Directed evolution of a magnetic resonance imaging contrast agent for noninvasive imaging of dopamine

The development of molecular probes that allow in vivo imaging of neural signaling processes with high temporal and spatial resolution remains challenging. Here we applied directed evolution techniques to create magnetic resonance imaging (MRI) contrast agents sensitive to the neurotransmitter dopamine. The sensors were derived from the heme domain of the bacterial cytochrome P450-BM3 (BM3h). Ligand binding to a site near BM3h's paramagnetic heme iron led to a drop in MRI signal enhancement and a shift in optical absorbance. Using an absorbance-based screen, we evolved the specificity of BM3h away from its natural ligand and toward dopamine, producing sensors with dissociation constants for dopamine of 3.3–8.9 μM. These molecules were used to image depolarization-triggered neurotransmitter release from PC12 cells and in the brains of live animals. Our results demonstrate the feasibility of molecular-level functional MRI using neural activity–dependent sensors, and our protein engineering approach can be generalized to create probes for other targets.Charles A. Dana Foundation. Brain and Immuno-ImagingRaymond and Beverley Sackler FoundationNational Institutes of Health (U.S.) (grant R01-DA28299)National Institutes of Health (U.S.) (grant DP2-OD2441)National Institutes of Health (U.S.) (grant R01-GM068664)Jacobs Institute for Molecular Engineering for Medicine. Jacobs Institute for Molecular Engineering for MedicineNational Institutes of Health (U.S.) (grant R01-DE013023

Rare germline variants in DNA repair genes and the angiogenesis pathway predispose prostate cancer patients to develop metastatic disease

Background Prostate cancer (PrCa) demonstrates a heterogeneous clinical presentation ranging from largely indolent to lethal. We sought to identify a signature of rare inherited variants that distinguishes between these two extreme phenotypes. Methods We sequenced germline whole exomes from 139 aggressive (metastatic, age of diagnosis < 60) and 141 non-aggressive (low clinical grade, age of diagnosis ≥60) PrCa cases. We conducted rare variant association analyses at gene and gene set levels using SKAT and Bayesian risk index techniques. GO term enrichment analysis was performed for genes with the highest differential burden of rare disruptive variants. Results Protein truncating variants (PTVs) in specific DNA repair genes were significantly overrepresented among patients with the aggressive phenotype, with BRCA2, ATM and NBN the most frequently mutated genes. Differential burden of rare variants was identified between metastatic and non-aggressive cases for several genes implicated in angiogenesis, conferring both deleterious and protective effects. Conclusions Inherited PTVs in several DNA repair genes distinguish aggressive from non-aggressive PrCa cases. Furthermore, inherited variants in genes with roles in angiogenesis may be potential predictors for risk of metastases. If validated in a larger dataset, these findings have potential for future clinical application

Single nucleotide polymorphism genotyping in polyploid wheat with the Illumina GoldenGate assay

Single nucleotide polymorphisms (SNPs) are indispensable in such applications as association mapping and construction of high-density genetic maps. These applications usually require genotyping of thousands of SNPs in a large number of individuals. Although a number of SNP genotyping assays are available, most of them are designed for SNP genotyping in diploid individuals. Here, we demonstrate that the Illumina GoldenGate assay could be used for SNP genotyping of homozygous tetraploid and hexaploid wheat lines. Genotyping reactions could be carried out directly on genomic DNA without the necessity of preliminary PCR amplification. A total of 53 tetraploid and 38 hexaploid homozygous wheat lines were genotyped at 96 SNP loci. The genotyping error rate estimated after removal of low-quality data was 0 and 1% for tetraploid and hexaploid wheat, respectively. Developed SNP genotyping assays were shown to be useful for genotyping wheat cultivars. This study demonstrated that the GoldenGate assay is a very efficient tool for high-throughput genotyping of polyploid wheat, opening new possibilities for the analysis of genetic variation in wheat and dissection of genetic basis of complex traits using association mapping approach

Association of IGF1 and KDM5A polymorphisms with performance, fatness and carcass traits in chickens

Two functional and positional candidate genes were selected in a region of chicken chromosome 1 (GGA1), based on their biological roles, and also where several quantitative trait loci (QTL) have been mapped and associated with performance, fatness and carcass traits in chickens. The insulin-like growth factor 1 (IGF1) gene has been associated with several physiological functions related to growth. The lysine (K)-specific demethylase 5A (KDM5A) gene participates in the epigenetic regulation of genes involved with the cell cycle. Our objective was to find associations of selected single-nucleotide polymorphisms (SNPs) in these genes with performance, fatness and carcass traits in 165 F chickens from a resource population. In the IGF1 gene, 17 SNPs were detected, and in the KDM5A gene, nine SNPs were detected. IGF1 SNP c. 47673G > A was associated with body weight and haematocrit percentage, and also with feed intake and percentages of abdominal fat and gizzard genotype × sex interactions. KDM5A SNP c. 34208C > T genotype × sex interaction affected body weight, feed intake, percentages of abdominal fat (p = 0. 0001), carcass, gizzard and haematocrit. A strong association of the diplotype × sex interaction (p < 0. 0001) with abdominal fat was observed, and also associations with body weight, feed intake, percentages of carcass, drums and thighs, gizzard and haematocrit. Our findings suggest that the KDM5A gene might play an important role in the abdominal fat deposition in chickens. The IGF1 and KDM5A genes are strong candidates to explain the QTL mapped in this region of GGA1

Evolution of Disease Response Genes in Loblolly Pine: Insights from Candidate Genes

BACKGROUND: Host-pathogen interactions that may lead to a competitive co-evolution of virulence and resistance mechanisms present an attractive system to study molecular evolution because strong, recent (or even current) selective pressure is expected at many genomic loci. However, it is unclear whether these selective forces would act to preserve existing diversity, promote novel diversity, or reduce linked neutral diversity during rapid fixation of advantageous alleles. In plants, the lack of adaptive immunity places a larger burden on genetic diversity to ensure survival of plant populations. This burden is even greater if the generation time of the plant is much longer than the generation time of the pathogen. METHODOLOGY/PRINCIPAL FINDINGS: Here, we present nucleotide polymorphism and substitution data for 41 candidate genes from the long-lived forest tree loblolly pine, selected primarily for their prospective influences on host-pathogen interactions. This dataset is analyzed together with 15 drought-tolerance and 13 wood-quality genes from previous studies. A wide range of neutrality tests were performed and tested against expectations from realistic demographic models. CONCLUSIONS/SIGNIFICANCE: Collectively, our analyses found that axr (auxin response factor), caf1 (chromatin assembly factor) and gatabp1 (gata binding protein 1) candidate genes carry patterns consistent with directional selection and erd3 (early response to drought 3) displays patterns suggestive of a selective sweep, both of which are consistent with the arm-race model of disease response evolution. Furthermore, we have identified patterns consistent with diversifying selection at erf1-like (ethylene responsive factor 1), ccoaoemt (caffeoyl-CoA-O-methyltransferase), cyp450-like (cytochrome p450-like) and pr4.3 (pathogen response 4.3), expected under the trench-warfare evolution model. Finally, a drought-tolerance candidate related to the plant cell wall, lp5, displayed patterns consistent with balancing selection. In conclusion, both arms-race and trench-warfare models seem compatible with patterns of polymorphism found in different disease-response candidate genes, indicating a mixed strategy of disease tolerance evolution for loblolly pine, a major tree crop in southeastern United States

Do the mutations of C1GALT1C1 gene play important roles in the genetic susceptibility to Chinese IgA nephropathy?

Background: The deficiency of beta 1,3 galactose in hinge region of IgA1 molecule played a pivotal role in pathogenesis of IgA nephropathy (IgAN). Cosmc, encoded by C1GALT1C1 gene, was indispensable to beta 1,3 galactosylation of IgA1. We designed a serial study to investigate the relationship between the mutations of C1GALT1C1 gene and the genetic susceptibility to IgAN. Methods: Nine hundred and thirty-eight subjects, including 661 patients with IgAN and 277 healthy controls were enrolled in the study. Firstly, single nucleotide polymorphisms (SNPs) in the promoter region of C1GALT1C1 gene were screened. Then the c.-347-190G&gt; A was analyzed by PCR-restriction fragment length polymorphism (PCR-RFLP) for further case-control association analysis. Secondly the somatic mutations of DNAs from peripheral blood B lymphocytes were detected in 15 patients and 7 normal controls. Results: No significant association was observed between the different alleles or genotypes of c.347-190G&gt;A and IgAN. The patients with different genotypes of C1GALT1C1 gene did not significantly associate with clinical manifestations, including hematuria, proteinuria, and serum creatinine of patients with IgAN. There was no somatic mutation detected in total 202 clones of 22 individuals. Conclusion: The c.-347-190G&gt;A polymorphism and the somatic mutation of encoding region of C1GALT1C1 gene were not significantly related to the genetic susceptibility to IgAN in Northern Chinese population.http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000271285100001&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=8e1609b174ce4e31116a60747a720701Genetics &amp; HereditySCI(E)PubMed1ARTICLE1011

Foliar water uptake: a common water acquisition strategy for plants of the redwood forest

Evaluations of plant water use in ecosystems around the world reveal a shared capacity by many different species to absorb rain, dew, or fog water directly into their leaves or plant crowns. This mode of water uptake provides an important water subsidy that relieves foliar water stress. Our study provides the first comparative evaluation of foliar uptake capacity among the dominant plant taxa from the coast redwood ecosystem of California where crown-wetting events by summertime fog frequently occur during an otherwise drought-prone season. Previous research demonstrated that the dominant overstory tree species, Sequoia sempervirens, takes up fog water by both its roots (via drip from the crown to the soil) and directly through its leaf surfaces. The present study adds to these early findings and shows that 80% of the dominant species from the redwood forest exhibit this foliar uptake water acquisition strategy. The plants studied include canopy trees, understory ferns, and shrubs. Our results also show that foliar uptake provides direct hydration to leaves, increasing leaf water content by 2–11%. In addition, 60% of redwood forest species investigated demonstrate nocturnal stomatal conductance to water vapor. Such findings indicate that even species unable to absorb water directly into their foliage may still receive indirect benefits from nocturnal leaf wetting through suppressed transpiration. For these species, leaf-wetting events enhance the efficacy of nighttime re-equilibration with available soil water and therefore also increase pre-dawn leaf water potentials

Cardiac autonomic neuropathy in patients with diabetes and no symptoms of coronary artery disease: comparison of 123I-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability

PURPOSE The purpose of this study was to evaluate the prevalence of cardiac autonomic neuropathy (CAN) in a cohort of patients with type 2 diabetes, truly asymptomatic for coronary artery disease (CAD), using heart rate variability (HRV) and (123)I-metaiodobenzylguanidine ((123)I-mIBG) myocardial scintigraphy. METHODS The study group comprised 88 patients with type 2 diabetes prospectively recruited from an outpatient diabetes clinic. In all patients myocardial perfusion scintigraphy, CAN by HRV and (123)I-mIBG myocardial scintigraphy were performed. Two or more abnormal tests were defined as CAN-positive (ECG-based CAN) and one or fewer as CAN-negative. CAN assessed by (123)I-mIBG scintigraphy was defined as abnormal if the heart-to-mediastinum ratio was 25%, or the total defect score was >13. RESULTS The prevalence of CAN in patients asymptomatic for CAD with type 2 diabetes and normal myocardial perfusion assessed by HRV and (123)I-mIBG scintigraphy was respectively, 27% and 58%. Furthermore, in almost half of patients with normal HRV, (123)I-mIBG scintigraphy showed CAN. CONCLUSION The current study revealed a high prevalence of CAN in patients with type 2 diabetes. Secondly, disagreement between HRV and (123)I-mIBG scintigraphy for the assessment of CAN was observed.Cardiovascular Aspects of Radiolog

The Genome of Borrelia recurrentis, the Agent of Deadly Louse-Borne Relapsing Fever, Is a Degraded Subset of Tick-Borne Borrelia duttonii

In an effort to understand how a tick-borne pathogen adapts to the body louse, we sequenced and compared the genomes of the recurrent fever agents Borrelia recurrentis and B. duttonii. The 1,242,163–1,574,910-bp fragmented genomes of B. recurrentis and B. duttonii contain a unique 23-kb linear plasmid. This linear plasmid exhibits a large polyT track within the promoter region of an intact variable large protein gene and a telomere resolvase that is unique to Borrelia. The genome content is characterized by several repeat families, including antigenic lipoproteins. B. recurrentis exhibited a 20.4% genome size reduction and appeared to be a strain of B. duttonii, with a decaying genome, possibly due to the accumulation of genomic errors induced by the loss of recA and mutS. Accompanying this were increases in the number of impaired genes and a reduction in coding capacity, including surface-exposed lipoproteins and putative virulence factors. Analysis of the reconstructed ancestral sequence compared to B. duttonii and B. recurrentis was consistent with the accelerated evolution observed in B. recurrentis. Vector specialization of louse-borne pathogens responsible for major epidemics was associated with rapid genome reduction. The correlation between gene loss and increased virulence of B. recurrentis parallels that of Rickettsia prowazekii, with both species being genomic subsets of less-virulent strains

Analyses of an Expressed Sequence Tag Library from Taenia solium, Cysticerca

A method used to describe expressed genes at a specific stage in an organism is an EST library. In this method mRNA from a specific organism is isolated, transcribed into cDNA and sequenced. The sequence will derive from the 5′-end of the cDNA. The library will not have sequences from all genes, especially if they are expressed in low amounts or not at all in the studied stage. Also the library will mostly not contain full length sequences from genes, but expression patterns can be established. If EST libraries are made from different stages of the same organisms these libraries can be compared and differently expressed genes can be identified. Described here is an analysis of an EST library from the pig cysticerca which is thought to be similar to the stage giving the human neglected disease neurocysticercosis. Novel genes together with putative drug targets are examples of data presented