Microsomal and cytosolic cytochrome P450 mediated benzo(a)pyrene hydroxylation in Pleurotus pulmonarius

Summary Microsomal and soluble fractions of Pleurotus pulmonarius exhibited a reduced carbon monoxide difference spectrum with P450 maxima at 448nm and 450–452nm respectively. Substrate induced Type I spectra were observed on addition of benzo(a)pyrene to both fractions. Benzo(a)pyrene hydroxylation was measured using the aryl hydrocarbon hydroxylase assay and was observed to be P450 dependent as indicated by carbon monoxide inhibition together with the substrate binding characteristics. The activity of the fractions were observed to give K_m of 200mM and 660mM and V_max of 1.25 nmol/min/nmol P450 and 0.57 nmol/min/nmol P450 for the microsomal and cytosolic fractions respectively.     

Getting Your Sea Legs

Sea travel mandates changes in the control of the body. The process by which we adapt bodily control to life at sea is known as getting one''s sea legs. We conducted the first experimental study of bodily control as maritime novices adapted to motion of a ship at sea. We evaluated postural activity (stance width, stance angle, and the kinematics of body sway) before and during a sea voyage. In addition, we evaluated the role of the visible horizon in the control of body sway. Finally, we related data on postural activity to two subjective experiences that are associated with sea travel; seasickness, and mal de debarquement. Our results revealed rapid changes in postural activity among novices at sea. Before the beginning of the voyage, the temporal dynamics of body sway differed among participants as a function of their (subsequent) severity of seasickness. Body sway measured at sea differed among participants as a function of their (subsequent) experience of mal de debarquement. We discuss implications of these results for general theories of the perception and control of bodily orientation, for the etiology of motion sickness, and for general phenomena of perceptual-motor adaptation and learning.     

Is local selection so widespread in river organisms? Fractal geometry of river networks leads to high bias in outlier detection

Identifying local adaptation is crucial in conservation biology to define ecotypes and establish management guidelines. Local adaptation is often inferred from the detection of loci showing a high differentiation between populations, the so‐called F_ST outliers. Methods of detection of loci under selection are reputed to be robust in most spatial population models. However, using simulations we showed that F_ST outlier tests provided a high rate of false‐positives (up to 60%) in fractal environments such as river networks. Surprisingly, the number of sampled demes was correlated with parameters of population genetic structure, such as the variance of F_STs, and hence strongly influenced the rate of outliers. This unappreciated property of river networks therefore needs to be accounted for in genetic studies on adaptation and conservation of river organisms.     

Regulation of the RYR1 and RYR2 Ca2+ release channel isoforms by Ca2+-insensitive mutants of calmodulin

Calmodulin (CaM) may function as a regulatory subunit of ryanodine receptor (RYR) channels, modulating both channel activation and inhibition by Ca2+; however, mechanisms underlying differences in CaM regulation of the RYR isoforms expressed in skeletal muscle (RYR1) and cardiac muscle (RYR2) are poorly understood. Here we use a series of CaM mutants deficient in Ca2+ binding to compare determinants of CaM regulation of the RYR1 and RYR2 isoforms. In submicromolar Ca2+, activation of the RYR1 isoform by each of the single-point CaM mutants was similar to that by wild-type apoCaM, whereas in micromolar Ca2+, RYR1 inhibition by Ca2+CaM was abolished by mutations targeting CaM's C-terminal Ca2+ sites. In contrast to the RYR1, no activation of the cardiac RYR2 isoform by wild-type CaM was observed, but rather CaM inhibited the RYR2 at all Ca2+ concentrations (100 nM to 1 mM). Consequently, whereas the apparent Ca2+ sensitivity of the RYR1 isoform was enhanced in the presence of CaM, the RYR2 displayed the opposite response (RYR2 Ca2+ EC50 increased 7-10-fold in the presence of 5 microM wild-type CaM). CaM inhibition of the RYR2 was nonetheless abolished by each of four mutations targeting individual CaM Ca2+ sites. Furthermore, a mutant CaM deficient in Ca2+ binding at all four Ca2+ sites significantly activated the RYR2 and acted as a competitive inhibitor of RYR2 regulation by wild-type Ca2+CaM. We conclude that Ca2+ binding to CaM determines the effect of CaM on both RYR1 and RYR2 channels and that isoform differences in CaM regulation reflect the differential tuning of Ca2+ binding sites on CaM when bound to the different RYRs. These results thus suggest a novel mechanism by which CaM may contribute to functional diversity among the RYR isoforms.     

Cleavage of preflagellins by an aspartic acid signal peptidase is essential for flagellation in the archaeon Methanococcus voltae

The differences between archaeal and bacterial flagella are becoming more apparent as research on the archaeal structure progresses. One crucial difference is the presence of a leader peptide on archaeal preflagellins, which is removed from the flagellin prior to its incorporation into the flagellar filament. The enzyme responsible for the removal of the flagellin leader peptide was identified as FlaK. FlaK of Methanococcus voltae retains its preflagellin peptidase activity when expressed in Escherichia coli and used in an in vitro assay. Homologous recombination of an integration vector into the chromosomal copy of flaK resulted in a non-motile, non-flagellated phenotype. The flagellins of the mutant had larger molecular weights than their wild-type counterparts, as expected if they retained their 11- to 12-amino-acid leader peptide. Membranes of the flaK mutant were unable to process preflagellin in the in vitro assay. Site-directed mutagenesis demonstrated that two aspartic acid residues conserved with ones in type IV prepilin peptidases were necessary for proper recognition or processing of the preflagellin. As bacterial flagellins lack a leader peptide and a peptidase is not required for export and assembly, the requirement for FlaK further emphasizes the similarity archaeal flagella have with type IV pili, rather than with bacterial flagella.     

Out of Africa and back again: nested cladistic analysis of human Y chromosome variation

We surveyed nine diallelic polymorphic sites on the Y chromosomes of 1,544 individuals from Africa, Asia, Europe, Oceania, and the New World. Phylogenetic analyses of these nine sites resulted in a tree for 10 distinct Y haplotypes with a coalescence time of approximately 150,000 years. The 10 haplotypes were unevenly distributed among human populations: 5 were restricted to a particular continent, 2 were shared between Africa and Europe, 1 was present only in the Old World, and 2 were found in all geographic regions surveyed. The ancestral haplotype was limited to African populations. Random permutation procedures revealed statistically significant patterns of geographical structuring of this paternal genetic variation. The results of a nested cladistic analysis indicated that these geographical associations arose through a combination of processes, including restricted, recurrent gene flow (isolation by distance) and range expansions. We inferred that one of the oldest events in the nested cladistic analysis was a range expansion out of Africa which resulted in the complete replacement of Y chromosomes throughout the Old World, a finding consistent with many versions of the Out of Africa Replacement Model. A second and more recent range expansion brought Asian Y chromosomes back to Africa without replacing the indigenous African male gene pool. Thus, the previously observed high levels of Y chromosomal genetic diversity in Africa may be due in part to bidirectional population movements. Finally, a comparison of our results with those from nested cladistic analyses of human mtDNA and beta-globin data revealed different patterns of inferences for males and females concerning the relative roles of population history (range expansions) and population structure (recurrent gene flow), thereby adding a new sex-specific component to models of human evolution.      

Exploratory Movement Generates Higher-Order Information That Is Sufficient for Accurate Perception of Scaled Egocentric Distance

Body movement influences the structure of multiple forms of ambient energy, including optics and gravito-inertial force. Some researchers have argued that egocentric distance is derived from inferential integration of visual and non-visual stimulation. We suggest that accurate information about egocentric distance exists in perceptual stimulation as higher-order patterns that extend across optics and inertia. We formalize a pattern that specifies the egocentric distance of a stationary object across higher-order relations between optics and inertia. This higher-order parameter is created by self-generated movement of the perceiver in inertial space relative to the illuminated environment. For this reason, we placed minimal restrictions on the exploratory movements of our participants. We asked whether humans can detect and use the information available in this higher-order pattern. Participants judged whether a virtual object was within reach. We manipulated relations between body movement and the ambient structure of optics and inertia. Judgments were precise and accurate when the higher-order optical-inertial parameter was available. When only optic flow was available, judgments were poor. Our results reveal that participants perceived egocentric distance from the higher-order, optical-inertial consequences of their own exploratory activity. Analysis of participants’ movement trajectories revealed that self-selected movements were complex, and tended to optimize availability of the optical-inertial pattern that specifies egocentric distance. We argue that accurate information about egocentric distance exists in higher-order patterns of ambient energy, that self-generated movement can generate these higher-order patterns, and that these patterns can be detected and used to support perception of egocentric distance that is precise and accurate.     

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

The online version of this article (doi:10.1186/s12931-015-0238-y) contains supplementary material, which is available to authorized users.     

HLA mismatches and hematopoietic cell transplantation: Structural simulations assess the impact of changes in peptide binding specificity on transplant outcome

The success of hematopoietic cell transplantation from an unrelated donor depends in part on the degree of Human Histocompatibility Leukocyte Antigen (HLA) matching between donor and patient. We present a structure-based analysis of HLA mismatching, focusing on individual amino acid mismatches and their effect on peptide binding specificity. Using molecular modeling simulations of HLA-peptide interactions, we find evidence that amino acid mismatches predicted to perturb peptide binding specificity are associated with higher risk of mortality in a large and diverse dataset of patient-donor pairs assembled by the International Histocompatibility Working Group in Hematopoietic Cell Transplantation consortium. This analysis may represent a first step toward sequence-based prediction of relative risk for HLA allele mismatches.