A frameshift mutation in the cubilin gene (CUBN) in Beagles with Imerslund–Gräsbeck syndrome (selective cobalamin malabsorption)

Mammals are unable to synthesize cobalamin or vitamin B₁₂ and rely on the uptake of dietary cobalamin. The cubam receptor expressed on the intestinal endothelium is required for the uptake of cobalamin from the gut. Cubam is composed of two protein subunits, amnionless and cubilin, which are encoded by the AMN and CUBN genes respectively. Loss‐of‐function mutations in either the AMN or the CUBN gene lead to hereditary selective cobalamin malabsorption or Imerslund–Gräsbeck syndrome (IGS). We investigated Beagles with IGS and resequenced the whole genome of one affected Beagle at 15× coverage. The analysis of the AMN and CUBN candidate genes revealed a homozygous deletion of a single cytosine in exon 8 of the CUBN gene (c.786delC). This deletion leads to a frameshift and early premature stop codon (p.Asp262Glufs*47) and is, thus, predicted to represent a complete loss‐of‐function allele. We tested three IGS‐affected and 89 control Beagles and found perfect association between the IGS phenotype and the CUBN:c.786delC variant. Given the known role of cubilin in cobalamin transport, which has been firmly established in humans and dogs, our data strongly suggest that the CUBN:c.786delC variant is causing IGS in the investigated Beagles.     

Resurvey studies of terricolous bryophytes and lichens indicate a widespread nutrient enrichment in German forests

Questions

Vegetation resurveys, both single studies and meta-analyses, have predominantly been based on vascular plant data while bryophytes and lichens have largely been neglected. Our study aims to fill this gap and addresses the following research questions: has the overall species richness of terricolous bryophytes and lichens in forests changed over time? Which are the winners and losers among single species and ecological species groups? Do the results give a signal of the impact of nutrient enrichment, of changes in the light regime and of climate change?

Location

Deciduous and coniferous forests in Germany.

Methods

We compiled 35 single resurvey data sets, including 1096 plots in total (each sampled twice). The time interval between initial surveys and resurveys ranged from 10 to 65 years. The differences between old and new plots were analysed with respect to the frequency of single species, total species richness, and the absolute and relative numbers of taxa in the species groups. Trend scores of species were related to ecological indicator values to identify the main environmental drivers behind the observed changes.

Results

Total species richness did not systematically change, while pleurocarpous mosses had increased at the expense of acrocarpous mosses and, in coniferous forests, of lichens. Weak changes were generally observed in deciduous forests on base-rich soils. In coniferous forests and in deciduous forests on acid soils, species with high nitrogen demand and high shade tolerance had increased, whereas those being typical for more infertile and open forest sites had decreased. There were trends towards a larger share of taxa with a more oceanic distribution.

Conclusions

The changes in the vegetation of terricolous bryophytes and lichens in the studied forests indicate nutrient enrichment and increasingly shady conditions in forests on acid soils, likely caused by nitrogen deposition and shrub layer closure.     

Selective decrease in cell surface expression and mRNA level of the 55-kDa tumor necrosis factor receptor during differentiation of HL-60 cells into macrophage-like but not granulocyte-like cells.

Expression of the two known receptors for TNF was studied in the promyelocytic leukemia cell line HL-60 before and after differentiation of the cells along the granulocyte lineage (induced by incubation with retinoic acid), or along the macrophage lineage (induced by incubation with the phorbol diester, PMA). The extent of inhibition of TNF binding by receptor-specific antisera, as well as the size of the complexes formed after cross-linking TNF to its receptors on intact cells, indicated that both receptor species were expressed on the surface of the undifferentiated HL60 cells. Differentiation into granulocyte-like cells resulted in some increase in TNF binding. The increase was apparently due to enhanced expression of the 75-kDa TNF-R, whereas the amounts of the 55-kDa TNF-R did not change significantly. In contrast, in HL-60 cells induced to differentiate into macrophage-like cells, expression of the 55-kDa TNF-R species was completely abolished. The pattern of TNF-R expression in the differentiated HL-60 cells was similar to that observed in leukocytes isolated from peripheral blood: on granulocytes, there were about equal amounts of both receptor species, whereas on monocytes the 75-kDa receptor was predominant. The loss of 55-kDa receptors during differentiation of HL-60 cells into macrophage-like cells was accompanied by a pronounced decrease in the level of the mRNA for that receptor, suggesting that at least part of the change in TNF-R expression is due to mechanisms that control the amounts of receptor mRNA. Although little is yet known regarding the functional differences between the two receptor species, marked changes in the pattern of their expression, as observed during HL-60 cell differentiation, are likely to alter the kind of response of the cells to TNF and may therefore play an important role in the coordination of TNF effects in the organism.     

Fish populations in the Paraná River: Lentic environments of Diamante and San Pedro Areas (Argentine Republic)

Data on the fish populations of lentic water bodies of the Lower Paraná River in the Diamante and San Pedro areas (33°40–31°55 S, 59°40'–60°40 W), sampled during 1971 are given. Limnological characteristics such as surface area and depth of ponds, water turbidity, and plant cover are included. Differences in fish population size, structure, specific diversity, dominance, and biomass of particular species are established between two areas in a dry season.     

Cdc42 is not essential for filopodium formation, directed migration, cell polarization, and mitosis in fibroblastoid cells

Cdc42 is a small GTPase involved in the regulation of the cytoskeleton and cell polarity. To test whether Cdc42 has an essential role in the formation of filopodia or directed cell migration, we generated Cdc42-deficient fibroblastoid cells by conditional gene inactivation. We report here that loss of Cdc42 did not affect filopodium or lamellipodium formation and had no significant influence on the speed of directed migration nor on mitosis. Cdc42-deficient cells displayed a more elongated cell shape and had a reduced area. Furthermore, directionality during migration and reorientation of the Golgi apparatus into the direction of migration was decreased. However, expression of dominant negative Cdc42 in Cdc42-null cells resulted in strongly reduced directed migration, severely reduced single cell directionality, and complete loss of Golgi polarization and of directionality of protrusion formation toward the wound, as well as membrane blebbing. Thus, our data show that besides Cdc42 additional GTPases of the Rho-family, which share GEFs with Cdc42, are involved in the establishment and maintenance of cell polarity during directed migration.     

Wound-healing defect of CD18(-/-) mice due to a decrease in TGF-beta1 and myofibroblast differentiation

We studied the mechanisms underlying the severely impaired wound healing associated with human leukocyte-adhesion deficiency syndrome-1 (LAD1) using a murine disease model. In CD18(-/-) mice, healing of full-thickness wounds was severely delayed during granulation-tissue contraction, a phase where myofibroblasts play a major role. Interestingly, expression levels of myofibroblast markers alpha-smooth muscle actin and ED-A fibronectin were substantially reduced in wounds of CD18(-/-) mice, suggesting an impaired myofibroblast differentiation. TGF-beta signalling was clearly involved since TGF-beta1 and TGF-beta receptor type-II protein levels were decreased, while TGF-beta(1) injections into wound margins fully re-established wound closure. Since, in CD18(-/-) mice, defective migration leads to a severe reduction of neutrophils in wounds, infiltrating macrophages might not phagocytose apoptotic CD18(-/-) neutrophils. Macrophages would thus be lacking their main stimulus to secrete TGF-beta1. Indeed, in neutrophil-macrophage cocultures, lack of CD18 on either cell type leads to dramatically reduced TGF-beta1 release by macrophages due to defective adhesion to, and subsequent impaired phagocytic clearance of, neutrophils. Our data demonstrates that the paracrine secretion of growth factors is essential for cellular differentiation in wound healing.     

Decorin, a novel player in the insulin-like growth factor system

Decorin is a multifunctional proteoglycan that is expressed by sprouting endothelial cells. Its expression supports capillary formation and cell survival. Previously, it was shown that some effects of decorin are mediated by protein kinase B and the cyclin-dependent kinase inhibitor, p21. However, the cell surface receptor responsible for these effects was unknown. We demonstrate that decorin binds to the insulin-like growth factor-I (IGF-I) receptor on endothelial cells with an affinity in the nanomolar range (K(D) = 18 nm), which is comparable with IGF-I (K(D) = 1.2 nm). Furthermore, decorin can bind IGF-I itself, but with a lower affinity (K(D) = 190 nm) than classical IGF-I-binding proteins. Decorin addition causes IGF-I receptor phosphorylation and activation, which is followed by receptor down-regulation. These effects are caused by the core protein of decorin, and the binding region could be mapped to the N terminus of the molecule. The physiological relevance of the decorin/IGF-I receptor interaction was corroborated in two animal models (e.g. inflammatory angiogenesis in the cornea and unilateral ureteral obstruction). In both models the IGF-I receptor was up-regulated in decorin-deficient mice compared with controls and the up-regulation could not compensate the decorin deficiency in the disease models. These data indicate that decorin is an important player in the IGF system and its loss cannot fully be compensated in different types of diseases.     

Imputation of sequence level genotypes in the Franches-Montagnes horse breed

Background

A cost-effective strategy to increase the density of available markers within a population is to sequence a small proportion of the population and impute whole-genome sequence data for the remaining population. Increased densities of typed markers are advantageous for genome-wide association studies (GWAS) and genomic predictions.

Methods

We obtained genotypes for 54 602 SNPs (single nucleotide polymorphisms) in 1077 Franches-Montagnes (FM) horses and Illumina paired-end whole-genome sequencing data for 30 FM horses and 14 Warmblood horses. After variant calling, the sequence-derived SNP genotypes (~13 million SNPs) were used for genotype imputation with the software programs Beagle, Impute2 and FImpute.

Results

The mean imputation accuracy of FM horses using Impute2 was 92.0%. Imputation accuracy using Beagle and FImpute was 74.3% and 77.2%, respectively. In addition, for Impute2 we determined the imputation accuracy of all individual horses in the validation population, which ranged from 85.7% to 99.8%. The subsequent inclusion of Warmblood sequence data further increased the correlation between true and imputed genotypes for most horses, especially for horses with a high level of admixture. The final imputation accuracy of the horses ranged from 91.2% to 99.5%.

Conclusions

Using Impute2, the imputation accuracy was higher than 91% for all horses in the validation population, which indicates that direct imputation of 50k SNP-chip data to sequence level genotypes is feasible in the FM population. The individual imputation accuracy depended mainly on the applied software and the level of admixture.

Electronic supplementary material

The online version of this article (doi:10.1186/s12711-014-0063-7) contains supplementary material, which is available to authorized users.