A novel model of double replications and random loss accounts for rearrangements in the Mitogenome of Samariscus latus (Teleostei: Pleuronectiformes)

Background

Although more than one thousand complete mitochondrial DNA (mtDNA) sequences have been determined in teleostean fishes, only a few gene rearrangements have been observed, and genome-scale rearrangements are even rarer. However, flatfishes (Pleuronectiformes) have been identified as having diverse types of mitochondrial gene rearrangements. It has been reported that tongue soles and the blue flounder mitogenomes exhibit different types of large-scale gene rearrangements.

Results

In the present study, the complete mitochondrial genome of another flatfish, Samariscus latus, was sequenced, and genome-scale rearrangements were observed. The genomic features of this flounder are different from those of any other studied vertebrates, including flatfish species too. The mitogenome of S. latus is characterized by the duplication and translocation of the control region (CR). The genes located between the two CRs are divided into two clusters in which their relative orders are maintained.

Conclusions

We propose a “Double Replications and Random Loss” model to explain the rearrangement events in S. latus mitogenome. This model consists of the following steps. First, the CR was duplicated and translocated. Subsequently, double replications of the mitogenome were successively initiated from the two CRs, leading to the duplication of the genes between the two CRs. Finally, one of each pair of duplicated genes was lost in a random event.

Electronic supplementary material

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Species-specific duplications driving the recent expansion of NBS-LRR genes in five Rosaceae species

Background

Disease resistance (R) genes from different Rosaceae species have been identified by map-based cloning for resistance breeding. However, there are few reports describing the pattern of R-gene evolution in Rosaceae species because several Rosaceae genome sequences have only recently become available.

Results

Since most disease resistance genes encode NBS-LRR proteins, we performed a systematic genome-wide survey of NBS-LRR genes between five Rosaceae species, namely Fragaria vesca (strawberry), Malus × domestica (apple), Pyrus bretschneideri (pear), Prunus persica (peach) and Prunus mume (mei) which contained 144, 748, 469, 354 and 352 NBS-LRR genes, respectively. A high proportion of multi-genes and similar Ks peaks (Ks = 0.1- 0.2) of gene families in the four woody genomes were detected. A total of 385 species-specific duplicate clades were observed in the phylogenetic tree constructed using all 2067 NBS-LRR genes. High percentages of NBS-LRR genes derived from species-specific duplication were found among the five genomes (61.81% in strawberry, 66.04% in apple, 48.61% in pear, 37.01% in peach and 40.05% in mei). Furthermore, the Ks and Ka/Ks values of TIR-NBS-LRR genes (TNLs) were significantly greater than those of non-TIR-NBS-LRR genes (non-TNLs), and most of the NBS-LRRs had Ka/Ks ratios less than 1, suggesting that they were evolving under a subfunctionalization model driven by purifying selection.

Conclusions

Our results indicate that recent duplications played an important role in the evolution of NBS-LRR genes in the four woody perennial Rosaceae species. Based on the phylogenetic tree produced, it could be inferred that species-specific duplication has mainly contributed to the expansion of NBS-LRR genes in the five Rosaceae species. In addition, the Ks and Ka/Ks ratios suggest that the rapidly evolved TNLs have different evolutionary patterns to adapt to different pathogens compared with non-TNL resistant genes.

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The role of bronchial epithelial cells in the pathogenesis of COPD in Z-alpha-1 antitrypsin deficiency

Background

Alpha-1 antitrypsin is the main inhibitor of neutrophil elastase in the lung. Although it is principally synthesized by hepatocytes, alpha-1 antitrypsin is also secreted by bronchial epithelial cells. Gene mutations can lead to alpha-1 antitrypsin deficiency, with the Z variant being the most clinically relevant due to its propensity to polymerize. The ability of bronchial epithelial cells to produce Z-variant protein and its polymers is unknown.We investigated the expression, accumulation, and secretion of Z-alpha-1 antitrypsin and its polymers in cultures of transfected cells and in cells originating from alpha-1 antitrypsin-deficient patients.

Methods

Experiments using a conformation-specific antibody were carried out on M- and Z-variant–transfected 16HBE cells and on bronchial biopsies and ex vivo bronchial epithelial cells from Z and M homozygous patients. In addition, the effect of an inflammatory stimulus on Z-variant polymer formation, elicited by Oncostatin M, was investigated. Comparisons of groups were performed using t-test or ANOVA. Non-normally distributed data were assessed by Mann–Whitney U test or the Kruskal-Wallis test, where appropriate. A P value of < 0.05 was considered to be significant.

Results

Alpha-1 antitrypsin polymers were found at a higher concentration in the culture medium of ex vivo bronchial epithelial cells from Z-variant homozygotes, compared with M-variant homozygotes (P < 0.01), and detected in the bronchial epithelial cells and submucosa of patient biopsies. Oncostatin M significantly increased the expression of alpha-1 antitrypsin mRNA and protein (P < 0.05), and the presence of Z-variant polymers in ex vivo cells (P < 0.01).

Conclusions

Polymers of Z-alpha-1 antitrypsin form in bronchial epithelial cells, suggesting that these cells may be involved in the pathogenesis of lung emphysema and in bronchial epithelial cell dysfunction.

Electronic supplementary material

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Promoter methylation analysis of WNT/β-catenin pathway regulators and its association with expression of DNMT1 enzyme in colorectal cancer

Background

Aberrant DNA methylation as the most important reason making epigenetic silencing of genes is a main mechanism of gene inactivation in patients with colorectal cancer. In this study, we decided to identify promoter methylation status of ten genes encoding WNT negative regulators, and measure the expression of DNMT1 enzyme in colorectal cancer samples.

Results

Aberrant methylation of APC gene was statistically significant associated with age over 50 (p = 0.017), DDK3 with male (p < 0.0001), SFRP4, WIF1, and WNT5a with increasing tumor stage (p = 0.004, p = 0.029, and p = 0.004), SFRP4 and WIF1 with tumor differentiation (p = 0.009 and p = 0.031) and SFRP2 and SFRP5 with histological type (p = 0.001 and p = 0.025). The increasing number of methylated genes correlated with the expression levels of the DNMT1 mRNA.

Conclusions

The rate of gene promoter methylation of WNT pathway regulators is high in colorectal cancer cells. Hyper-methylation is associated with increased expression of the DNMT1 enzyme.     

Association between risk of oral precancer and genetic variations in microRNA and related processing genes

Background

MicroRNAs have been implicated in cancer but studies on their role in precancer, such as leukoplakia, are limited. Sequence variations at eight miRNA and four miRNA processing genes were studied in 452 healthy controls and 299 leukoplakia patients to estimate risk of disease.

Results

Genotyping by TaqMan assay followed by statistical analyses showed that variant genotypes at Gemin3 and mir-34b reduced risk of disease [OR = 0.5(0.3–0.9) and OR = 0.7(0.5–0.9) respectively] in overall patients as well as in smokers [OR = 0.58(0.3–1) and OR = 0.68(0.5–0.9) respectively]. Among chewers, only mir29a significantly increased risk of disease [OR = 1.8(1–3)]. Gene-environment interactions using MDR-pt program revealed that mir29a, mir34b, mir423 and Xpo5 modulated risk of disease (p < 0.002) which may be related to change in expression of these genes as observed by Real-Time PCR assays. But association between polymorphisms and gene expressions was not found in our sample set as well as in larger datasets from open access platforms like Genevar and 1000 Genome database.

Conclusion

Variations in microRNAs and their processing genes modulated risk of precancer but further in-depth study is needed to understand mechanism of disease process.     

Impaired macrophage phagocytosis of bacteria in severe asthma

Background

Bacteria are frequently cultured from sputum samples of severe asthma patients suggesting a defect in bacterial clearance from the airway. We measured the capacity of macrophages from patients with asthma to phagocytose bacteria.

Methods

Phagocytosis of fluorescently-labelled polystyrene beads, Haemophilus influenzae or Staphylococcus aureus by broncholaveolar lavage alveolar macrophages (AM) and by monocyte-derived macrophages (MDM) from non-asthmatics, mild-moderate and severe asthmatic patients was assessed using fluorimetry.

Results

There were no differences in phagocytosis of polystyrene beads by AMs or MDMs from any of the subject groups. There was reduced phagocytosis of Haemophilus influenzae and Staphylococcus aureus in MDMs from patients with severe asthma compared to non-severe asthma (p < 0.05 and p < 0.01, respectively) and healthy subjects (p < 0.01and p < 0.001, respectively). Phagocytosis of Haemophilus influenzae and Staphylococcus aureus by AM was also reduced in severe asthma compared to normal subjects (p < 0.05). Dexamethasone and formoterol did not suppress phagocytosis of bacteria by MDMs from any of the groups.

Conclusions

Persistence of bacteria in the lower airways may result partly from a reduced phagocytic capacity of macrophages for bacteria. This may contribute to increased exacerbations, airway colonization and persistence of inflammation.     

Decrease in quality of life predicts mortality in adult patients with pulmonary arterial hypertension due to congenital heart disease

Background

Decrease in quality of life (QoL) in left-sided heart failure precedes poor survival, which can be reversed with exercise training. We investigated whether QoL is associated with mortality in pulmonary arterial hypertension due to congenital heart disease (PAH-CHD) patients.

Methods

In this observational study, PAH-CHD adults referred for PAH-specific therapy were included. QoL surveys (SF36) were recorded during 2 years of therapy. Based on shift in SF36 scores during this period, patients had either decreased or non-decreased QoL. Subsequently, the patients were followed for mortality.

Results

Thirty-nine PAH-CHD patients (mean age 42, 44 % male, 49 % Down’s syndrome) were analysed. Following PAH-specific therapy, SF36 physical component summary (PCS) decreased in 13 (35–31 points, p = 0.001) and showed no decrease in 26 patients (34–43 points, mean values, p < 0.001). Post-initiation phase, median follow-up was 4.5 years, during which 12 deaths occurred (31 %), 10 (56 %) in the decreased and 2 (10 %) in the non-decreased group (p = 0.002). Cox regression showed a decrease in SF36 PCS predicted mortality (HR 3.4, 95 % CI 1.03–11, p = 0.045).

Conclusions

In PAH-CHD patients, decrease in SF36 PCS following initiation of PAH-specific therapy is a determinant of mortality.

Electronic supplementary material

The online version of this article (doi:10.1007/s12471-015-0666-9) contains supplementary material, which is available to authorized users.     

A high density GBS map of bread wheat and its application for dissecting complex disease resistance traits

Background

Genotyping-by-sequencing (GBS) is a high-throughput genotyping approach that is starting to be used in several crop species, including bread wheat. Anchoring GBS tags on chromosomes is an important step towards utilizing them for wheat genetic improvement. Here we use genetic linkage mapping to construct a consensus map containing 28644 GBS markers.

Results

Three RIL populations, PBW343 × Kingbird, PBW343 × Kenya Swara and PBW343 × Muu, which share a common parent, were used to minimize the impact of potential structural genomic variation on consensus-map quality. The consensus map comprised 3757 unique positions, and the average marker distance was 0.88 cM, obtained by calculating the average distance between two adjacent unique positions. Significant variation of segregation distortion was observed across the three populations. The consensus map was validated by comparing positions of known rust resistance genes, and comparing them to wheat reference genome sequences recently published by the International Wheat Genome Sequencing Consortium, Rye and Ae. tauschii genomes. Three well-characterized rust resistance genes (Sr58/Lr46/Yr29, Sr2/Yr30/Lr27, and Sr57/Lr34/Yr18) and 15 published QTLs for wheat rusts were validated with high resolution. Fifty-two per cent of GBS tags on the consensus map were successfully aligned through BLAST to the right chromosomes on the wheat reference genome sequence.

Conclusion

The consensus map should provide a useful basis for analyzing genome-wide variation of complex traits. The identified genes can then be explored as genetic markers to be used in genomic applications in wheat breeding.

Electronic supplementary material

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Ozone exposure,vitamin C intake,and genetic susceptibility of asthmatic children in Mexico City: a cohort study

Background

We previously reported that asthmatic children with GSTM1 null genotype may be more susceptible to the acute effect of ozone on the small airways and might benefit from antioxidant supplementation. This study aims to assess the acute effect of ozone on lung function (FEF_25-75) in asthmatic children according to dietary intake of vitamin C and the number of putative risk alleles in three antioxidant genes: GSTM1, GSTP1 (rs1695), and NQO1 (rs1800566).

Methods

257 asthmatic children from two cohort studies conducted in Mexico City were included. Stratified linear mixed models with random intercepts and random slopes on ozone were used. Potential confounding by ethnicity was assessed. Analyses were conducted under single gene and genotype score approaches.

Results

The change in FEF_25-75 per interquartile range (60 ppb) of ozone in persistent asthmatic children with low vitamin C intake and GSTM1 null was −91.2 ml/s (p = 0.06). Persistent asthmatic children with 4 to 6 risk alleles and low vitamin C intake showed an average decrement in FEF_25-75 of 97.2 ml/s per 60 ppb of ozone (p = 0.03). In contrast in children with 1 to 3 risk alleles, acute effects of ozone on FEF_25-75 did not differ by vitamin C intake.

Conclusions

Our results provide further evidence that asthmatic children predicted to have compromised antioxidant defense by virtue of genetic susceptibility combined with deficient antioxidant intake may be at increased risk of adverse effects of ozone on pulmonary function.     

Identification of a common risk haplotype for canine idiopathic epilepsy in the ADAM23 gene

Background

Idiopathic epilepsy is a common neurological disease in human and domestic dogs but relatively few risk genes have been identified to date. The seizure characteristics, including focal and generalised seizures, are similar between the two species, with gene discovery facilitated by the reduced genetic heterogeneity of purebred dogs. We have recently identified a risk locus for idiopathic epilepsy in the Belgian Shepherd breed on a 4.4 megabase region on CFA37.

Results

We have expanded a previous study replicating the association with a combined analysis of 157 cases and 179 controls in three additional breeds: Schipperke, Finnish Spitz and Beagle (p_c = 2.9e–07, p_GWAS = 1.74E-02). A targeted resequencing of the 4.4 megabase region in twelve Belgian Shepherd cases and twelve controls with opposite haplotypes identified 37 case-specific variants within the ADAM23 gene. Twenty-seven variants were validated in 285 cases and 355 controls from four breeds, resulting in a strong replication of the ADAM23 locus (p_raw = 2.76e–15) and the identification of a common 28 kb-risk haplotype in all four breeds. Risk haplotype was present in frequencies of 0.49–0.7 in the breeds, suggesting that ADAM23 is a low penetrance risk gene for canine epilepsy.

Conclusions

These results implicate ADAM23 in common canine idiopathic epilepsy, although the causative variant remains yet to be identified. ADAM23 plays a role in synaptic transmission and interacts with known epilepsy genes, LGI1 and LGI2, and should be considered as a candidate gene for human epilepsies.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1651-9) contains supplementary material, which is available to authorized users.     

A novel approach for multi-domain and multi-gene family identification provides insights into evolutionary dynamics of disease resistance genes in core eudicot plants

Background

Recent advances in DNA sequencing techniques resulted in more than forty sequenced plant genomes representing a diverse set of taxa of agricultural, energy, medicinal and ecological importance. However, gene family curation is often only inferred from DNA sequence homology and lacks insights into evolutionary processes contributing to gene family dynamics. In a comparative genomics framework, we integrated multiple lines of evidence provided by gene synteny, sequence homology and protein-based Hidden Markov Modelling to extract homologous super-clusters composed of multi-domain resistance (R)-proteins of the NB-LRR type (for NUCLEOTIDE BINDING/LEUCINE-RICH REPEATS), that are involved in plant innate immunity.

Results

To assess the diversity of R-proteins within and between species, we screened twelve eudicot plant genomes including six major crops and found a total of 2,363 NB-LRR genes. Our curated R-proteins set shows a 50% average for tandem duplicates and a 22% fraction of gene copies retained from ancient polyploidy events (ohnologs). We provide evidence for strong positive selection and show significant differences in molecular evolution rates (Ka/Ks-ratio) among tandem- (mean = 1.59), ohnolog (mean = 1.36) and singleton (mean = 1.22) R-gene duplicates. To foster the process of gene-edited plant breeding, we report species-specific presence/absence of all 140 NB-LRR genes present in the model plant Arabidopsis and describe four distinct clusters of NB-LRR “gatekeeper” loci sharing syntenic orthologs across all analyzed genomes.

Conclusion

By curating a near-complete set of multi-domain R-protein clusters in an eudicot-wide scale, our analysis offers significant insight into evolutionary dynamics underlying diversification of the plant innate immune system. Furthermore, our methods provide a blueprint for future efforts to identify and more rapidly clone functional NB-LRR genes from any plant species.

Electronic supplementary material

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Disruption of Mycobacterium avium subsp. paratuberculosis-specific genes impairs in vivo fitness

Background

Mycobacterium avium subsp. paratuberculosis (MAP) is an obligate intracellular pathogen that infects many ruminant species. The acquisition of foreign genes via horizontal gene transfer has been postulated to contribute to its pathogenesis, as these genetic elements are absent from its putative ancestor, M. avium subsp. hominissuis (MAH), an environmental organism with lesser pathogenicity. In this study, high-throughput sequencing of MAP transposon libraries were analyzed to qualitatively and quantitatively determine the contribution of individual genes to bacterial survival during infection.

Results

Out of 52384 TA dinucleotides present in the MAP K-10 genome, 12607 had a MycoMarT7 transposon in the input pool, interrupting 2443 of the 4350 genes in the MAP genome (56%). Of 96 genes situated in MAP-specific genomic islands, 82 were disrupted in the input pool, indicating that MAP-specific genomic regions are dispensable for in vitro growth (odds ratio = 0.21). Following 5 independent in vivo infections with this pool of mutants, the correlation between output pools was high for 4 of 5 (R = 0.49 to 0.61) enabling us to define genes whose disruption reproducibly reduced bacterial fitness in vivo. At three different thresholds for reduced fitness in vivo, MAP-specific genes were over-represented in the list of predicted essential genes. We also identified additional genes that were severely depleted after infection, and several of them have orthologues that are essential genes in M. tuberculosis.

Conclusions

This work indicates that the genetic elements required for the in vivo survival of MAP represent a combination of conserved mycobacterial virulence genes and MAP-specific genes acquired via horizontal gene transfer. In addition, the in vitro and in vivo essential genes identified in this study may be further characterized to offer a better understanding of MAP pathogenesis, and potentially contribute to the discovery of novel therapeutic and vaccine targets.

Electronic supplementary material

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Global gene expression in endometrium of high and low fertility heifers during the mid-luteal phase of the estrous cycle

Background

In both beef and dairy cattle, the majority of early embryo loss occurs within the first 14 days following insemination. During this time-period, embryos are completely dependent on their maternal uterine environment for development, growth and ultimately survival, therefore an optimum uterine environment is critical to their survival. The objective of this study was to investigate whether differences in endometrial gene expression during the mid-luteal phase of the estrous cycle exist between crossbred beef heifers ranked as either high (HF) or low fertility (LF) (following four rounds of artificial insemination (AI)) using the Affymetrix® 23 K Bovine Gene Chip.

Results

Conception rates for each of the four rounds of AI were within a normal range: 70–73.3%. Microarray analysis of endometrial tissue collected on day 7 of the estrous cycle detected 419 differentially expressed genes (DEG) between HF (n = 6) and LF (n = 6) animals. The main gene pathways affected were, cellular growth and proliferation, angiogenesis, lipid metabolism, cellular and tissue morphology and development, inflammation and metabolic exchange. DEG included, FST, SLC45A2, MMP19, FADS1 and GALNT6.

Conclusions

This study highlights, some of the molecular mechanisms potentially controlling uterine endometrial function during the mid-luteal phase of the estrous cycle, which may contribute to uterine endometrial mediated impaired fertility in cattle. Differentially expressed genes are potential candidate genes for the identification of genetic variation influencing cow fertility, which may be incorporated into future breeding programmes.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-234) contains supplementary material, which is available to authorized users.     

Association of 25-Hydroxyvitamin D status and genetic variation in the vitamin D metabolic pathway with FEV1 in the Framingham Heart Study

Background

Vitamin D is associated with lung function in cross-sectional studies, and vitamin D inadequacy is hypothesized to play a role in the pathogenesis of chronic obstructive pulmonary disease. Further data are needed to clarify the relation between vitamin D status, genetic variation in vitamin D metabolic genes, and cross-sectional and longitudinal changes in lung function in healthy adults.

Methods

We estimated the association between serum 25-hydroxyvitamin D [25(OH)D] and cross-sectional forced expiratory volume in the first second (FEV₁) in Framingham Heart Study (FHS) Offspring and Third Generation participants and the association between serum 25(OH)D and longitudinal change in FEV₁ in Third Generation participants using linear mixed-effects models. Using a gene-based approach, we investigated the association between 241 SNPs in 6 select vitamin D metabolic genes in relation to longitudinal change in FEV₁ in Offspring participants and pursued replication of these findings in a meta-analyzed set of 4 independent cohorts.

Results

We found a positive cross-sectional association between 25(OH)D and FEV₁ in FHS Offspring and Third Generation participants (P = 0.004). There was little or no association between 25(OH)D and longitudinal change in FEV₁ in Third Generation participants (P = 0.97). In Offspring participants, the CYP2R1 gene, hypothesized to influence usual serum 25(OH)D status, was associated with longitudinal change in FEV₁ (gene-based P < 0.05). The most significantly associated SNP from CYP2R1 had a consistent direction of association with FEV₁ in the meta-analyzed set of replication cohorts, but the association did not reach statistical significance thresholds (P = 0.09).

Conclusions

Serum 25(OH)D status was associated with cross-sectional FEV₁, but not longitudinal change in FEV₁. The inconsistent associations may be driven by differences in the groups studied. CYP2R1 demonstrated a gene-based association with longitudinal change in FEV₁ and is a promising candidate gene for further studies.

Electronic supplementary material

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Effectiveness of moxibustion treatment as adjunctive therapy in osteoarthritis of the knee: a randomized,double-blinded,placebo-controlled clinical trial

Introduction

Our objective was to compare the effectiveness and safety of traditional Chinese moxibustion to that of sham moxibustion in patients with chronic knee osteoarthritis (KOA) pain.

Methods

We conducted a randomized placebo-controlled trial involving 110 patients with KOA who met the inclusion criteria. These patients randomly received either active moxibustion (n = 55) or sham moxibustion control (n = 55) at acupoints Dubi (ST 35), extra-point Neixiyan (EX-LE 4), and an Ashi (tender) point three times a week for 6 weeks. Effects were evaluated with Western Ontario and McMaster Universities’ Osteoarthritis Index (WOMAC VA 3.1) criteria at the end of the course of treatment and 3, 12, and 24 weeks after the initial treatment.

Results

The WOMAC pain scores showed greater improvement in the active treatment group than in control at weeks 3 (P = 0.012), 6 (P <0.001), 12 (P = 0.002), and 24 (P = 0.002) as did WOMAC physical function scores of the active treatment group at week 3 (P = 0.002), 6 (P = 0.015), and 12 (P <0.001) but not 24 (P = 0.058). Patients and practitioners were blinded successfully, and no significant adverse effects were found during the trial.

Conclusions

A 6-week course of moxibustion seems to relieve pain effectively and improve function in patients with KOA for up to 18 weeks after the end of treatment. Moxibustion treatment appears to be safe, and the usefulness of the novel moxa device was validated.

Trial registration

Current controlled trial: ISRCTN68475405. Registered 4 April 2014.