Gene Linkage Analysis Research Articles

Identification of a biomarker for Bacillus thuringiensis strains with high toxicity against Spodoptera frugiperda based on insecticidal gene linkage analysis.

Bacillus thuringiensis (Bt) is a Gram-positive bacterium that produces various insecticidal proteins used to control insect pests. Spodoptera frugiperda is a global insect pest which causes serious damage to crops, but bio-insecticides currently available to control this pest have limited activity and so new ones are always being sought. In this study we have tested the hypothesis that a biomarker for strain toxicity could be found that would greatly facilitate the identification of new potential products. Using genomic sequencing data we constructed a linkage network of insecticidal genes from 1957 Bt genomes and found that four gene families, namely cry1A, cry1I, cry2A and vip3A, showed strong linkage. For 95 strains isolated from soil samples we assayed them for toxicity towards S. frugiperda and for the presence of the above gene families. All of the strains that showed high toxicity also contained a member of the vip3A gene family. Two of them were more toxic than a commercially available strain and genomic sequencing identified a number of potentially novel toxin-encoding genes. The presence of a vip3A gene in the genome of a Bt strain proved to be a strong indicator of toxicity towards S. frugiperda validating this biomarker approach as a strategy for future discovery programs. © 2024 Society of Chemical Industry.

Abstract 151: Single Cell Transcriptional And Epigenetic Map Of Human Coronary Artery Disease In A Multi-ethnic Population

Identifying target genes that are causally linked to coronary artery disease (CAD) remains an elusive problem despite the plethora of CAD genome-wide association study (GWAS) risk loci. Here, we performed single nucleus multiomic (RNA + ATAC) sequencing in 44 human coronary arteries and bulk Hi-C in 16 coronaries from multi-ethnic participants. From sequencing 126,804 nuclei, we identified cell type specific transcriptional signatures, chromatin peaks, and gene regulatory networks through peak to gene linkage analysis. Single-cell quantitative trait loci (QTL) analysis identified cell-specific caQTL’s including cell-type specific caQTLs within CAD GWAS loci. The overlap of CAD GWAS loci with ATAC peaks demonstrated that vascular smooth muscle and endothelial cells harbor the greatest genetic risk. To identify proximal and distal putative target genes for CAD loci in a cell type specific manner, we built a enhancer-gene map by leveraging coronary artery ChiP-seq, cell specific RNA/ATAC-seq, and Hi-C based chromatin contact loops. Using this map, we identified distal target genes linked to CAD GWAS loci. Finally, given the effect of ancestry on CAD GWAS risk interpretation, we leveraged Hi-C data to compare the chromatin architecture in coronary tissue from 8 European Americans vs 8 African Americans. A meta-map, sample level, and local ancestry analysis identified ancestry-specific differences in chromatin looping and topologically associated domains. Collectively, our results provide a cell-type specific transcriptomic and epigenetic map which along with bulk chromatin contacts can be used to interpret and prioritize CAD GWAS candidates and nominate cell specific causal target genes.

Cellular Plasticity As Mechanisms to Escape from NOTCH1-Inhibition in T-ALL

Cellular plasticity has recently emerged as an underlying mechanism for treatment refraction in cancer. Hence, characterizing cell states and the transcriptional changes required for state transitions is critical for the development of efficient targeted therapies. Acute lymphoblastic leukemia (T-ALL) is an aggressive hematopoietic malignancy in children and adolescents that is associated with high rates of treatment failure and early relapse. T-ALL patients frequently harbor NOTCH1 activating mutations as the driving oncogene in this disease. Using T-ALL patient-derived xenograft (PDX) models carrying activating NOTCH1 mutations, we aim to u First, to determine how cell state diversity drives treatment refraction in T-ALL, we performed full-length single-cell transcriptome analysis on 3188 immune cells collected from 2 sensitive and 3 refractory PDX models. PDX models were treated with either with NOTCH inhibitor DBZ (Dipenzazepine; 10 μM/kg every other day intravenously) or vehicle. Profiled cells were collected at both a short (5 days) and a late time point (x-150 days) post-treatment to assess the effects of DBZ. Analysis of early hematopoietic and thymic precursor signatures demonstrate expression of immature hematopoietic signatures and acquisition of alternative lineage identities coexisting within the same cell, predominantly in refractory models. Analyses of pre-treatment samples indicated that disruption of the normal developmental hierarchy was associated with blockade at an early stage of T-cell commitment and can predict response to NOTCH inhibition in vivo. Upon treatment, cells underwent dramatic transcriptional reprogramming resulting in expansion of 2 distinct populations that greatly differ in both differentiation stage and developmental trajectory. One population expresses lymphoid differentiation programs, whereas the other population is enriched in genes associated with RNA processing and more immature hematopoietic signatures. Cell fate trajectories assessment using Monocle 2, revealed that consistent with their immature hematopoietic signature, refractory cells map at the start point of the differentiation trajectory, while sensitive cells clustered at the endpoint. We next sought to characterize the differentiation hierarchies that define the immature states. Developmental potential as assessed by CytoTRACE was higher in refractory cells when compared to their sensitive counterpart. To identify determinants of cell fate in a functionally relevant manner, we inferred gene regulatory network configurations using SCENIC and defined state specific regulon activity. Among the most prominently expressed regulons present in mature states, we identified transcriptional regulators involved in T-cell differentiation such as IKAROS, GATA3, and SOX5, while in immature states we observed high transcriptional regulon activity in transcription factors associated with renewal of hematopoietic progenitors such as ATF4, BCLAF1, and MYC in addition to the chromatin remodelers SMARCA4 and EZH2. To assess whether transcriptional rewiring in refractory cells results from greater chromatin accessibility, we performed single-cell ATAC sequencing (10x Genomics) on splenic cells from 1 sensitive and 1 refractory T-ALL PDX model treated with DBZ or vehicle. After integration with corresponding single-cell RNA-sequencing data, annotated ATAC profiles demonstrated greater chromatin accessibility, predominantly in active promotor and enhancer regions, in the refractory model compared to the sensitive model. Peak to gene linkage analysis revealed widespread alterations in cis-regulatory regions near genes associated with immature hematopoietic precursor signatures such as CD33. Furthermore, greater chromatin accessibility in refractory cells was associated with an increased number of enhancer loops per expressed gene as determined by H3K27ac HiChIP analysis. In conclusion, we demonstrated that presence of highly plastic cellular states defined by aberrant differentiation trajectories, distinct transcriptional circuitries, and remodeled chromatin architecture results in treatment escape in T-ALL.

Synergistic effect of amino acid substitutions in CYP51B for prochloraz resistance in Fusarium fujikuroi

Prochloraz has been used to control Fusarium fujikuroi, the causative pathogen of rice bakanae disease. Linkage analysis of FfCYP51 genes in the progenies obtained from crossing prochloraz moderately resistant and sensitive strains suggested that the FfCYP51B gene is involved in prochloraz resistance. Sequence comparison revealed that the prochloraz-resistant strain had an F511S or S312T/F511S substitution in FfCYP51B compared with the sensitive strains. The contribution of the S312T and F511S substitutions in FfCYP51B to prochloraz resistance was investigated by creating S/F-, T/F-, or T/S- types at 312/511 codons from the S/S-type, which is a natural moderately resistant strain, using a gene-editing technique. T/S exhibited the highest prochloraz resistance, followed by S/S-, T/F-, and S/F-types. These results indicated that the S312T and F511S substitutions in FfCYP51B had a synergistic effect on prochloraz resistance in F. fujikuroi.

Comparative co-expression analysis of RNA-Seq transcriptome revealing key genes, miRNA and transcription factor in distinct metabolic pathways in diabetic nerve, eye, and kidney disease.

Diabetes and its related complications are associated with long term damage and failure of various organ systems. The microvascular complications of diabetes considered in this study are diabetic retinopathy, diabetic neuropathy, and diabetic nephropathy. The aim is to identify the weighted co-expressed and differentially expressed genes (DEGs), major pathways, and their miRNA, transcription factors (TFs) and drugs interacting in all the three conditions. The primary goal is to identify vital DEGs in all the three conditions. The overlapped five genes (AKT1, NFKB1, MAPK3, PDPK1, and TNF) from the DEGs and the co-expressed genes were defined as key genes, which differentially expressed in all the three cases. Then the protein-protein interaction network and gene set linkage analysis (GSLA) of key genes was performed. GSLA, gene ontology, and pathway enrichment analysis of the key genes elucidates nine major pathways in diabetes. Subsequently, we constructed the miRNA-gene and transcription factor-gene regulatory network of the five gene of interest in the nine major pathways were studied. hsa-mir-34a-5p, a major miRNA that interacted with all the five genes. RELA, FOXO3, PDX1, and SREBF1 were the TFs interacting with the major five gene of interest. Finally, drug-gene interaction network elucidates five potential drugs to treat the genes of interest. This research reveals biomarker genes, miRNA, TFs, and therapeutic drugs in the key signaling pathways, which may help us, understand the processes of all three secondary microvascular problems and aid in disease detection and management.

A family-based study to identify genetic biomarkers of fibromyalgia: consideration of patients' subgroups.

Evidence from genome-wide and candidate gene association studies, familial aggregation and linkage analyses demonstrate the genetic contribution to fibromyalgia (FM) disease. This study aimed to identify genetic biomarkers of FM and its related comorbid disorders, by exploring 41 polymorphisms potentially involved in FM pathogenesis in families with at least one patient with FM. Core symptoms were assessed, and blood samples collected from 556 patients with FM and 395 healthy relatives. For the genetic study, a final sample of 401 FM patients and 232 healthy controls was selected, discarding patients with concomitant pathologies and controls with chronic pain. A family-based approach using DFAM test (Plink) and SNPs (single nucleotide polymorphisms) combination analyses to compare FM patients vs. controls were first applied. Second, the genotypic distribution of subgroups of FM patients, stratified by severe vs. mild symptoms of pain, depression and sleep impairment, was considered. No evidence of associations with FM per se were detected, using either a family-based approach or SNPs combination analyses. However, considering the subgroups of FM patients, the SNP rs6454674 (CNR1, cannabinoid receptor 1 gene) was found as a potential genetic marker of FM correlated with depression (p<.001). No significant associations using either the family-based analysis or the SNPs combination tests dissociated FM patients and their healthy relatives. FM patients with and without depression showed a significant difference in the genotypic distribution related to the SNP rs6454674 in the cannabinoid receptor 1 gene (CNR1) indicating that FM is not a homogenous disorder.

Linkage Analysis of Genes Controlling the Qualitative Characteristics of Barley Plants in Chromosome 6

On the basis of the genetic analysis of F2 by F3 in the combination M13 × r “e” + o, the recombination frequencies between genetic factors of chromosome 6 (=6H) barley were estimated. The study included the following genes: r “e”(smooth awns “e”), sex 1 (shrunken endosperm xenia 1), o (orange lemma 1), gs 4 (glossy sheath 4), and Amy 1–7 (α-amylase 1, type 7). The Amy 1 locus was identified by separation of alpha-amylase by electrophoresis at pH 8.3. On the basis of the five-point test, the following order to location of the studied loci in chromosome 6 (=6H) was established: 6S–r “e” (=raw 5)–gs 4 (=gsh 4)–o (=rob 1)–cen–Sex 1–Amy 1–6L. The linkage estimation and literature analysis showed that, among the indicated genes, the Amy 1 and Sex 1 loci are located in the long arm of chromosome 6. The other genetic factors are associated with the short arm of this chromosome. The centromere is between the genetic factors of Sex 1 and o (=rob 1). In general, the data presented indicate the need to review the genetic linkage map of chromosome 6 (=6H) barley.

Specific Evolution and Gene Family Expansion of Complement 3 and Regulatory Factor H in Fish.

The complement system comprises a large family of plasma proteins that play a central role in innate and adaptive immunity. To better understand the evolution of the complement system in vertebrates and the contribution of complement to fish immunity comprehensive in silico and expression analysis of the gene repertoire was made. Particular attention was given to C3 and the evolutionary related proteins C4 and C5 and to one of the main regulatory factors of C3b, factor H (Cfh). Phylogenetic and gene linkage analysis confirmed the standing hypothesis that the ancestral c3/c4/c5 gene duplicated early. The duplication of C3 (C3.1 and C3.2) and C4 (C4.1 and C4.2) was likely a consequence of the (1R and 2R) genome tetraploidization events at the origin of the vertebrates. In fish, gene number was not conserved and multiple c3 and cfh sequence related genes were encountered, and phylogenetic analysis of each gene generated two main clusters. Duplication of c3 and cfh genes occurred across the teleosts in a species-specific manner. In common, with other immune gene families the c3 gene expansion in fish emerged through a process of tandem gene duplication. Gilthead sea bream (Sparus aurata), had nine c3 gene transcripts highly expressed in liver although as reported in other fish, extra-hepatic expression also occurs. Differences in the sequence and protein domains of the nine deduced C3 proteins in the gilthead sea bream and the presence of specific cysteine and N-glycosylation residues within each isoform was indicative of functional diversity associated with structure. The diversity of C3 and other complement proteins as well as Cfh in teleosts suggests they may have an enhanced capacity to activate complement through direct interaction of C3 isoforms with pathogenic agents.

Using genomic resources for linkage analysis in Peromyscus with an application for characterizing Dominant Spot

BackgroundPeromyscus are the most common mammalian species in North America and are widely used in both laboratory and field studies. The deer mouse, P. maniculatus and the old-field mouse, P. polionotus, are closely related and can generate viable and fertile hybrid offspring. The ability to generate hybrid offspring, coupled with developing genomic resources, enables researchers to conduct linkage analysis studies to identify genomic loci associated with specific traits.ResultsWe used available genomic data to identify DNA polymorphisms between P. maniculatus and P. polionotus and used the polymorphic data to identify the range of genetic complexity that underlies physiological and behavioral differences between the species, including cholesterol metabolism and genes associated with autism. In addition, we used the polymorphic data to conduct a candidate gene linkage analysis for the Dominant spot trait and determined that Dominant spot is linked to a region of chromosome 20 that contains a strong candidate gene, Sox10. During the linkage analysis, we found that the spot size varied quantitively in affected Peromyscus based on genetic background.ConclusionsThe expanding genomic resources for Peromyscus facilitate their use in linkage analysis studies, enabling the identification of loci associated with specific traits. More specifically, we have linked a coat color spotting phenotype, Dominant spot, with Sox10, a member the neural crest gene regulatory network, and that there are likely two genetic modifiers that interact with Dominant spot. These results establish Peromyscus as a model system for identifying new alleles of the neural crest gene regulatory network.

Genetic diversity and population structure of four Chinese rabbit breeds.

There are a few well-known indigenous breeds of Chinese rabbits in Sichuan and Fujian provinces, for which the genetic diversity and population structure have been poorly investigated. In the present study, we successfully employed the restriction-site-associated DNA sequencing (RAD-seq) approach to comprehensively discover genome-wide SNPs of 104 rabbits from four Chinese indigenous breeds: 30 Sichuan White, 34 Tianfu Black, 32 Fujian Yellow and eight Fujian Black. A total of 7,055,440 SNPs were initially obtained, from which 113,973 high-confidence SNPs (read depth ≥ 3, calling rate = 100% and biallelic SNPs) were selected to study the genetic diversity and population structure. The mean polymorphism information content (PIC) and nucleotide diversity (π) of each breed slightly varied with ranging from 0.2000 to 0.2281 and from 0.2678 to 0.2902, respectively. On the whole, Fujian Yellow rabbits showed the highest genetic diversity, which was followed by Tianfu Black and Sichuan White rabbits. The principal component analysis (PCA) revealed that the four breeds were clearly distinguishable. Our results first reveal the genetic differences among these four rabbit breeds in the Sichuan and Fujian provinces and also provide a high-confidence set of genome-wide SNPs for Chinese indigenous rabbits that could be employed for gene linkage and association analyses in the future.

17p13.3 genomic rearrangement in a Chinese family with split-hand/foot malformation with long bone deficiency: report of a complicated duplication with marked variation in phenotype

BackgroundSplit hand/foot malformation (SHFM) is a genetically heterogeneous limb malformation with variable expressivity. SHFM with tibia or femur aplasia is called SHFM with long bone deficiency (SHFLD). 17p13.3 duplications containing BHLHA9 are associated with SHFLD. Cases with variable SHFLD phenotype and different 17p13.3 duplicated regions are reported. The severity of long bone defect could not be simply explained by BHLHA9 overdosage or 17p13.3 duplication.MethodsA four-generation Chinese SHFM family was recruited. Three family members have long bone defects, one male was severely affected with hypoplasia or aplasia in three of four limbs. Linkage analysis and direct sequencing of candidate genes were used to exclude six responsible genes/loci for isolated SHFM. Array comparative genomic hybridization (CGH) was performed to detect copy number variations on a genome-wide scale, and quantitative real-time polymerase chain reaction (qPCR) assays were designed to validate the identified copy number variation in the index and other family members.ResultsNo mutations were found in genes or loci linked to isolated SHFM. A ~ 966 kb duplication was identified in 17p13.3 by array CGH, in which BHLHA9 surrounding region presented as triplication. The qPCR assays confirmed the indicated 17p13.3 duplication as well as BHLHA9 triplication in all available affected family members and other two asymptomatic carriers. Given the incomplete penetrance in SHFLD, those two carriers were regarded as non-penetrant, which suggested that the genomic rearrangement was co-segregated with malformation in this family.ConclusionsThe present study reports an additional SHFLD family case with 17p13.3 genomic rearrangement. To our knowledge, the 966 kb genomic rearrangement is larger in size than any previously reported SHFLD-associated 17p13.3 duplication, and the present family shows marked phenotypic variability with two asymptomatic carriers and one patient with an extremely severe phenotype. This rare case provides the opportunity to identify underlying genotype-phenotype correlations between SHFLD and 17p13.3 genomic rearrangement.

Advance in molecular genetic research on X-linked syndromic hearing impairment

In addition to hearing impairment, syndromic hearing impairment is often accompanied by disorders of urinary, skeletal, muscular, nervous, and ocular systems. Genetic factors have shown to play an important role in the pathogenesis of deafness. Mutations of X-linked genes may cause syndromic hearing impairment. Gene mapping, linkage analysis and next-generation sequencing may facilitate delineation of the pathogenesis of X-linked syndromic hearing impairment. This article reviews recent progress in molecular genetic research on X-linked syndromic hearing impairment, which may shed light for the diagnosis and treatment of these diseases.

Gene expression and linkage analysis implicate CBLB as a mediator of rituximab resistance.

Elucidating resistance mechanisms for therapeutic monoclonal antibodies (MAbs) is challenging, because they are difficult to study in non-human models. We therefore developed a strategy to genetically map in vitro drug sensitivity, identifying genes that alter responsiveness to rituximab, a therapeutic anti-CD20 MAb that provides significant benefit to patients with B-cell malignancies. We discovered novel loci with genome-wide mapping analyses and functionally validated one of these genes, CBLB, which causes rituximab resistance when knocked down in lymphoma cells. This study demonstrates the utility of genome-wide mapping to discover novel biological mechanisms of potential clinical advantage.

UGT1A1 gene linkage analysis: application of polymorphic markers rs4148326/rs4124874 in the Iranian population.

Objective(s):Mutations in the UGT1A1 gene are responsible for hyperbilirubinemia syndromes including Crigler-Najjar type 1 and 2 and Gilbert syndrome. In view of the genetic heterogeneity and involvement of large numbers of the disease causing mutations, the application of polymorphic markers in the UGTA1 gene could be useful in molecular diagnosis of the disease.Materials and Methods:In the present study, two polymorphic markers including rs4148326 and rs4124874 in the UGT1A1 gene region were characterized. The markers were selected using bioinformatics analysis of the UGT1A1 gene region and genotyped in 212 unrelated healthy individuals and 13 family trios in the Iranian population using Tetra-Primer ARMS PCR technique. The allele frequency and population status of the alleles were estimated using GENEPOP, FBAT, PowerMarker and Arlequin software.Results:The results indicated that in the case of rs4148326 marker, allele frequency for T and C allele was 66.04% and 33.96%, respectively. For rs4124874 marker, allele frequency for G and T alleles was 39.4% and 60.6%, respectively. The values of heterozygosity index for the markers examined were 64.1 for rs4148326 and 72.1 for rs4124874, respectively. The haplotype estimation analysis of the markers resulted in three informative haplotypes with frequencies ≥0.05. Moreover, the results suggested the presence of linkage disequilibrium between two markers.Conclusion:Altogether, the data suggested that rs4148326 and rs4124874 could be introduced as informative markers for molecular diagnosis of Crigler-Najjar type 1 and 2 and Gilbert syndrome in the Iranian population.

Abstract 1487: Gene expression and linkage analysis implicates CBLB as a mediator of rituximab resistance

Abstract Drug resistance remains one of the largest challenges in the curative treatment of cancer. We investigated the problem of monoclonal antibody resistance in vitro in a high throughput, complement-dependent cytotoxic (CDC) assay using immortalized as well as cancerous cell lines. First, the heritability of rituximab and ofatumumab sensitivity was explored using Epstein Barr Virus-immortalized human lymphoblastoid cell lines from two independent sources: a collection of trios (95 samples, 13 trios) from the Centre d’Etude du Polymorphisme Humain and an unrelated collection (486 samples) from the Children's Hospital Oakland Research Initiative. Cell viability in the presence or absence of 10ug/ml rituximab or 10ug/ml ofatumumab treatment was quantitated using AlamarBlue. Heritability was estimated using variance component analysis using MERLIN: H2 = 33.11% (p&amp;lt;0.001) and H2 = 31.11% (p&amp;lt;0.001) for rituximab and ofatumumab, resp. Next, public information on genotype and gene expression was used to analyze associations between loci/genes and drug response with three methods: genome-wide association, linkage, and gene expression analysis. GWAS were performed on the 486 unrelated samples: (a) rituximab responses, (b) ofatumumab responses, and (c) the vector of both responses modeled jointly. There was one suggestive (-log10(P)&amp;gt;6) result across all three analyses; this result for rituximab response occurred within the gene, SMOC2. Next, linkage analysis on the 95 related samples revealed one significant linkage peak on chromosome 12 for rituximab and two significant peaks for ofatumumab, on chromosome 12 and 3. Also, we identified genes whose mRNA expression level was correlated with the degree of either rituximab or ofatumumab sensitivity. Quantitative Significance Analysis of Microarrays revealed 13 genes whose expression was correlated with rituximab sensitivity and 25 genes whose expression was correlated with ofatumumab sensitivity (false discovery rate cutoff&amp;lt;0.001%). CBLB, present on both gene lists, was the only gene that was located in either the chromosome 3 or chromosome 12 linkage peak. As such, CBLB was chosen for further functional validation. We used RNA silencing in several cell line and cancerous models to functionally validate the role of CBLB expression on rituximab resistance. Knockdown of CBLB in cell lines, both LCLs and lymphoma, led to increased resistant to both rituximab and ofatumumab. Additionally, we performed immunofluorescence in CBLB knockdown cells. CD20 localization was altered in cells with CBLB knockdown. Our current work has uncovered new mechanistic insight into the relationship between anti-CD20 antibody susceptibility, CBLB, and CD20, illustrating the power of comprehensive genetic analyses to discover a previously unknown mediator of rituxumab response. Additional studies are ongoing to further elucidate the mechanisms of CBLB-mediated resistance as well as its clinical relevance. Citation Format: John Jack, George Small, Tammy Havener, Chad Brown, Howard McLeod, Alison Motsinger, Kristy L. Richards. Gene expression and linkage analysis implicates CBLB as a mediator of rituximab resistance. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1487.

The evolving spectrum of PRRT2-associated paroxysmal diseases.

Next-generation sequencing has identified mutations in the PRRT2 (proline-rich transmembrane protein 2) gene as the leading cause for a wide and yet evolving spectrum of paroxysmal diseases. PRRT2 mutations are found in the majority of patients with benign familial infantile epilepsy, infantile convulsions and choreoathetosis and paroxysmal kinesigenic dyskinesia, confirming a common disease spectrum that had previously been suggested based on gene linkage analyses and shared clinical features. Beyond these clinical entities, PRRT2 mutations have been described in other childhood-onset movement disorders, different forms of seizures, headache disorders, and intellectual disability. PRRT2 encodes a protein that is expressed in the central nervous system and is thought to be involved in the modulation of synaptic neurotransmitter release. The vast majority of mutations lead to a truncated protein or no protein at all and thus to a haploinsufficient state. The subsequent reduction of PRRT2 protein may lead to altered synaptic neurotransmitter release and dysregulated neuronal excitability in various regions of the brain, resulting in paroxysmal movement disorders and seizure phenotypes. In this review, we examine the genetics and neurobiology of PRRT2 and summarize the evolving clinical and molecular spectrum of PRRT2-associated diseases. Through a comprehensive review of 1444 published cases, we provide a detailed assessment of the demographics, disease characteristics and genetic findings of patients with PRRT2 mutations. Benign familial infantile epilepsy (41.7%; n = 602), paroxysmal kinesigenic dyskinesia (38.7%; n = 560) and infantile convulsions and choreoathetosis (14.3%; n = 206) constitute the vast majority of PRRT2-associated diseases, leaving 76 patients (5.3%) with a different primary diagnosis. A positive family history is present in 89.1% of patients; and PRRT2 mutations are familial in 87.1% of reported cases. Seventy-three different disease-associated PRRT2 mutations (35 truncating, 22 missense, three extension mutations, six putative splice site changes, and seven changes that lead to a complete PRRT2 deletion) have been described to date, with the c.649dupC frameshift mutation accounting for the majority of cases (78.5%). Expanding the genetic landscape, 15 patients with biallelic PRRT2 mutations and six patients with 16p11.2 microdeletions and a paroxysmal kinesigenic dyskinesia phenotype have been reported. Probing the phenotypic boundaries of PRRT2-associated disorders, several movement, seizure and headache disorders have been linked to PRRT2 mutations in a subset of patients. Of these, hemiplegic migraine emerges as a novel PRRT2-associated phenotype. With this comprehensive review of PRRT2-associated diseases, we hope to provide a scientific resource for informing future research, both in laboratory models and in clinical studies.

Applying knowledge of autism to brain cancer management: what do we know?

Autism meets glioblastoma What do two seemingly unrelated disorders of the brain have in common? Autism encompasses a heterogeneous collection of neurodevelopmental dysfunctions ranging from impaired social behavior, stereotyped interests and language deficit. Diagnosed at increasingly high rates of one in 68 children (one in 42 for boys), autism spectrum disorders are a major impediment to childhood development worldwide. Current treatment options for autism are limited [1]. On the other hand, glioblastoma multiforme (GBM) is the most common and aggressive form of adult primary brain cancer with extremely poor 3-year survival rates of less than 10%. In GBM, clinical trials of drugs inhibiting the EGF receptor (EGFR) kinase, the most prominent oncogenic target, have been disappointing and as a result, patient outcomes have failed to show substantive improvement over median survival times of 12 months [2]. Thus, there is a great, unmet need for effective therapy in both disorders and innovative approaches are urgently needed. In order to identify new drug targets, we need a transdisciplinary approach that breaches the conventional boundaries confining each area of research. There is growing recognition that glioblastoma and many brain disorders share the same fundamental pathophysiological mechanisms at a cellular and molecular level. As a consequence, mutations or gene expression changes in the same gene family can underlie multiple, distinct clinical phenotypes. A compelling example is that of the SLC9A6 and SLC9A9 genes encoding endosomal Na/H exchangers NHE6 and NHE9, respectively [1]. Genome-wide association studies, gene linkage analysis, homozygosity mapping, transcription profiling and mining patient databases have linked these genes to a collection of neurological disorders, including autism, attention deficit hyperactivity disorder, epilepsy, intellectual disability, Alzheimer’s disease and other neurodegenerative disorders, and most recently and unexpectedly brain cancer [1–4]. “Meaningful future therapies are likely to require cocktails of drugs that target the proliferation/invasion of cancer and the development of chemoresistance.”

Genome-Wide Association Study in Thai Tsunami Survivors Identified Risk Alleles for Posttraumatic Stress Disorder

Posttraumatic stress disorder (PTSD) is a psychiatric disorder found in individuals afflicted by a traumatic event. Multiple environmental and genetic factors can contribute to PTSD susceptibility. Since it is rare to find members of the same family afflicted by the same catastrophic event, it is not practical to determine PTSD susceptibility genes by a gene linkage analysis. A natural disaster, such as the 2004 Tsunami, provided us with a rare chance for a genetic analysis of PTSD. To identify SNPs associated with PTSD susceptibility, we conducted a genome-association study (GWAS) in Thai-Tsunami survivors. Initial phase of the study with 396 chronic PTSD patients and 457 controls, we identified top ninety SNPs (P -4), which were further assessed in the second phase with 395 chronic PTSD patients and 798 controls. Two SNPs (rs267950 and rs954406), were identified in the second phase, and subjected to fine mapping using a data set from both phases. SNP rs267943 showed the strongest association with PTSD susceptibility and was in complete linkage disequilibrium with SNP rs267950 with P = 6.15 × 10-8, OR = 1.46 and 95% CI = 1.19 - 1.79, reaching genome-wide significance. SNP rs267943 is located on chromosome 5 in the intron of the death-associated protein 1 (DAP1) gene and, when linked to a synthetic promoter, could regulate transcription. To our knowledge, this is the first GWAS for PTSD susceptibility in an Asian population which could provide an important insight into the genetic contribution of PTSD and may lead to new treatment strategies for PTSD.

What genetics has taught us about keratoconus

AbstractKeratoconus most likely represents complex disease, with different modes of inheritance, genetic heterogeneity, and environmental factors contributing to the disease manifestation. The relative contribution of each is subject to debate and likely varies between individuals. At one end of the spectrum, disease in some individuals is probably due entirely to environmental influences, yet at the other end is solely under the control of genetic mechanisms. However the majority of cases will most likely be the result of a genetic predisposition with precipitating environmental stressors or modifiers Over the last decade significant progress has been made in identifying genetic risk factors for keratoconus. These genetic risk factors have been identified using multiple approaches including candidate gene studies, linkage analysis, genome‐wide studies and new sequencing technologies. This presentation will summarize how the genes implicated provide clues to the molecular etiology and pathways involved in the pathogenesis of keratoconus.

English

Modern dairy cattle breeding strategies depend on linkage analysis and quantitative trait loci (QTL) of genes involved in milk yield and composition. This is because of their biological desired quantitative traits that play key roles in milk production. In this study, three genes directly related to milk production: prolactin (PRL), bovine kappa-casein (K-CN) and the pituitary-specii¬c transcription factor (PIT-1) were analyzed in 144 cows. The aim of this study was to identify polymorphisms in the Holstein-Friesian cattle breed in Palestine in relation to the genetic markers and allelic variants of the three genes. Collection of samples depended on an experimental design that was completely randomized (CRD) and blood samples were collected from different cities across the West Bank, Palestine. The genotypes were determined through the polymerase chain reaction-restriction fragments length polymorphism (PCR-RFLP) technique. The amplified fragments of PRL (294-bp), K-CN (530-bp) and PIT-1 (451-bp) were digested with RsaI, HindIII and HinfI, respectively. Statistical analysis found that the prolactin allelic substitution (AG, GG) played a role in milk production with a p-value of 0.00643 and α (0.001**), the AG allele of PRL being more favorable for milk production as compared to the GG allele. Genetic variants of the bovine K-CN gene played a role in milk production with a p-value of 0.04071 and α (0.01*), the AA allele possessing more positive effect than the BB and AB alleles. Similarly, the allelic substitution of the PIT-1 gene affected milk production with a p-value of 2.274e-05 and α (0***), the AA allele exercising a more positive effect followed by the AB and BB alleles, respectively. Among the three studied breeds (Friesian, hybrid and local), results show that the Friesian breed possesses higher overall milk production in Palestine as compared to the other two breeds. Key words: Prolactin (PRL), bovine kappa-casein (K-CN), pituitary-specii¬c transcription factor (PIT-1), polymerase chain reaction-restriction fragments length polymorphism (PCR-RFLP).