Cellular localization and allele-selective inhibition of mutant huntingtin protein by peptide nucleic acid oligomers containing the fluorescent nucleobase [bis-o-(aminoethoxy)phenyl]pyrrolocytosine

Peptide nucleic acid (PNA) is a successful DNA/RNA mimic. A major challenge for research is to invent chemically modified PNAs that retain the favorable properties of the parent compound while improving biological recognition. Here, we test modified PNAs containing [bis-o-(aminoethoxy)phenyl]pyrrolocytosine bases designed to engage guanine with an additional hydrogen bond. We observe elevated melting temperatures, localization to cellular compartments, and allele-selective inhibition of mutant huntingtin protein expression.     

Endogenous clock gene expression in the suprachiasmatic nuclei of previsual newborn rabbits is entrained by nursing

The rabbit is particularly suitable for investigating the development of mammalian circadian function. Blind at birth, the pups are only visited by the mother to be nursed once every 24 h for about 3 min and so can be studied largely without maternal interference. They anticipate the mother's visit with increased behavioral arousal and with a rise in body temperature, both of which represent endogenous circadian rhythms. We now report that in newborn pups the suprachiasmatic nuclei of the hypothalamus (SCN; the main circadian pacemaker in mammals) show endogenous 24‐h rhythmicity in the expression of the clock genes Per1, Per2, and Bmal1. Pups nursed from postnatal days 1 to 7 and fasted to day 9 showed the same rhythms of clock gene expression as normally nursed controls. We also report that these rhythms are entrained by nursing. Pups killed on postnatal days 3–4 showed the same rhythms in gene expression as pups in the previous experiment, whereas littermates subsequently nursed from postnatal days 4 to 7 with nursing delayed 6 h showed a corresponding shift in the diurnal pattern of clock gene expression. Consistent with this, two groups of pups implanted with telemetric thermal sensors and nursed 6 h apart had daily patterns in body temperature synchronized with the two different nursing times. We conclude that the expression of clock genes associated with the newborn rabbit's circadian system is entrained by nonphotic cues accompanying nursing, the exact nature of which now needs to be clarified. © 2008 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

Genetic Variation in the Familial Mediterranean Fever Gene (MEFV) and Risk for Crohn's Disease and Ulcerative Colitis

Background and Aims

The familial Mediterranean fever (FMF) gene (MEFV) encodes pyrin, a major regulator of the inflammasome platform controlling caspase-1 activation and IL-1β processing. Pyrin has been shown to interact with the gene product of NLRP3, NALP3/cryopyrin, also an important active member of the inflammasome. The NLRP3 region was recently reported to be associated with Crohn''s disease (CD) susceptibility. We therefore sought to evaluate MEFV as an inflammatory bowel disease (IBD) susceptibility gene.

Methodology and Results

MEFV colonic mucosal gene expression was significantly increased in experimental colitis mice models (TNBS p<0.0003; DSS p<0.006), in biopsies from CD (p<0.02) and severe ulcerative colitis (UC) patients (p<0.008). Comprehensive genetic screening of the MEFV region in the Belgian exploratory sample set (440 CD trios, 137 UC trios, 239 CD cases, 96 UC cases, and 107 healthy controls) identified SNPs located in the MEFV 5′ haplotype block that were significantly associated with UC (rs224217; p = 0.003; A allele frequency: 56% cases, 45% controls), while no CD associations were observed. Sequencing and subsequent genotyping of variants located in this associated haplotype block identified three synonymous variants (D102D/rs224225, G138G/rs224224, A165A/rs224223) and one non-synonymous variant (R202Q/rs224222) located in MEFV exon 2 that were significantly associated with UC (rs224222: p = 0.0005; A allele frequency: 32% in cases, 23% in controls). No consistent associations were observed in additional Canadian (256 CD trios, 91 UC trios) and Scottish (495 UC, 370 controls) sample sets. We note that rs224222 showed marginal association (p = 0.012; G allele frequency: 82% in cases, 70% in controls) in the Canadian sample, but with a different risk allele. None of the NLRP3 common variants were associated with UC in the Belgian-Canadian UC samples and no significant interactions were observed between NLRP3 and MEFV that could explain the observed flip-flop of the rs224222 risk allele.

Conclusion

The differences in association levels observed between the sample sets may be a consequence of distinct founder effects or of the relative small sample size of the cohorts evaluated in this study. However, the results suggest that common variants in the MEFV region do not contribute to CD and UC susceptibility.     

Impact of egg harvesting on breeding success of black-headed gulls,Larus ridibundus

Gull colonies world-wide have been harvested for their eggs for centuries with minimal knowledge of the impacts on breeding. Although most Laridae can replace lost eggs, they have comparatively high energetic demands for egg production. In this paper we assess the impacts of a licensed egg harvest on the breeding success of black-headed gulls Larus ridibundus, which nest colonially in an EU Special Protection Area in Hampshire, southern England. We compared egg volume, hatching and chick survival from harvested and un-harvested nests in central and fringe positions within colonies of various sizes, including colonies with no harvesting activity. Eggs from various laying stages were collected from harvested and un-harvested colonies of similar pre-harvest intrinsic quality, for comparison of their volumes, yolk-to-albumen ratios and eggshell thickness. Egg volume and the yolk-to-albumen ratio depended on laying time and location, with the largest eggs laid during the peak period by birds breeding in central positions on large colonies. Eggs produced by these peak layers also had the largest yolk-to-albumen ratios. Harvested sites were characterised by reductions in egg volume, yolk-to-albumen ratio and eggshell thickness, which translated to poorer hatching success and chick survival. Harvested sites also had a higher proportion of abnormal eggs, particularly taking the forms of small yolkless eggs and unpigmented eggs. The reduced breeding success on harvested colonies is likely to be linked to depletion of the female's endogenous reserves which can also reduce future survival and breeding propensity.     

Evolution of Allometry in Antirrhinum

Correlated variation in shape and size (allometry) is a major component of natural diversity. We examined the evolutionary and genetic basis for allometry using leaves and flower petals of snapdragon species (Antirrhinum). A computational method was developed to capture shape and size variation in both types of organ within the Antirrhinum species group. The results show that the major component of variation between species involves positively correlated changes in leaf and petal size. The correlation was maintained in an F2 population derived from crossing two species with organs of different sizes, suggesting that developmental constraints were involved. Identification of the underlying genes as quantitative trait loci revealed that the larger species carried alleles that increased organ size at all loci. Although this was initially taken as evidence that directional selection has driven diversity in both leaf and petal size, simulations revealed that evolution without consistent directional selection, an undirected walk, could also account for the parental distribution of organ size alleles.     

Occasional intraguild predation structuring small mammal assemblages: the marsupial Didelphis aurita in the Atlantic Forest of Brazil

The didelphid marsupial, Didelphis aurita, is suggested as an intraguild predator and as key‐species in small mammal assemblages of the Atlantic Forest of Brazil. The field experiments required to test this hypothesis are complex to implement, but the recent revival of regression methods offers a viable alternative. Here we use the dynamic and static regression methods to determine the importance of D. aurita as a competitor and intraguild predator. Capture–recapture data from two localities in the Rio de Janeiro State were used, Garrafão (municipality of Guapimirim), a coastal forest of the Serra do Mar, and Barra de Maricá, a costal sand dune vegetation. Population and microhabitat variables were monitored from April 1997 to April 2003 in Garrafão, and from January 1986 to July 1990 in Barra de Maricá. Microhabitat variables were related to Canopy, Plant, Litter and Rock covers, Obstruction from 0 to 1.5 m, and Number of logs. Exploitation competition was tested by the dynamic method, which models the effects of D. aurita on the per capita growth rate of a species. Interference by predation or competition was tested by the static method, where the abundance of D. aurita at trap stations was regressed against the abundance of other small mammals, after removal of any variation associated with microhabitat factors. Exploitation competition was not detected, but the interference of D. aurita was pervasive, affecting all small mammals studied in the two localities. The clear avoidance of D. aurita by all small mammals tested in two localities of different physiognomies indicates that it functions as an intraguild predator, even if actual predation by D. aurita is an occasional event.     

Expression and evolutionary conservation of the tescalcin gene during development

The tescalcin gene (Tesc) encodes an EF-hand calcium-binding protein that interacts with the sodium/hydrogen exchanger, NHE1. Previous studies indicated that Tesc was expressed in mouse embryonic testis, but not in ovary, during the critical period of testis and ovary determination. In this paper we compared the expression of Tesc in embryonic tissues of chicken and mouse. Tesc expression was sexually dimorphic in the embryonic gonads of both mouse and chicken. Tescalcin (TESC) was detected in both Sertoli cells and germ cells. In the embryonic brain of both mouse and chicken, Tesc was highly expressed in the nasal placode and in fibers extending from the olfactory epithelium to the primordial olfactory bulb. Tesc was expressed in the embryonic heart of both chicken and mouse. In mouse Tesc expression was also detected in embryonic adrenal. These studies indicate very specific expression of Tesc in various tissues in chicken and mouse during embryologic development, and conservation of Tesc expression in both species.     

Fine mapping the soybean aphid resistance gene Rag1 in soybean

The soybean aphid (Aphis glycines Matsumura) is an important soybean [Glycine max (L.) Merr.] pest in North America. The dominant aphid resistance gene Rag1 was previously mapped from the cultivar ‘Dowling’ to a 12 cM marker interval on soybean chromosome 7 (formerly linkage group M). The development of additional genetic markers mapping closer to Rag1 was needed to accurately position the gene to improve the effectiveness of marker-assisted selection (MAS) and to eventually clone it. The objectives of this study were to identify single nucleotide polymorphisms (SNPs) near Rag1 and to position these SNPs relative to Rag1. To generate a fine map of the Rag1 interval, 824 BC₄F₂ and 1,000 BC₄F₃ plants segregating for the gene were screened with markers flanking Rag1. Plants with recombination events close to the gene were tested with SNPs identified in previous studies along with new SNPs identified from the preliminary Williams 82 draft soybean genome shotgun sequence using direct re-sequencing and gene-scanning melt-curve analysis. Progeny of these recombinant plants were evaluated for aphid resistance. These efforts resulted in the mapping of Rag1 between the two SNP markers 46169.7 and 21A, which corresponds to a physical distance on the Williams 82 8× draft assembly (Glyma1.01) of 115 kilobase pair (kb). Several candidate genes for Rag1 are present within the 115-kb interval. The markers identified in this study that are closely linked to Rag1 will be a useful resource in MAS for this important aphid resistance gene.     

Carboxylated N‐glycans on RAGE promote S100A12 binding and signaling

The receptor for advanced glycation end products (RAGE) is a signaling receptor protein of the immunoglobulin superfamily implicated in multiple pathologies. It binds a diverse repertoire of ligands, but the structural basis for the interaction of different ligands is not well understood. We earlier showed that carboxylated glycans on the V‐domain of RAGE promote the binding of HMGB1 and S100A8/A9. Here we study the role of these glycans on the binding and intracellular signaling mediated by another RAGE ligand, S100A12. S100A12 binds carboxylated glycans, and a subpopulation of RAGE enriched for carboxylated glycans shows more than 10‐fold higher binding potential for S100A12 than total RAGE. When expressed in mammalian cells, RAGE is modified by complex glycans predominantly at the first glycosylation site (N25IT) that retains S100A12 binding. Glycosylation of RAGE and maximum binding sites for S100A12 on RAGE are also cell type dependent. Carboxylated glycan‐enriched population of RAGE forms higher order multimeric complexes with S100A12, and this ability to multimerize is reduced upon deglycosylation or by using non‐glycosylated sRAGE expressed in E. coli. mAbGB3.1, an antibody against carboxylated glycans, blocks S100A12‐mediated NF‐κB signaling in HeLa cells expressing full‐length RAGE. These results demonstrate that carboxylated N‐glycans on RAGE enhance binding potential and promote receptor clustering and subsequent signaling events following oligomeric S100A12 binding. J. Cell. Biochem. 110: 645–659, 2010. © 2010 Wiley‐Liss, Inc.     

Evidence that αC region is origin of low modulus, high extensibility, and strain stiffening in fibrin fibers

Fibrin fibers form the structural scaffold of blood clots and perform the mechanical task of stemming blood flow. Several decades of investigation of fibrin fiber networks using macroscopic techniques have revealed remarkable mechanical properties. More recently, the microscopic origins of fibrin's mechanics have been probed through direct measurements on single fibrin fibers and individual fibrinogen molecules. Using a nanomanipulation system, we investigated the mechanical properties of individual fibrin fibers. The fibers were stretched with the atomic force microscope, and stress-versus-strain data was collected for fibers formed with and without ligation by the activated transglutaminase factor XIII (FXIIIa). We observed that ligation with FXIIIa nearly doubled the stiffness of the fibers. The stress-versus-strain behavior indicates that fibrin fibers exhibit properties similar to other elastomeric biopolymers. We propose a mechanical model that fits our observed force extension data, is consistent with the results of the ligation data, and suggests that the large observed extensibility in fibrin fibers is mediated by the natively unfolded regions of the molecule. Although some models attribute fibrin's force-versus-extension behavior to unfolding of structured regions within the monomer, our analysis argues that these models are inconsistent with the measured extensibility and elastic modulus.