functionality of the tissue.JLGR acknowledge the support of the Spanish Ministry of Education through project No. MAT2010-21611-C03-01 (including the FEDER financial support). The support of the Instituto de Salud Carlos III (ISCIII) through the CIBER initiative of the Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) is also acknowledged

Functional Reconstitution of a Tunable E3-Dependent Sumoylation Pathway in Escherichia coli

SUMO (small ubiquitin-related modifier) is a reversible post-translational protein modifier that alters the localization, activity, or stability of proteins to which it is attached. Many enzymes participate in regulated SUMO-conjugation and SUMO-deconjugation pathways. Hundreds of SUMO targets are currently known, with the majority being nuclear proteins. However, the dynamic and reversible nature of this modification and the large number of natively sumoylated proteins in eukaryotic proteomes makes molecular dissection of sumoylation in eukaryotic cells challenging. Here, we have reconstituted a complete mammalian SUMO-conjugation cascade in Escherichia coli cells that involves a functional SUMO E3 ligase, which effectively biases the sumoylation of both native and engineered substrate proteins. Our sumo-engineered E. coli cells have several advantages including efficient protein conjugation and physiologically relevant sumoylation patterns. Overall, this system provides a rapid and controllable platform for studying the enzymology of the entire sumoylation cascade directly in living cells

A Novel ‘Gene Insertion/Marker Out’ (GIMO) Method for Transgene Expression and Gene Complementation in Rodent Malaria Parasites

Research on the biology of malaria parasites has greatly benefited from the application of reverse genetic technologies, in particular through the analysis of gene deletion mutants and studies on transgenic parasites that express heterologous or mutated proteins. However, transfection in Plasmodium is limited by the paucity of drug-selectable markers that hampers subsequent genetic modification of the same mutant. We report the development of a novel ‘gene insertion/marker out’ (GIMO) method for two rodent malaria parasites, which uses negative selection to rapidly generate transgenic mutants ready for subsequent modifications. We have created reference mother lines for both P. berghei ANKA and P. yoelii 17XNL that serve as recipient parasites for GIMO-transfection. Compared to existing protocols GIMO-transfection greatly simplifies and speeds up the generation of mutants expressing heterologous proteins, free of drug-resistance genes, and requires far fewer laboratory animals. In addition we demonstrate that GIMO-transfection is also a simple and fast method for genetic complementation of mutants with a gene deletion or mutation. The implementation of GIMO-transfection procedures should greatly enhance Plasmodium reverse-genetic research

SUMO modification of PCNA is controlled by DNA

Post-translational modification by the ubiquitin-like protein SUMO is often regulated by cellular signals that restrict the modification to appropriate situations. Nevertheless, many SUMO-specific ligases do not exhibit much target specificity, and—compared with the diversity of sumoylation substrates—their number is limited. This raises the question of how SUMO conjugation is controlled in vivo. We report here an unexpected mechanism by which sumoylation of the replication clamp protein, PCNA, from budding yeast is effectively coupled to S phase. We find that loading of PCNA onto DNA is a prerequisite for sumoylation in vivo and greatly stimulates modification in vitro. To our surprise, however, DNA binding by the ligase Siz1, responsible for PCNA sumoylation, is not strictly required. Instead, the stimulatory effect of DNA on conjugation is mainly attributable to DNA binding of PCNA itself. These findings imply a change in the properties of PCNA upon loading that enhances its capacity to be sumoylated

Mutations at the Subunit Interface of Yeast Proliferating Cell Nuclear Antigen Reveal a Versatile Regulatory Domain

Acknowledgments We thank Szilvia Minorits for technical assistance. I.U. conceived and designed the project and wrote the manuscript. All authors participated in designing and performing the experiments, and analyzing the results. The authors declare no competing financial interests. This work was also supported by a grant from the National Research, Development and Innovation Office GINOP-2.3.2-15-2016-00001. Funding: This work was supported by Hungarian Science Foundation Grant OTKA 109521 and National Research Development and Innovation Office GINOP-2.3.2-15-2016-00001. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

Cohesin Is Limiting for the Suppression of DNA Damage–Induced Recombination between Homologous Chromosomes

Double-strand break (DSB) repair through homologous recombination (HR) is an evolutionarily conserved process that is generally error-free. The risk to genome stability posed by nonallelic recombination or loss-of-heterozygosity could be reduced by confining HR to sister chromatids, thereby preventing recombination between homologous chromosomes. Here we show that the sister chromatid cohesion complex (cohesin) is a limiting factor in the control of DSB repair and genome stability and that it suppresses DNA damage–induced interactions between homologues. We developed a gene dosage system in tetraploid yeast to address limitations on various essential components in DSB repair and HR. Unlike RAD50 and RAD51, which play a direct role in HR, a 4-fold reduction in the number of essential MCD1 sister chromatid cohesion subunit genes affected survival of gamma-irradiated G2/M cells. The decreased survival reflected a reduction in DSB repair. Importantly, HR between homologous chromosomes was strongly increased by ionizing radiation in G2/M cells with a single copy of MCD1 or SMC3 even at radiation doses where survival was high and DSB repair was efficient. The increased recombination also extended to nonlethal doses of UV, which did not induce DSBs. The DNA damage–induced recombinants in G2/M cells included crossovers. Thus, the cohesin complex has a dual role in protecting chromosome integrity: it promotes DSB repair and recombination between sister chromatids, and it suppresses damage-induced recombination between homologues. The effects of limited amounts of Mcd1and Smc3 indicate that small changes in cohesin levels may increase the risk of genome instability, which may lead to genetic diseases and cancer