Fine-mapping the HOXB region detects common variants tagging a rare coding allele : evidence for synthetic association in prostate cancer

The HOXB13 gene has been implicated in prostate cancer (PrCa) susceptibility. We performed a high resolution fine-mapping analysis to comprehensively evaluate the association between common genetic variation across the HOXB genetic locus at 17q21 and PrCa risk. This involved genotyping 700 SNPs using a custom Illumina iSelect array (iCOGS) followed by imputation of 3195 SNPs in 20,440 PrCa cases and 21,469 controls in The PRACTICAL consortium. We identified a cluster of highly correlated common variants situated within or closely upstream of HOXB13 that were significantly associated with PrCa risk, described by rs117576373 (OR 1.30, P = 2.62×10(-14)). Additional genotyping, conditional regression and haplotype analyses indicated that the newly identified common variants tag a rare, partially correlated coding variant in the HOXB13 gene (G84E, rs138213197), which has been identified recently as a moderate penetrance PrCa susceptibility allele. The potential for GWAS associations detected through common SNPs to be driven by rare causal variants with higher relative risks has long been proposed; however, to our knowledge this is the first experimental evidence for this phenomenon of synthetic association contributing to cancer susceptibility.

An organization of visual and auditory fear conditioning in the lateral amygdala

Pavlovian fear conditioning is an evolutionary conserved and extensively studied form of associative learning and memory. In mammals, the lateral amygdala (LA) is an essential locus for Pavlovian fear learning and memory. Despite significant progress unraveling the cellular mechanisms responsible for fear conditioning, very little is known about the anatomical organization of neurons encoding fear conditioning in the LA. One key question is how fear conditioning to different sensory stimuli is organized in LA neuronal ensembles. Here we show that Pavlovian fear conditioning, formed through either the auditory or visual sensory modality, activates a similar density of LA neurons expressing a learning-induced phosphorylated extracellular signal-regulated kinase (p-ERK1/2). While the size of the neuron population specific to either memory was similar, the anatomical distribution differed. Several discrete sites in the LA contained a small but significant number of p-ERK1/2-expressing neurons specific to either sensory modality. The sites were anatomically localized to different levels of the longitudinal plane and were independent of both memory strength and the relative size of the activated neuronal population, suggesting some portion of the memory trace for auditory and visually cued fear conditioning is allocated differently in the LA. Presenting the visual stimulus by itself did not activate the same p-ERK1/2 neuron density or pattern, confirming the novelty of light alone cannot account for the specific pattern of activated neurons after visual fear conditioning. Together, these findings reveal an anatomical distribution of visual and auditory fear conditioning at the level of neuronal ensembles in the LA.

Age-related changes in perception of movement in driving scenes

Purpose Age-related changes in motion sensitivity have been found to relate to reductions in various indices of driving performance and safety. The aim of this study was to investigate the basis of this relationship in terms of determining which aspects of motion perception are most relevant to driving. Methods Participants included 61 regular drivers (age range 22–87 years). Visual performance was measured binocularly. Measures included visual acuity, contrast sensitivity and motion sensitivity assessed using four different approaches: (1) threshold minimum drift rate for a drifting Gabor patch, (2) Dmin from a random dot display, (3) threshold coherence from a random dot display, and (4) threshold drift rate for a second-order (contrast modulated) sinusoidal grating. Participants then completed the Hazard Perception Test (HPT) in which they were required to identify moving hazards in videos of real driving scenes, and also a Direction of Heading task (DOH) in which they identified deviations from normal lane keeping in brief videos of driving filmed from the interior of a vehicle. Results In bivariate correlation analyses, all motion sensitivity measures significantly declined with age. Motion coherence thresholds, and minimum drift rate threshold for the first-order stimulus (Gabor patch) both significantly predicted HPT performance even after controlling for age, visual acuity and contrast sensitivity. Bootstrap mediation analysis showed that individual differences in DOH accuracy partly explained these relationships, where those individuals with poorer motion sensitivity on the coherence and Gabor tests showed decreased ability to perceive deviations in motion in the driving videos, which related in turn to their ability to detect the moving hazards. Conclusions The ability to detect subtle movements in the driving environment (as determined by the DOH task) may be an important contributor to effective hazard perception, and is associated with age, and an individuals' performance on tests of motion sensitivity. The locus of the processing deficits appears to lie in first-order, rather than second-order motion pathways.

Opsin clines in butterflies suggest novel roles for insect photopigments

Opsins are ancient molecules that enable animal vision by coupling to a vitamin-derived chromophore to form lightsensitive photopigments. The primary drivers of evolutionary diversification in opsins are thought to be visual tasks related to spectral sensitivity and color vision. Typically, only a few opsin amino acid sites affect photopigment spectral sensitivity. We show that opsin genes of the North American butterfly Limenitis arthemis have diversified along a latitudinal cline, consistent with natural selection due to environmental factors. We sequenced single nucleotide(SNP) polymorphisms in the coding regions of the ultraviolet (UVRh), blue (BRh), and long-wavelength (LWRh) opsin genes from ten butterfly populations along the eastern United States and found that a majority of opsin SNPs showed significant clinal variation. Outlier detection and analysis of molecular variance indicated that many SNPs are under balancing selection and show significant population structure. This contrasts with what we found by analysing SNPs in the wingless and EF-1 alpha loci, and from neutral amplified fragment length polymorphisms, which show no evidence of significant locus-specific or genome-wide structure among populations. Using a combination of functional genetic and physiological approaches, including expression in cell culture, transgenic Drosophila, UV-visible spectroscopy, and optophysiology, we show that key BRh opsin SNPs that vary clinally have almost no effect on spectral sensitivity. Our results suggest that opsin diversification in this butterfly is more consistent with natural selection unrelated to spectral tuning. Some of the clinally varying SNPs may instead play a role in regulating opsin gene expression levels or the thermostability of the opsin protein. Lastly, we discuss the possibility that insect opsins might have important, yet-to-be elucidated, adaptive functions in mediating animal responses to abiotic factors, such as temperature or photoperiod.

Variation in the complex carbohydrate biosynthesis loci of Acinetobacter baumannii genomes

Extracellular polysaccharides are major immunogenic components of the bacterial cell envelope. However, little is known about their biosynthesis in the genus Acinetobacter, which includes A. baumannii, an important nosocomial pathogen. Whether Acinetobacter sp. produce a capsule or a lipopolysaccharide carrying an O antigen or both is not resolved. To explore these issues, genes involved in the synthesis of complex polysaccharides were located in 10 complete A. baumannii genome sequences, and the function of each of their products was predicted via comparison to enzymes with a known function. The absence of a gene encoding a WaaL ligase, required to link the carbohydrate polymer to the lipid A-core oligosaccharide (lipooligosaccharide) forming lipopolysaccharide, suggests that only a capsule is produced. Nine distinct arrangements of a large capsule biosynthesis locus, designated KL1 to KL9, were found in the genomes. Three forms of a second, smaller variable locus, likely to be required for synthesis of the outer core of the lipid A-core moiety, were designated OCL1 to OCL3 and also annotated. Each K locus includes genes for capsule export as well as genes for synthesis of activated sugar precursors, and for glycosyltransfer, glycan modification and oligosaccharide repeat-unit processing. The K loci all include the export genes at one end and genes for synthesis of common sugar precursors at the other, with a highly variable region that includes the remaining genes in between. Five different capsule loci, KL2, KL6, KL7, KL8 and KL9 were detected in multiply antibiotic resistant isolates belonging to global clone 2, and two other loci, KL1 and KL4, in global clone 1. This indicates that this region is being substituted repeatedly in multiply antibiotic resistant isolates from these clones.

Structure of the K2 capsule associated with the KL2 gene cluster of Acinetobacter baumannii

The repeat unit structure of the K2 capsule from an extensively antibiotic-resistant Acinetobacter baumannii global clone 2 (GC2) strain was determined. The oligosaccharide contains three simple sugars, d-glucopyranose, d-galatopyranose and N-acetyl-d-galactosamine, and the complex sugar, 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-l-manno-non-2-ulosonic acid (Pse5Ac7Ac or pseudaminic acid), which has not previously been reported in any A. baumannii capsule. The strain was found to carry all the genes required for the synthesis of the sugars and construction of the K2 structure. The linkages catalyzed by the initiating transferase, three glycosyltransferases and the Wzy polymerase were also predicted. Examination of publicly available A. baumannii genome sequences revealed that the same gene cluster, KL2, often occurs in extensively antibiotic-resistant GC2 isolates and in further strain types. The gene module responsible for the synthesis of pseudaminic acid was also detected in four other K loci. A related module including genes for an acylated relative of pseudaminic acid was also found in two new KL types. A polymerase chain reaction scheme was developed to detect all modules containing genes for sugars based on pseudaminic acid and to specifically detect KL2.

A conjugative plasmid carrying the carbapenem resistance gene blaOXA-23 in AbaR4 in an extensively resistant GC1 Acinetobacter baumannii isolate

OBJECTIVES: To locate the acquired bla(OXA-23) carbapenem resistance gene in an Australian A. baumannii global clone 1 (GC1) isolate. METHODS: The genome of the extensively antibiotic-resistant GC1 isolate A85 harbouring bla(OXA-23) in Tn2006 was sequenced using Illumina HiSeq, and the reads were used to generate a de novo assembly. PCR was used to assemble relevant contigs. Sequences were compared with ones in GenBank. Conjugation experiments were conducted. RESULTS: The sporadic GC1 isolate A85, recovered in 2003, was extensively resistant, exhibiting resistance to imipenem, meropenem and ticarcillin/clavulanate, to cephalosporins and fluoroquinolones and to the older antibiotics gentamicin, kanamycin and neomycin, sulfamethoxazole, trimethoprim and tetracycline. Genes for resistance to older antibiotics are in the chromosome, in an AbaR3 resistance island. A second copy of the ampC gene in Tn6168 confers cephalosporin resistance and the gyrA and parC genes have mutations leading to fluoroquinolone resistance. An 86 335 bp repAci6 plasmid, pA85-3, carrying bla(OXA-23) in Tn2006 in AbaR4, was shown to transfer imipenem, meropenem and ticarcillin/clavulanate resistance into a susceptible recipient. A85 also contains two small cryptic plasmids of 2.7 and 8.7 kb. A85 is sequence type ST126 (Oxford scheme) and carries a novel KL15 capsule locus and the OCL3 outer core locus. CONCLUSIONS: A85 represents a new GC1 lineage identified by the novel capsule locus but retains AbaR3 carrying genes for resistance to older antibiotics. Resistance to imipenem, meropenem and ticarcillin/clavulanate has been introduced into A85 by pA85-3, a repAci6 conjugative plasmid carrying Tn2006 in AbaR4.

Modified MLVA for genotyping Queensland invasive Streptococcus pneumoniae

Background Globally, over 800 000 children under five die each year from infectious diseases caused by Streptococcus pneumoniae. To understand genetic relatedness between isolates, study transmission routes, assess the impact of human interventions e.g. vaccines, and determine infection sources, genotyping methods are required. The ‘gold standard’ genotyping method, Multi-Locus Sequence Typing (MLST), is useful for long-term and global studies. Another genotyping method, Multi-Locus Variable Number of Tandem Repeat Analysis (MLVA), has emerged as a more discriminatory, inexpensive and faster technique; however there is no universally accepted method and it is currently suitable for short-term and localised epidemiology studies. Currently Australia has no national MLST database, nor has it adopted any MLVA method for short-term or localised studies. This study aims to improve S. pneumoniae genotyping methods by modifying the existing MLVA techniques to be more discriminatory, faster, cheaper and technically less demanding than previously published MLVA methods and MLST. Methods Four different MLVA protocols, including a modified method, were applied to 317 isolates of serotyped invasive S. pneumoniae isolated from sterile body sites of Queensland children under 15 years from 2007–2012. MLST was applied to 202 isolates for comparison. Results The modified MLVA4 is significantly more discriminatory than the ‘gold standard’ MLST method. MLVA4 has similar discrimination compared to other MLVA techniques in this study). The failure to amplify particular loci in previous MLVA methods were minimised in MLVA4. Failure to amplify BOX-13 and Spneu19 were found to be serotype specific. Conclusion We have modified a highly discriminatory MLVA technique for genotyping Queensland invasive S. pneumoniae. MLVA4 has the ability to enhance our understanding of the pneumococcal epidemiology and the changing genetics of the pneumococcus in localised and short-term studies.

Differential interaction between the C2B domain of synaptotagmin-I and the Drosophila stonedA and stonedB proteins

The stoned locus in Drosophila encodes two proteins StonedA (STNA) and StonedB (STNB), both of which have been suggested to act as adaptins in mediating synaptic vesicle recycling. A combination of immunological, genetic and biochemical studies have shown an interaction of STNA and STNB with the C2B domain of Synaptotagmin-I (SYT-1), an integral synaptic vesicle protein that mediates Ca2+-dependent exocytosis, as well as endocytosis. The C2B domain of SYT-1 contains an AP-2 binding site that controls the size of recycled vesicles, and a C-terminal tryptophan-containing motif that acts as an internalization signal. Investigation of SYT-1 mutations in Drosophila has shown that altering the Ca2+ binding region of the C2B domain, results in a reduction in the rate of vesicle recycling, implicating this region in SYT-I endocytosis. In this poster, we report the molecular dissection of the interactions between the STNA and STNB proteins and the C2B domain of SYT-1. Deletion of the AP-2 binding site decreased the binding of both STNA and STNB. However, C-terminal deletions of the C2B domain significantly increased STNB binding. In contrast, the same C-terminal deletions reduced the affinity of the C2B domain for STNA. The possible interactions of both STNB and STNA with the Ca2+ binding region of SYT-1 will be also investigated.

A perfusion fMRI investigation of thematic and categorical context effects in the spoken production of object names

The context in which objects are presented influences the speed at which they are named. We employed the blocked cyclic naming paradigm and perfusion functional magnetic resonance imaging (fMRI) to investigate the mechanisms responsible for interference effects reported for thematicallyand categorically related compared to unrelated contexts. Naming objects in categorically homogeneous contexts induced a significant interference effect that accumulated from the second cycle onwards. This interference effect was associated with significant perfusion signal decreases in left middle and posterior lateral temporal cortex and the hippocampus. By contrast, thematically homogeneous contexts facilitated naming latencies significantly in the first cycle and did not differ from heterogeneous contexts thereafter, nor were they associated with any perfusion signal changes compared to heterogeneous contexts. These results are interpreted as being consistent with an account in which the interference effect both originates and has its locus at the lexical level, with an incremental learning mechanism adapting the activation levels of target lexical representations following access. We discuss the implications of these findings for accounts that assume thematic relations can be active lexical competitors or assume mandatory involvement of top-down control mechanisms in interference effects during naming.

Classic identity negative priming involves accessing semantic representations in the left anterior temporal cortex

Classic identity negative priming (NP) refers to the finding that when an object is ignored, subsequent naming responses to it are slower than when it has not been previously ignored (Tipper, S.P., 1985. The negative priming effect: inhibitory priming by ignored objects. Q. J. Exp. Psychol. 37A, 571-590). It is unclear whether this phenomenon arises due to the involvement of abstract semantic representations that the ignored object accesses automatically. Contemporary connectionist models propose a key role for the anterior temporal cortex in the representation of abstract semantic knowledge (e.g., McClelland, J.L., Rogers, T.T., 2003. The parallel distributed processing approach to semantic cognition. Nat. Rev. Neurosci. 4, 310-322), suggesting that this region should be involved during performance of the classic identity NP task if it involves semantic access. Using high-field (4 T) event-related functional magnetic resonance imaging, we observed increased BOLD responses in the left anterolateral temporal cortex including the temporal pole that was directly related to the magnitude of each individual's NP effect, supporting a semantic locus. Additional signal increases were observed in the supplementary eye fields (SEF) and left inferior parietal lobule (IPL).

Orthographic/phonological facilitation of naming responses in the picture-word task: An event-related fMRI study using overt vocal responding

In the picture-word interference task, naming responses are facilitated when a distractor word is orthographically and phonologically related to the depicted object as compared to an unrelated word. We used event-related functional magnetic resonance imaging (fMRI) to investigate the cerebral hemodynamic responses associated with this priming effect. Serial (or independent-stage) and interactive models of word production that explicitly account for picture-word interference effects assume that the locus of the effect is at the level of retrieving phonological codes, a role attributed recently to the left posterior superior temporal cortex (Wernicke's area). This assumption was tested by randomly presenting participants with trials from orthographically related and unrelated distractor conditions and acquiring image volumes coincident with the estimated peak hemodynamic response for each trial. Overt naming responses occurred in the absence of scanner noise, allowing reaction time data to be recorded. Analysis of this data confirmed the priming effect. Analysis of the fMRI data revealed blood oxygen level-dependent signal decreases in Wernicke's area and the right anterior temporal cortex, whereas signal increases were observed in the anterior cingulate, the right orbitomedial prefrontal, somatosensory, and inferior parietal cortices, and the occipital lobe. The results are interpreted as supporting the locus for the facilitation effect as assumed by both classes of theoretical model of word production. In addition, our results raise the possibilities that, counterintuitively, picture-word interference might be increased by the presentation of orthographically related distractors, due to competition introduced by activation of phonologically related word forms, and that this competition requires inhibitory processes to be resolved. The priming effect is therefore viewed as being sufficient to offset the increased interference. We conclude that information from functional imaging studies might be useful for constraining theoretical models of word production.

The semantic interference effect in the picture-word paradigm: An event-related fMRI study employing overt responses

We used event-related functional magnetic resonance imaging (fMRI) to investigate neural responses associated with the semantic interference (SI) effect in the picture-word task. Independent stage models of word production assume that the locus of the SI effect is at the conceptual processing level (Levelt et al. [1999]: Behav Brain Sci 22:1-75), whereas interactive models postulate that it occurs at phonological retrieval (Starreveld and La Heij [1996]: J Exp Psychol Learn Mem Cogn 22:896-918). In both types of model resolution of the SI effect occurs as a result of competitive, spreading activation without the involvement of inhibitory links. These assumptions were tested by randomly presenting participants with trials from semantically-related and lexical control distractor conditions and acquiring image volumes coincident with the estimated peak hemodynamic response for each trial. Overt vocalization of picture names occurred in the absence of scanner noise, allowing reaction time (RT) data to be collected. Analysis of the RT data confirmed the SI effect. Regions showing differential hemodynamic responses during the SI effect included the left mid section of the middle temporal gyrus, left posterior superior temporal gyrus, left anterior cingulate cortex, and bilateral orbitomedial prefrontal cortex. Additional responses were observed in the frontal eye fields, left inferior parietal lobule, and right anterior temporal and occipital cortex. The results are interpreted as indirectly supporting interactive models that allow spreading activation between both conceptual processing and phonological retrieval levels of word production. In addition, the data confirm that selective attention/response suppression has a role in resolving the SI effect similar to the way in which Stroop interference is resolved. We conclude that neuroimaging studies can provide information about the neuroanatomical organization of the lexical system that may prove useful for constraining theoretical models of word production.

Obesity gene NEGR1 associated with white matter integrity in healthy young adults

Obesity is a crucial public health issue in developed countries, with implications for cardiovascular and brain health as we age. A number of commonly-carried genetic variants are associated with obesity. Here we aim to see whether variants in obesity-associated genes - NEGR1, FTO, MTCH2, MC4R, LRRN6C, MAP2K5, FAIM2, SEC16B, ETV5, BDNF- AS, ATXN2L, ATP2A1, KCTD15, and TNN13K - are associated with white matter microstructural properties, assessed by high angular resolution diffusion imaging (HARDI) in young healthy adults between 20 and 30. years of age from the Queensland Twin Imaging study (QTIM). We began with a multi-locus approach testing how a number of common genetic risk factors for obesity at the single nucleotide polymorphism (SNP) level may jointly influence white matter integrity throughout the brain and found a wide spread genetic effect. Risk allele rs2815752 in NEGR1 was most associated with lower white matter integrity across a substantial portion of the brain. Across the area of significance in the bilateral posterior corona radiata, each additional copy of the risk allele was associated with a 2.2% lower average FA. This is the first study to find an association between an obesity risk gene and differences in white matter integrity. As our subjects were young and healthy, our results suggest that NEGR1 has effects on brain structure independent of its effect on obesity.

Common genetic variants influence human subcortical brain structures

The highly complex structure of the human brain is strongly shaped by genetic influences. Subcortical brain regions form circuits with cortical areas to coordinate movement, learning, memory and motivation, and altered circuits can lead to abnormal behaviour and disease. To investigate how common genetic variants affect the structure of these brain regions, here we conduct genome-wide association studies of the volumes of seven subcortical regions and the intracranial volume derived from magnetic resonance images of 30,717 individuals from 50 cohorts. We identify five novel genetic variants influencing the volumes of the putamen and caudate nucleus. We also find stronger evidence for three loci with previously established influences on hippocampal volume and intracranial volume. These variants show specific volumetric effects on brain structures rather than global effects across structures. The strongest effects were found for the putamen, where a novel intergenic locus with replicable influence on volume (rs945270; P = 1.08×10 -33; 0.52% variance explained) showed evidence of altering the expression of the KTN1 gene in both brain and blood tissue. Variants influencing putamen volume clustered near developmental genes that regulate apoptosis, axon guidance and vesicle transport. Identification of these genetic variants provides insight into the causes of variability in human brain development, and may help to determine mechanisms of neuropsychiatric dysfunction.