Inclusion of soil organic carbon changes can reverse results of historical energy flux quantifications of Pampean agriculture.

The questions that guided this work were: 1) How do energy inputs, outputs, and energy indices evolve during the last four decades along the rainfall gradient of the Pampas, 2) How does present agrochemical and fertilizer use in Argentina resemble, or differ, from other main grain producing regions with large no-till surfaces?, and 3) How do energy fluxes vary when soil organic carbon (SOC) changes during the last four decades are included? Energy balances (outputs - inputs), energy efficiencies (outputs/inputs) and energy intensities (inputs/yield) were calculated. Inputs comprised agrochemicals and fertilizers, machinery used for soil tillage and fuel use and gathered from different information sources. Outputs included yield of main crops from national statistics. Calculations were performed for four areas along a rainfall gradient during the 1970-2015 period. Energy coefficients were collected from literature. Soil organic carbon changes of the upper soil profile meter were available from a previous publication. Total input averages per area were low although increased 62% after four decades, from 6.6 GJ ha-1 in the past up to 10.5 GJ ha-1 at present with no marked differences between areas. Agrochemicals comprised 49% of total energy input, a very large proportion compared to other regions mainly related to the large surface under no-tillage while fertilizer rates were low. Average energy outputs increased 51% with time and all energy balances were positive. Energy efficiencies had an optimum during 1995 of 4.8 decreasing afterwards down to 3.7. Energy intensities decreased and at present 14% less input energy was needed per t DM yield produced. Two areas gained SOC and one lost large amounts. Inclusion of SOC losses in energy quantifications turned all energy indices to negative values therefore providing a real scenario of what happened with energy fluxes after four decades of agriculture which otherwise would be ignored.

Identification of novel alleles associated with insulin resistance in childhood obesity using pooled-DNA genome-wide association study approach.

BACKGROUND: Recently, we witnessed great progress in the discovery of genetic variants associated with obesity and type 2 diabetes (T2D), especially in adults. Much less is known regarding genetic variants associated with insulin resistance (IR). We hypothesized that novel IR genes could be efficiently detected in a population of obese children and adolescents who may not exhibit comorbidities and other confounding factors. OBJECTIVES: This study aimed to determine whether a genome-wide association study (GWAS), using a DNA-pooling approach, could identify novel genes associated with IR. SUBJECTS: The pooled-DNA GWAS analysis included Slovenian obese children and adolescents with and without IR matched for body mass index, gender and age. A replication study was conducted in another independent cohort with or without IR. METHODS: For the pooled-DNA GWAS, we used HumanOmni5-Quad SNP array (Illumina). Allele frequency distributions were compared with modified t-tests and χ2-tests and ranked using PLINK. Top single nucleotide polymorphisms (SNPs) were validated using individual genotyping by high-resolution melting analysis and TaqMan assay. RESULTS: We identified five top-ranking SNPs from the pooled-DNA GWAS analysis within the ECE1, IL1R2, GNPDA1, HLA-J and PYGB loci. All except SNP rs9261108 (HLA-J locus) were confirmed in the validation phase using individual genotyping. The SNP rs2258617 within PYGB remained statistically significant for both recessive and additive models in both cohorts and in a merged analysis of both cohorts and present the strongest novel candidate gene for IR. CONCLUSION: We report for the first time a pooled-DNA GWAS approach to identify five novel SNPs or genes for IR in a paediatric population. The four loci confirmed in the second validation phase study warrant further studies, especially the strongest SNP rs2258617 within PYGB, and provide targets for further basic research of IR mechanisms and for the development of potential new IR and T2D therapies.

HIF1A gene polymorphisms and human diseases: Graphical review of 97 association studies.

Hypoxia-inducible factors (HIFs) belong to a family of transcription factors (TF) responsive to a low O2 availability, which is often a characteristic feature of solid tumors. The alpha subunit of the HIF heterodimer is O2 -sensitive, and once stabilized in hypoxia, it functions as a master regulator of various genes involved in hypoxia pathway. Changes in the HIF1A (hypoxia inducible factor 1, alpha subunit) nucleotide sequence or expression has been shown to be associated with the development of several diseases. Because of increasing research interest in HIF1A gene a review of association studies was needed. We here reviewed published data on single nucleotide polymorphisms (SNPs) in HIF1A in various diseases; in total, 34 SNPs were tested for an association with 49 phenotypes, and the results were visualized using the Cytoscape software. Among all collected polymorphisms 16 SNPs showed significant associations with 40 different phenotypes, including six SNPs associated with 14 cancer types. Missense SNPs (rs11549465 and rs11549467) within the oxygen-dependent degradation domain were most frequently studied. The study provides a comprehensive tool for researchers working in this area and may contribute to more accurate disease diagnosis and identification of therapeutic targets.

FTO gene variants are associated with growth and carcass traits in cattle.

An important aim in animal breeding is the improvement of growth and meat quality traits. Previous studies have demonstrated that genetic variants in the fat mass and obesity associated (FTO) gene have a relatively large effect on human obesity as well as on body composition in rodents and, more recently, in livestock. Here, we examined the effects of the FTO gene variants on growth and carcass traits in the Slovenian population of Simmental (SS) and Brown (SB) cattle. To validate and identify new polymorphisms, we used sequencing, PCR-RFLP analysis and TaqMan assays in the SS breed and FTO gene variants data from the Illumina BovineSNP50 v1 array for the SB breed. Sequencing of the eight samples of progeny-tested SS sires detected 108 single nucleotide polymorphisms (SNPs) in the bovine FTO gene. Statistical analyses between growth and carcass traits and 34 FTO polymorphisms revealed significant association of FTO variants with lean meat percentage in both breeds. Additionally, FTO SNPs analyzed in SS cattle were associated with fat percentage, bone weight and live weight at slaughter. The FTO gene can thus be regarded as a candidate gene for the marker-assisted selection programs in our and possibly other populations of cattle. Future studies in cattle might reveal novel roles for the FTO gene in shaping carcass traits in livestock species as well as body composition control in other mammals.

Administration of probiotics Lactobacillus rhamnosus GG and Lactobacillus gasseri K7 during pregnancy and lactation changes mouse mesenteric lymph nodes and mammary gland microbiota.

The milk and mammary gland (MG) microbiome can be influenced by several factors, such as mode of delivery, breastfeeding, maternal lifestyle, health status, and diet. An increasing number of studies show a variety of positive effects of consumption of probiotics during pregnancy and breastfeeding on the mother and the newborn. The aim of this study was to investigate the effect of oral administration of probiotics Lactobacillus gasseri K7 (LK7) and Lactobacillus rhamnosus GG (LGG) during pregnancy and lactation on microbiota of the mouse mesenteric lymph nodes (MLN), MG, and milk. Pregnant FVB/N mice were fed skim milk or probiotics LGG or LK7 resuspended in skim milk during gestation and lactation. On d 3 and 8 postpartum, blood, feces, MLN, MG, and milk were analyzed for the presence of LGG or LK7. The effects of probiotics on MLN, MG, and milk microbiota was evaluated by real-time PCR and by 16S ribosomal DNA 454-pyrosequencing. In 5 of 8 fecal samples from the LGG group and in 5 of 8 fecal samples from the LK7 group, more than 1 × 10(3) of live LGG or LK7 bacterial cells were detected, respectively, whereas no viable LGG or LK7 cells were detected in the control group. Live lactic acid bacteria but no LGG or LK7 were detected in blood, MLN, and MG. Both probiotics significantly increased the total bacterial load as assessed by copies of 16S ribosomal DNA in MLN, and a similar trend was observed in MG. Metagenomic sequencing revealed that both probiotics increased the abundance of Firmicutes in MG, especially the abundance of lactic acid bacteria. The Lactobacillus genus appeared exclusively in MG from probiotic groups. Both probiotics influenced MLN microbiota by decreasing diversity (Chao1) and increasing the distribution of species (Shannon index). The LGG probiotic also affected the MG microbiota as it increased diversity and distribution of species and proportions of the genera Lactobacillus and Bifidobacterium. These results provide evidence that probiotics can modulate the bacterial composition of MLN and MG microbiota in ways that could improve the health of the MG and, ultimately, the health of the newborn.

Genome-wide in silico screening for microRNA genetic variability in livestock species.

MicroRNAs are a class of non-coding RNAs that post-transcriptionally regulate target gene expression. Previous studies have shown that microRNA gene variability can interfere with its function, resulting in phenotypic variation. Polymorphisms within microRNA genes present a source of novel biomarkers for phenotypic traits in animal breeding. However, little is known about microRNA genetic variability in livestock species, which is also due to incomplete data in genomic resource databases. Therefore, the aim of this study was to perform a genome-wide in silico screening of genomic sources and determine the genetic variability of microRNA genes in livestock species using mirna sniper 3.0 (http://www.integratomics-time.com/miRNA-SNiPer/), a new version of our previously developed tool. By examining Ensembl and miRBase genome builds, it was possible to design a tool-based generated search of 16 genomes including four livestock species: pig, horse, cattle and chicken. The analysis revealed 65 polymorphisms located within mature microRNA regions in these four species, including 28% within the seed region in cattle and chicken. Polymorphic microRNA genes in cattle and chicken were further examined for mapping to quantitative trait loci regions associated with production and health traits. The developed bioinformatics tool enables the analysis of polymorphic microRNA genes and prioritization of potential regulatory polymorphisms and therefore contributes to the development of microRNA-based biomarkers in livestock species. The assembled catalog and the developed tool can serve the animal science community to efficiently select microRNA SNPs for further quantitative and molecular genetic evaluations of their phenotypic effects and causal associations with livestock production traits.

Computational structure-activity study directs synthesis of novel antitumor enkephalin analogs.

The capability of a Support Vector Machines QSAR model to predict the antiproliferative ability of small peptides was evaluated by screening a virtual library of enkephalin-like analogs modified by incorporation of the (R,S)-(1-adamantyl)glycine (Aaa) residue. From an initial set of 390 compounds, the peptides, Tyr-Aaa-Gly-Phe-Met (2), Tyr-Aaa-Gly-Phe-Phe (3), Phe-Aaa-Gly-Phe-Phe (4) and Phe-Aaa-Gly-Phe-Met (5) were selected, synthesized and their antitumor activity was tested and compared to that of Met-enkephalin (1). The antiproliferative activity correlated with the computational prediction and with the foldamer-forming ability of the studied peptides. The most active compounds were the hydrophobic peptides, Phe-Aaa-Gly-Phe-Phe (4) and Phe-Aaa-Gly-Phe-Met (5), having a greater propensity to adopt folded structures than the other peptides.

Identification of chromosomal regions associated with growth and carcass traits in an F(3) full sib intercross line originating from a cross of chicken lines divergently selected on body weight.

An F(3) resource population originating from a cross between two divergently selected lines for high (D+ line) or low (D- line) body weight at 8-weeks of age (BW55) was generated and used for Quantitative Trait Locus (QTL) mapping. From an initial cross of two founder F(0) animals from D(+) and D(-) lines, progeny were randomly intercrossed over two generations following a full sib intercross line (FSIL) design. One hundred and seventy-five genome-wide polymorphic markers were employed in the DNA pooling and selective genotyping of F(3) to identify markers with significant effects on BW55. Fifty-three markers on GGA2, 5 and 11 were then genotyped in the whole F(3) population of 503 birds, where interval mapping with GridQTL software was employed. Eighteen QTL for body weight, carcass traits and some internal organ weights were identified. On GGA2, a comparison between 2-QTL vs. 1-QTL analysis revealed two separate QTL regions for body, feet, breast muscle and carcass weight. Given co-localization of QTL for some highly correlated traits, we concluded that there were 11 distinct QTL mapped. Four QTL localized to already mapped QTL from other studies, but seven QTL have not been previously reported and are hence novel and unique to our selection line. This study provides a low resolution QTL map for various traits and establishes a genetic resource for future fine-mapping and positional cloning in the advanced FSIL generations.

Co-expression of p21(Waf1/Cip1) in adenovirus vectors improves expression of a second transgene.

First-generation adenoviral (Ad) vectors are frequently used vectors for experimental and clinical gene transfer. Earlier it has been shown that parallel overexpression of the cell cycle regulator p21(Waf1/Cip1) (p21) or antiapoptotic bcl-2 from a second vector reduces cytotoxicity and improves transgene expression. Here, we investigate whether the co-expression of p21 and alpha(1)-antitrypsin from a single vector improves vector safety and alpha(1)-antitrypsin expression. Cell lines (A549 and HeLa) and primary cells (small airway epithelial cells and hepatocytes) were infected with adenovirus vectors transducing alpha(1)-antitrypsin with (AdCMV.p21-RSV.hAAT) or without (AdRSV.hAAT) p21. alpha(1)-Antitrypsin expression and cytotoxicity were analyzed using western blot/ELISA and LDH/ALT/AST assays, respectively. Cell cycle profiles were determined by flow cytometry. Co-expression of p21 strongly increased the alpha(1)-antitrypsin expression in all cell types and at all doses tested. No changes in ALT/AST from hepatocytes and only minor increases in the LDH release in A549 and HeLa were observed with either vector. Cell cycle profiles were also not affected adversely. Incorporation of p21 in Ad vectors together with a gene of interest improves the vector performance; such vectors will allow the application of lower doses and thereby reduce immunological side effects.

Increased bone mass, altered trabecular architecture and modified growth plate organization in the growing skeleton of SOCS2 deficient mice.

Suppressor of cytokine signalling-2 (SOCS2) negatively regulates the signal transduction of several cytokines. Socs2(-/-) mice show increased longitudinal skeletal growth associated with deregulated GH/IGF-1 signalling. The present study examined the role of SOCS2 in endochondral ossification and trabecular and cortical bone formation, and investigated whether pro-inflammatory cytokines associated with pediatric chronic inflammatory disorders mediate their effects through SOCS2. Seven-week-old Socs2(-/-) mice were heavier (27%; P < 0.001) and longer (6%; P < 0.001) than wild-type mice. Socs2(-/-) tibiae were longer (8%; P < 0.001) and broader (18%; P < 0.001) than that of wild-type mice, and the Socs2(-/-) mice had wider growth plates (24%; P < 0.001) with wider proliferative and hypertrophic zones (10% (P < 0.05) and 14% (P < 0.001) respectively). Socs2(-/-) mice showed increased total cross-sectional bone area (16%: P < 0.001), coupled to increased total tissue area (17%; P < 0.05) compared to tibia from wild-type mice. Socs2(-/-) mice showed increased percent bone volume (101%; P < 0.001), trabecular number (82%; P < 0.001) and trabecular thickness (11%; P < 0.001), with associated decreases in trabecular separation (19%; P < 0.001). TNFalpha exposure to growth plate chondrocytes for 48 h increased SOCS2 protein expression. Growth of metatarsals from 1-day-old Socs2(-/-) and Socs2(+/+) mice, as well as expression of Aggrecan, Collagen Type II and Collagen Type X, were inhibited by TNFalpha, with no effect of genotype. Our data indicate that physiological levels of SOCS2 negatively regulate bone formation and endochondral growth. Our results further suggest that pro-inflammatory cytokines mediate their inhibitory effects on longitudinal bone growth through a mechanism that is independent of SOCS2.

[Studies on local anesthetics. Part 183: micellization and thermodynamic parameters of heptacainium chloride in the solution of potassium bromide]. / Stúdium lokálnych anestetik. Cast' 183: micelizácia a termodynamické parametre heptakaíniumchloridu v prostredí roztoku KBr.

The critical micellar concentration (CMC) of the local anaesthetic agent heptacainium chloride in the solution of KBr was determined by the spectrophotometric method in the UV region of the spectrum at the temperature range of t = 20-40 degrees C and pH = 4.5-5.0. The dependence of CMC on the temperature T turned out forming the U-shape with the minimum at the temperature of t = 25 degrees C. The parabolic dependence of CMC on the temperature T was drawn by the fitting of the values using the polynomial function and the so-called power law equation. The CMC dependence on the temperature T was fitted by the second degree polynomial function. The obtained parabolic equations were applied to the "phase separation model", so the following thermodynamic parameters could be calculated: standard Gibbs free energy (deltaG), enthalpy (deltaH degrees), and entropy (deltaS degrees). The thermodynamic parameters were further used to determine the so-called entropy-enthalpy compensation of the systems under study. The compensation temperature was in the following range: (301 +/- 1-303 +/- 3)K. Then the temperature dependence of the enthalpy (deltaH degrees) and entropy (-TdeltaS degrees) contributions to the standard Gibbs free energy (deltaG degrees) for all prepared concentrations of the compound were calculated.

Reactivity and oxidative potential of fructose and glucose in enkephalin-sugar model systems.

The reactions of Leu- and Met-enkephalin (Tyr-Gly-Gly-Phe-Leu/Met) with fructose resulted in the parallel formation of Heyns compounds (N-peptidyl-D-mannosamine and -D-glucosamine) and sugar-peptide generated imidazolidinone diastereomers. Glucose showed higher level of reactivity than fructose with respect to the extent of glycated product formation. The presence of fructose in the incubation mixtures makes Met residue more susceptible to oxidation than glucose.

In situ analysis of Raidd-beta-galactosidase fusion gene expression in transgenic mouse midgestation embryos.

Apoptosis and differentiation are tightly intertwined processes occurring at organ formation and remodelling during embryonic development. RAIDD (receptor-interacting protein [RIP]-associated ICH-1/CED-3-homologous protein with a death domain), a dual-domain adaptor protein has been shown to mediate the recruitment of CASPASE-2 to tumour necrosis factor receptor-1 (TNF-R1) signalling complex through RIP kinase. However, Raidd overexpression studies suggest that apart from the established role in apoptosis, Raidd may have an additional function in cell differentiation. In this study, we could not generate Raidd null adult mice suggesting that lack of function of Raidd might be embryonic lethal. Thus, to elucidate the role of Raidd during mouse embryogenesis when the processes of organogenesis are most dynamic, we studied the Raidd expression pattern in midgestation mouse embryos. We generated Raidd+/- transgenic mice with a reporter transgene encoding the bacterial Beta-galactosidase (beta-gal) under the control of Raidd promoter. During the midgestation period (E8.5-E12.5), Raidd is expressed in developing organs derived from the ectoderm such as lens, structures of the inner ear and the fourth brain ventricle in regions where differentiation takes place implicating Raidd role in this process. In addition, Raidd expression was found in developing mesenchyme organs like heart and kidney and in the endothelial lining of the midgut at the time when profound morphological changes take place in these organs. In developing heart and kidney Raidd expression patterns overlapped with known zones of cell death suggesting Raidd may be involved in apoptosis-mediated remodelling. The observed lethality of mice targeted at both Raidd alleles and Raidd expression patterns during midgestation period strongly suggest that Raidd plays an important role in mammalian development.

Embryonic activation and developmental expression of the murine prion protein gene.

While it is well established that cellular prion protein (PrP(C)) expression is required for the development of transmissible spongiform encephalopathies (TSEs), the physiological function of PrP(C) has yet to be determined. A number of studies have examined PrP expression in different tissues and in the later stages of embryonic development. However, the relative levels of expression of PrP RNA and protein in tissues outside the central nervous system (CNS) is not well documented and the exact point of transcriptional activation of PrP during embryogenesis is unknown. We have studied PrP mRNA expression in murine embryos and both mRNA and protein expression in a variety of adult tissues. PrP RNA was detected at different levels in all tissues tested while PrP(C) protein was detectable in all adult tissues tested with the exception of kidney and liver. RNA and protein levels were also assessed at four points during postnatal brain development and levels of both were seen to increase with development. We also established that, during embryogenesis, induction of PrP RNA expression occurs between E8.5 and E9, during the period of transition from anaerobic to aerobic metabolism. Preliminary experiments investigating the effects of superoxide radicals on PrP expression in cultured neuroblastoma and astrocyte cells support the suggestion that PrP(C) forms part of a cellular antioxidant defense mechanism.

Overexpression of Raidd cDNA inhibits differentiation of mouse preadipocytes.

RAIDD (RIP-associated ICH-1 homologous protein with a death domain) is an adaptor molecule that mediates the action of cysteine proteases involved in apoptosis. To study the possibility of a novel system of cell ablation mediated by RAIDD, a preadipocyte cell line (3T3L1) was stably transfected with a plasmid containing the murine Raidd cDNA under the control of the adipocyte specific promoter aP2. Instead of the expected apoptosis, a blockage to differentiation upon hormonal induction was observed as judged by an absence of lipid accumulation, a lack of expression of adipocyte-specific genes and a fibroblastic appearance. Proliferation rate of Raidd-transfected clones remained unaffected. Overexpression of Raidd cDNA in 3T3L1 cell therefore inhibited differentiation, suggesting that Raidd plays a role in controlling differentiation of mouse preadipocytes and, perhaps, in other cell types, in addition to its established role in apoptosis.

Molecular basis of the length variation in the N-terminal part of Mycoplasma synoviae hemagglutinin.

Mycoplasma synoviae is a major avian pathogen that synthesizes hemagglutinin VlhA, an abundant immunodominant surface lipoprotein. In most M. synoviae strains, the VlhA protein cleaves into the N-terminal part, a lipoprotein MSPB, and a C-terminal part MSPA, which mediates binding to erythrocytes. VlhA is encoded by the vlhA gene of which the 5'-end is present in the genome as a single copy, which does not change its sequence during recombination of the vlhA gene with pseudogenes. In this study, sequence analyses of the 5'-end vlhA sequences of 30 M. synoviae isolates revealed a highly polymorphic region encoding the proline-rich repeats (PRR) in the N-terminal part of MSPB. Pathogenic strain K1968 had an insertion encoding sequence DNPQNPN in PRR, whereas strains F10-2AS, K2581, K3344 and five strains belonging to two related clusters of strains isolated recently from chickens in Slovenia lacked one PRR repeat of 19 amino acids. The predicted length variations correlated well with the lengths of the corresponding MSPB proteins detected in immunoblots with specific antibodies. Comparison of the 5'-end vlhA sequences of 30 M. synoviae strains showed 11 different types of vlhA sequences indicating that the analysis of this vlhA part is useful for strain differentiation. Distinct sequence motifs seem to be characteristic for vlhA genes of individual M. synoviae strains or clusters of strains and can be used as markers for tracing their spreading between poultry farms.

Lack of Socs2 expression causes the high-growth phenotype in mice.

Characterizing causal molecular defects in mouse models of overgrowth or dwarfism helps to identify the key genes and pathways that regulate the growth process. We report here the molecular basis for high growth (hg), a spontaneous mutation that causes a 30-50% increase in postnatal growth. We conclude that hg is an allele of the suppressor of cytokine signaling 2 (Socs2), a member of a family of regulators of cytokine signal transduction. We demonstrate mapping of Socs2 to the hg region, lack of Socs2 mRNA expression, a disruption of the Socs2 locus in high-growth (HG) mice, and a similarity of phenotypes of HG mice and Socs2(-/-) mice generated by gene targeting. Characteristics of the HG phenotype suggest that Socs2 deficiency affects growth prenatally and postnatally most likely through deregulating the growth hormone (GH)/insulin-like growth factor I (IGF1). These results demonstrate a critical role for Socs2 in controlling growth.

The early glycation products of the Maillard reaction: mass spectrometric characterization of novel imidazolidinones derived from an opioid pentapeptide and glucose.

Glucose-substituted imidazolidinones related to the endogenous opioid peptide leucine-enkephalin have been investigated using fast atom bombardment tandem mass spectrometry (FAB-MS/MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS). In addition to Amadori compounds, the studied imidazolidinones represent a novel type of the early glycation products formed in the Maillard reaction. To obtain insight into the fragmentation behavior of these carbohydrate-peptide adducts, we also studied synthetic precursors of the glucose-substituted imidazolidinones as well as the corresponding isopropylidene derivatives. The collision-induced dissociation (CID) spectra of [M + H](+) ions of all these imidazolidinones have been compared. Detailed analysis showed that fragmentation of each compound generates two ions at m/z 566 and m/z 598 which are characteristic and undoubtedly confirm the imidazolidinone-type structure. These two significant ions were identified as the M + 10 and M + 42 modifications of the N-terminus of the parent opioid pentapeptide effected by the carbohydrate moiety. Furthermore, the ion at m/z 178 is identified as the M + 42 modification of the immonium ion of the N-terminal amino acid (tyrosine) also effected by the carbohydrate moiety. They can be used as diagnostic ions for imidazolidinone-type compounds in studying the Maillard reaction. Thus, we have demonstrated the utility of FAB-MS/MS and ESI-MS/MS in the structural determination and identification of such novel peptide-carbohydrate adducts, useful in understanding the details of the mechanism of non-enzymatic glycation in vivo.

Quantitative trait loci affecting growth in high growth (hg) mice.

A genome-wide scan was performed in order to identify Quantitative Trait Loci (QTL) associated with growth in a population segregating high growth (hg), a partially recessive mutation that enhances growth rate and body size in the mouse. A sample of 262 hg/hg mice was selected from a C57BL/6J-hg/hg x CAST/EiJ F2 cross and typed with 79 SSLP markers distributed across the genome. Eight significant loci were identified through interval mapping. Loci on Chromosomes (Chrs) 2 and 8 affected the growth rate of F2 mice. Loci on Chr 2 and 11 affected growth rate and carcass lean mass (protein and ash). A locus on Chr 9 modified femur length and another one in Chr 17 affected both carcass lean mass and femur length, but none of these had significant effects on growth rate. Loci on Chrs 5 and 9 modified carcass fat content. Additive effects were positive for C57BL/6J alleles, except for the two loci affecting carcass fatness. Typing of selected markers in 274 +/+ F2 mice revealed significant interactions between hg and other growth QTL, which were detected as changes in gene action (additive or dominant) and in allele substitution effects. Knowledge about interactions between loci, especially when major genes are involved, will help in the identification of positional candidate genes and in the understanding of the complex genetic regulation of growth rate and body size in mammals.

Spectroscopic evidence of beta-turn in N-glycated peptidomimetics related to leucine-enkephalin.

The conformational differences caused by N-glycation of the amide bond in endogenous opioid pentapeptide leucine-enkephalin (Tyr-Gly-Gly-Phe-Leu) have been explored in solution using FTIR spectroscopy, NMR and molecular modelling. The compounds studied include protected and unprotected enkephalin analogues N-alkylated at the second (Gly2) amino acid residue with a 6-deoxy-D-galactose moiety (1-3). Comparison of the amide I component bands in the FTIR spectra, measured in trifluoroethanol (TFE), CHCl3 and DMSO, revealed significant differences in the intensity as well as shifts in component band frequencies for glycopeptides 1-3. We found that only the FTIR spectrum of the fully protected compound 1 indicated the presence of a higher population of beta-turns, while the spectra of the partially protected and unprotected glycopeptides 2 and 3 reflected the dominance of unordered or open structures, with some low population of turns. The observed NOE connectivities in CDCl3 for both isomers of the fully protected compound 1, the all-trans one and another with Tyr1-Gly2 peptide bond in cis conformation, indicate the presence of a beta-like turn conformation. Molecular dynamics simulations of the glycopeptide 1 obtained by unconstrained energy minimization of trans- and cis-1 shows that one of trans form conformations is consistent with beta-turn whereas cis isomer has revealed less-compact turn.