The Clinical and Molecular Assessment of Iranian Families with Severe Congenital Neutropenia, Identification of HYOU1 and SHOC2 as Potential Novel Gene Defects.

Neutropenia congenita grave (SCN) is a rare disease with a genetically and clinically heterogeneous nature, usually diagnosed in childhood, with an elevated risk of infections such as otitis, skin infections, pneumonia, deep abscesses, and septicemia. Patients with SCN also have an increased risk of leukemia, and mutations in the ELANE and the HAX1 genes have been observed in those patients. This study was conducted to genetically screen six Iranian families with SCN who have at least one affected person. In the first step, all exons and intron boundaries of ELANE and HAX1 genes were sequenced in probands. Cases with no pathogenic mutations were tested through whole-exome sequencing (WES). Analysis showed five different variants in ELANE (c.377 C>T), HAX1 (c.130_131 insA), HYOU1 (c.69 G>C and c.2744 G>A) and SHOC2 (c.4 A>G) genes in four families. We found that two out of six families had mutations in ELANE and HAX1 genes. Moreover, we found two novel mutations at the HYOU1 gene that had not previously been reported, as well as a pathogenic mutation at SHOC2 with multiple phenotypes, that will contribute to determining the genetic basis for SCN. Our study revealed that WES could help diagnose SCN, improve the classification of neutropenia, and rule out other immunodeficiencies such as autoimmune neutropenia, primary immunodeficiency diseases, and inherited bone marrow failure syndromes.

Assessment of Association between miR-146a Polymorphisms and Expression of miR-146a, TRAF-6, and IRAK-1 Genes in Patients with Brucellosis.

BACKGROUND: Brucellosis is a major zoonosis all over the world. MicroRNAs are significant gene expression regulators and could be involved during the infections and also genetic alterations in the miRNAs sequence can affect primary miRNAs and precursor miRNAs processing and thus alter miRNAs expression. Current research studied the impact of the miR-146a polymorphism on miR-146a, TRAF-6, and IRAK-1 genes expression in patients with brucellosis illness. METHODS AND RESULTS: In this research, 25 patients with brucellosis and 25 healthy participants with determined genotypes for miR-SNP rs2910164 and miR-SNP rs57095329 were recruited. IRAK-1, TRAF-6, and miR-146a expressions in peripheral blood mononuclear cells (PBMCs) were specified by quantitative real- time PCR (qRT-PCR). Moreover, interleukin-1ß (IL-1ß) and tumor necrosis factor- alpha (TNF-α) serum levels were assessed by a sandwich enzyme-linked immunosorbent assay (ELISA) technique. There was no significant difference in the expression level of miR-146a, IRAK-1, and TRAF-6, among the patients with brucellosis and control group. TRAF-6 PBMCs expression levels in the distinctive genotypes of rs2910164 were significantly observed in patients (P = 0.048). No significant distinctions were found in miR-146a, IRAK-1, and TRAF-6 expression levels and among the rs57095329 different genotypes in brucellosis patients and controls. Meanwhile, no significant relationship was found between the rs2910164 and rs57095329 genotypes and the serum level of cytokines mentioned between the two groups. We did not find any association between expression of TRAF-6, miR-146a, and IRAK-1 in PBMCs, and cytokines serum levels with two single nucleotide polymorphisms (SNPs) in miR-146a. CONCLUSIONS: To the best of writers' knowledge, this research is the first one evaluating the probable link between the miR-146a rs2910164 and rs57095329 variant with miRNAs, relevant cytokine levels, and target genes in brucellosis.

Integrative assessment of CIP2A overexpression and mutational effects in human malignancies identifies possible deleterious variants.

KIAA1524 is the gene encoding the human cancerous inhibitor of PP2A (CIP2A) protein which is regarded as a novel target for cancer therapy. It is overexpressed in 65%-90% of tissues in almost all studied human cancers. CIP2A expression correlates with cancer progression, disease aggressivity in lung cancer besides poor survival and resistance to chemotherapy in breast cancer. Herein, a pan-cancer analysis of public gene expression datasets was conducted showing significant upregulation of CIP2A in cancerous and metastatic tissues. CIP2A overexpression also correlated with poor survival of cancer patients. To determine the non-coding variants associated with CIP2A overexpression, 5'UTR and 3'UTR variants were annotated and scored using RegulomeDB and Enformer deep learning model. The 5'UTR variants rs1239349555, rs1576326380, and rs1231839144 were predicted to be potential regulators of CIP2A overexpression scoring best on RegulomeDB annotations with a high "2a" rank of supporting experimental data. These variants also scored the highest on Enformer predictions. Analysis of the 3'UTR variants of CIP2A predicted rs56255137 and rs58758610 to alter binding sites of hsa-miR-500a-5 and (hsa-miR-3671, hsa-miR-5692a) respectively. Both variants were also found in linkage disequilibrium with rs11709183 and rs147863209 respectively at r2 ≥ 0.8. The aforementioned variants were found to be eQTL hits significantly associated with CIP2A overexpression. Further, analysis of rs11709183 and rs147863209 revealed a high "2b" rank on RegulomeDB annotations indicating a probable effect on DNAse transcription factors binding. The MuTarget analysis indicated that somatic mutations in TP53 are significantly associated with upregulated CIP2A in human cancers. Analysis of missense SNPs on CIP2A solved structure predicted seven deleterious effects. Four of these variants were also predicted as structurally and functionally destabilizing to CIP2A including; rs375108755, rs147942716, rs368722879, and rs367941403. Variant rs1193091427 was predicted as a potential intronic splicing mutation that might be responsible for the novel CIP2A variant (NOCIVA) in multiple myeloma. Finally, Enrichment of the Wnt/ß-catenin pathway within the CIP2A regulatory gene network suggested potential of therapeutic combinations between FTY720 with Wnt/ß-catenin, Plk1 and/or HDAC inhibitors to downregulate CIP2A which has been shown to be essential for the survival of different cancer cell lines.

Discovery of a hidden transient state in all bromodomain families.

Bromodomains (BDs) are small protein modules that interact with acetylated marks in histones. These posttranslational modifications are pivotal to regulate gene expression, making BDs promising targets to treat several diseases. While the general structure of BDs is well known, their dynamical features and their interplay with other macromolecules are poorly understood, hampering the rational design of potent and selective inhibitors. Here, we combine extensive molecular dynamics simulations, Markov state modeling, and available structural data to reveal a transiently formed state that is conserved across all BD families. It involves the breaking of two backbone hydrogen bonds that anchor the ZA-loop with the αA helix, opening a cryptic pocket that partially occludes the one associated to histone binding. By analyzing more than 1,900 experimental structures, we unveil just two adopting the hidden state, explaining why it has been previously unnoticed and providing direct structural evidence for its existence. Our results suggest that this state is an allosteric regulatory switch for BDs, potentially related to a recently unveiled BD-DNA-binding mode.

Race, Ethnicity, and Clinical Outcomes in Hormone Receptor-Positive, HER2-Negative, Node-Negative Breast Cancer in the Randomized TAILORx Trial.

BACKGROUND: Black race is associated with worse outcomes in early breast cancer. We evaluated clinicopathologic characteristics, the 21-gene recurrence score (RS), treatment delivered, and clinical outcomes by race and ethnicity among women who participated in the Trial Assigning Individualized Options for Treatment. METHODS: The association between clinical outcomes and race (White, Black, Asian, other or unknown) and ethnicity (Hispanic vs non-Hispanic) was examined using proportional hazards models. All P values are 2-sided. RESULTS: Of 9719 eligible women with hormone receptor-positive, HER2-negative, node-negative breast cancer, there were 8189 (84.3%) Whites, 693 (7.1%) Blacks, 405 (4.2%) Asians, and 432 (4.4%) with other or unknown race. Regarding ethnicity, 889 (9.1%) were Hispanic. There were no substantial differences in RS or ESR1, PGR, or HER2 RNA expression by race or ethnicity. After adjustment for other covariates, compared with White race, Black race was associated with higher distant recurrence rates (hazard ratio [HR] = 1.60, 95% confidence intervals [CI] = 1.07 to 2.41) and worse overall survival in the RS 11-25 cohort (HR = 1.51, 95% CI = 1.06 to 2.15) and entire population (HR = 1.41, 95% CI = 1.05 to 1.90). Hispanic ethnicity and Asian race were associated with better outcomes. There was no evidence of chemotherapy benefit for any racial or ethnic group in those with a RS of 11-25. CONCLUSIONS: Black women had worse clinical outcomes despite similar 21-gene assay RS results and comparable systemic therapy in the Trial Assigning Individualized Options for Treatment. Similar to Whites, Black women did not benefit from adjuvant chemotherapy if the 21-gene RS was 11-25. Further research is required to elucidate the basis for this racial disparity in prognosis.

Application of a glycinated bile acid biomarker for diagnosis and assessment of response to treatment in Niemann-pick disease type C1.

Niemann-Pick disease type C (NPC) is a neurodegenerative disease in which mutation of NPC1 or NPC2 gene leads to lysosomal accumulation of unesterified cholesterol and sphingolipids. Diagnosis of NPC disease is challenging due to non-specific early symptoms. Biomarker and genetic tests are used as first-line diagnostic tests for NPC. In this study, we developed a plasma test based on N-(3ß,5α,6ß-trihydroxy-cholan-24-oyl)glycine (TCG) that was markedly increased in the plasma of human NPC1 subjects. The test showed sensitivity of 0.9945 and specificity of 0.9982 to differentiate individuals with NPC1 from NPC1 carriers and controls. Compared to other commonly used biomarkers, cholestane-3ß,5α,6ß-triol (C-triol) and N-palmitoyl-O-phosphocholine (PPCS, also referred to as lysoSM-509), TCG was equally sensitive for identifying NPC1 but more specific. Unlike C-triol and PPCS, TCG showed excellent stability and no spurious generation of marker in the sample preparation or aging of samples. TCG was also elevated in lysosomal acid lipase deficiency (LALD) and acid sphingomyelinase deficiency (ASMD). Plasma TCG was significantly reduced after intravenous (IV) 2-hydroxypropyl-ß-cyclodextrin (HPßCD) treatment. These results demonstrate that plasma TCG was superior to C-triol and PPCS as NPC1 diagnostic biomarker and was able to evaluate the peripheral treatment efficacy of IV HPßCD treatment.

A pan-cancer assessment of alterations of the kinase domain of ULK1, an upstream regulator of autophagy.

Autophagy is a key clearance process to recycle damaged cellular components. One important upstream regulator of autophagy is ULK1 kinase. Several three-dimensional structures of the ULK1 catalytic domain are available, but a comprehensive study, including molecular dynamics, is missing. Also, an exhaustive description of ULK1 alterations found in cancer samples is presently lacking. We here applied a framework which links -omics data to structural protein ensembles to study ULK1 alterations from genomics data available for more than 30 cancer types. We predicted the effects of mutations on ULK1 function and structural stability, accounting for protein dynamics, and the different layers of changes that a mutation can induce in a protein at the functional and structural level. ULK1 is down-regulated in gynecological tumors. In other cancer types, ULK2 could compensate for ULK1 downregulation and, in the majority of the cases, no marked changes in expression have been found. 36 missense mutations of ULK1, not limited to the catalytic domain, are co-occurring with mutations in a large number of ULK1 interactors or substrates, suggesting a pronounced effect of the upstream steps of autophagy in many cancer types. Moreover, our results pinpoint that more than 50% of the mutations in the kinase domain of ULK1, here investigated, are predicted to affect protein stability. Three mutations (S184F, D102N, and A28V) are predicted with only impact on kinase activity, either modifying the functional dynamics or the capability to exert effects from distal sites to the functional and catalytic regions. The framework here applied could be extended to other protein targets to aid the classification of missense mutations from cancer genomics studies, as well as to prioritize variants for experimental validation, or to select the appropriate biological readouts for experiments.

Multi-state recognition pathway of the intrinsically disordered protein kinase inhibitor by protein kinase A.

In the nucleus, the spatiotemporal regulation of the catalytic subunit of cAMP-dependent protein kinase A (PKA-C) is orchestrated by an intrinsically disordered protein kinase inhibitor, PKI, which recruits the CRM1/RanGTP nuclear exporting complex. How the PKA-C/PKI complex assembles and recognizes CRM1/RanGTP is not well understood. Using NMR, SAXS, fluorescence, metadynamics, and Markov model analysis, we determined the multi-state recognition pathway for PKI. After a fast binding step in which PKA-C selects PKI's most competent conformations, PKI folds upon binding through a slow conformational rearrangement within the enzyme's binding pocket. The high-affinity and pseudo-substrate regions of PKI become more structured and the transient interactions with the kinase augment the helical content of the nuclear export sequence, which is then poised to recruit the CRM1/RanGTP complex for nuclear translocation. The multistate binding mechanism featured by PKA-C/PKI complex represents a paradigm on how disordered, ancillary proteins (or protein domains) are able to operate multiple functions such as inhibiting the kinase while recruiting other regulatory proteins for nuclear export.

Decision tree learning to predict overweight/obesity based on body mass index and gene polymporphisms.

The new technologies for data analysis, such as decision tree learning, may help to predict the risk of developing diseases. The aim of the present work was to develop a pilot decision tree learning to predict overweight/obesity based on the combination of six single nucleotide polymorphisms (SNP) located in feeding-associated genes. Genotype study was performed in 151 healthy individuals, who were anonymized and randomly selected from the TALAVERA study. The decision tree analysis was performed using the R package rpart. The learning process was stopped when 15 or less observation was found in a node. The participant group consisted of 78 men and 73 women, who 100 individuals showed body mass index (BMI) ≥ 25 kg/m2 and 51 BMI < 25 kg/m2. Chi-square analysis revealed that individuals with BMI ≥ 25 kg/m2 showed higher frequency of the allelic variation Ala67Ala in AgRP rs5030980 with respect to those with BMI <25 kg/m2. However, the variant Thr67Ala in AgRP rs5030980 was the most frequently found in individuals with BMI <25 kg/m2. There were no statistical differences in the other analyzed SNPs. Decision tree learning revealed that carriers of the allelic variants AgRP (rs5030980) Ala67Ala, ADRB2 (rs1042714) Gln27Glu or Glu27Glu, INSIG2 (rs7566605) 73 + 9802 with CC or GG genotypes and PPARG (rs1801282) with the allelic variants of Ala12Ala or Pro12Pro, will most likely develop overweight/obesity (BMI ≥ 25 kg/m2). Moreover, the decision tree learning indicated that age and gender may change the developed three decision learning associated with overweight/obesity development. The present work should be considered as a pilot demonstrative study to reinforce the broad field of application of new data analysis technologies, such as decision tree learning, as useful tools for diseases prediction. This technology may achieve a potential applicability in the design of early strategies to prevent overweight/obesity.

KIBRA and APOE Gene Variants Affect Brain Functional Network Connectivity in Healthy Older People.

Genetic factors play a critical role in the development of Alzheimer's disease (AD). Kidney and brain expressed protein (KIBRA) and apolipoprotein E (APOE) are involved in episodic memory performance and AD. However, the interactions between KIBRA and APOE on brain functional network connectivity (FNC) remain unknown in healthy older people. Using independent component analysis, we systematically investigated additive and epistatic interactions of KIBRA rs1707045 and APOE on FNC in 170 healthy older Chinese people of Han ethnicity. We found significant additive KIBRA-APOE interactions on brain FNC in the right medial prefrontal cortex, the posterior cingulate cortex in the default-mode network, and the dorsal anterior cingulate cortex in the salience network. We also found significant epistatic KIBRA-APOE interactions on brain FNC in the left superior frontal gyrus and left angular gyrus in default-mode network. No significant KIBRA-APOE interactions were detected in other brain resting-state networks. These findings suggest that healthy older people have additive and epistatic interactions of KIBRA and APOE gene variants, which modulate brain FNC and may partly elucidate their association with episodic memory performance and AD.

The mutation-dependent pathogenicity of NPHS2 p.R229Q: A guide for clinical assessment.

NPHS2, encoding podocin, is the major gene implicated in steroid-resistant nephrotic syndrome. Its c.686G>A, p.R229Q variant is the first human variant with a mutation-dependent pathogenicity; it is only pathogenic when trans-associated to specific mutations. Secondary to its high allele frequency in the European, South Asian, African, and Latino populations, its benign trans-associations can be accidentally identified in affected patients. Distinguishing pathogenic and benign p.R229Q associations can be challenging. In this paper, we present the currently known pathogenic and benign associations, and show that a rare p.R229Q association can be considered pathogenic if the variant in trans meets the following criteria; it affects the 270-351 residues and alters but does not disrupt the oligomerization, its p.R229Q association is found in a family with slowly progressing focal segmental glomerulosclerosis, but is expected to be rare in the general population (<1:106 ). We show that >15% of the p.R229Q associations identified so far in patients are benign.

A novel Tetra-primer ARMS-PCR based assay for genotyping SNP rs12303764(G/T) of human Unc-51 like kinase 1 gene.

Various case-control studies have shown association of single nucleotide polymorphism rs12303764(G/T) in ULK1 with crohn's disease. The techniques used in these studies were time consuming, complicated and require sophisticated/expensive instruments. Therefore, in order to overcome these problems, we have developed a new, rapid and cost effective Tetra-primer ARMS-PCR assay to genotype single nucleotide polymorphism rs12303764(G/T) of ULK1 gene. We manually designed allele specific primers. DNA fragment amplified using outer primers was sequenced to obtain samples with known genotypes (GG, GT and TT) for further use in the development of T-ARMS-PCR assay. Amplification conditions were optimized for parameters; annealing temperature, Taq DNA polymerase and primers. The developed T-ARMS-PCR assay was applied to genotype one hundred samples from healthy individuals. Genotyping results of 10 DNA samples from healthy individuals for rs12303764(G/T) by T-ARMS-PCR assay and sequencing were concordant. The newly developed assay was further applied to genotype samples from 100 healthy individuals of North Indian origin. Genotype frequencies were 9, 34 and 57 % for GG, GT and TT, respectively. Allele frequencies were 0.26 and 0.74 for G and T, respectively. The allele frequencies were in Hardy-Weinberg's equilibrium (p = 0.2443). T-ARMS-PCR assay developed in our laboratory for genotyping rs12303764 (G/T) of ULK1 gene is time saving and cost-effective as compared to the available methods. Furthermore, this is the first study reporting allelic and genotype frequencies of ULK1 rs12303764 (G/T) variants in North Indian population.

Isolation of RNA from milk somatic cells as an alternative to biopsies of mammary tissue for nutrigenomic studies in dairy ewes.

Nutrigenomic studies of mammary lipogenesis in ruminants often rely on the use of mammary tissue (MT) collected either by biopsy or at slaughter. However, isolating RNA from milk would be a useful and cost-effective technique that may avoid distress to the animal and facilitate the collection of samples in time series experiments. This assay was therefore conducted to test the hypothesis that RNA extracted from milk somatic cells (MSC) in dairy sheep would be a feasible alternative to the performance of MT biopsies for nutrigenomic analyses. To meet this objective, 8 lactating Assaf ewes were divided in 2 groups and offered a total mixed ration without supplementation (control) or supplemented with 2.4% dry matter of fish oil, which was known not only to elicit milk fat depression but also to downregulate the expression of some candidate genes involved in mammary lipogenesis. Total RNA was extracted from MSC and biopsied MT to examine whether the potential changes in the abundance of transcripts was similarly detected with both RNA sources. Milk fatty acid profile was also analyzed by gas chromatography, and variations in mRNA abundance were determined by reverse transcription quantitative PCR. Values of RNA integrity number were always ≥7.7. The expected and designed decrease of milk fat concentration with fish oil (-29%), was associated with a lower transcript abundance of genes coding for enzymes involved in fatty acid activation (ACSS1), de novo synthesis (ACACA and FASN), uptake from plasma lipids (LPL), and esterification of fatty acids to glycerol (LPIN1), as well as of a transcription factor that may regulate their expression (INSIG1). Stable mRNA levels were showed in other candidate genes, such as FABP3, GPAT4, or SCD. Changes due to the dietary treatment were similarly detected with both RNA sources (MSC and MT biopsies), which supports the initial hypothesis and would validate the use of milk as an alternative RNA source for nutrigenomic analyses in dairy sheep.

In silico Structural characterization of podocin and assessment of nephrotic syndrome-associated podocin mutants.

Nephrotic syndrome (NS) is manifested by hyperproteinuria, hypoalbuminemia, and edema. NPHS2 that encodes podocin was found to have most mutations among the genes that are involved in the pathophysiology of NS. Podocin, an integral membrane protein belonging to stomatin family, is expressed exclusively in podocytes and is localized to slit-diaphragm (SD). Mutations in podocin are known to be associated with steroid-resistant NS and rapid progression to end-stage renal disease, thus signifying its role in maintaining SD integrity and podocyte function. The structural insights of podocin are not known, and the precise mechanism by which podocin contributes to the architecture of SD is yet to be elucidated. In this study, we deduced a model for human podocin, discussed the details of transmembrane localization and intrinsically unstructured regions, and provide an understanding of how podocin interacts with other SD components. Intraprotein interactions were assessed in wild-type podocin and in some of its mutants that are associated with idiopathic NS. Mutations in podocin alter the innate intraprotein interactions affecting the native structure of podocin and its ability to form critical complex with subpodocyte proteins. © 2016 IUBMB Life, 68(7):578-588, 2016.

Quantitative assessment of the association between GAK rs1564282 C/T polymorphism and the risk of Parkinson's disease.

The aim of this meta-analysis was to evaluate the association between cyclin G-associated kinase (GAK) rs1564282 C/T polymorphism and Parkinson's disease (PD) susceptibility. GAK modifies α-synuclein expression levels and affects susceptibility to PD. Genetic variation in GAK may influence the risk of occurrence and progression of PD. Many studies have evaluated the association between GAK rs1564282 C/T polymorphism and the risk of PD. However, published data are still controversial. Nine case-control studies with a total of 8159 PD patients and 12,747 controls were included in the meta-analysis. The summary odds ratio with 95% confidence interval was calculated to estimate this association. Both the minor allele frequencies and the genotype distributions of rs1564282 within GAK were different between the two groups when all studies were pooled. Subgroup analysis by ethnicity showed GAK rs1564282 C/T polymorphism was significantly associated with increased risk in both Asian and Caucasian populations. This meta-analysis suggests that GAK rs1564282 C/T polymorphism is associated with increased susceptibility to PD.

Response to cyclosporine in steroid-resistant nephrotic syndrome: discontinuation is possible.

BACKGROUND: Steroid-resistant nephrotic syndrome (SRNS) is still regarded as a serious disease although treatment with cyclosporine (CSA) has improved outcome. However, the duration of treatment in responders is unclear, and treatment of patients with genetic causes is a matter of debate. METHODS: Thirty-six patients with SRNS were studied retrospectively. Median age at presentation was 3.2 (range, 0.06-15.0) and median follow-up 15.5 years (range, 1.8-27.7), respectively; 23 (64%) had focal segmental glomerulosclerosis (FSGS) on biopsy. In 33/36 patients (92%), genetic testing was performed for at least three most common genes known to be mutated in SRNS. RESULTS: Nineteen patients (53%), especially those with minimal change nephrotic syndrome (MCNS) at initial biopsy (p < 0.002), entered complete remission with CSA monotherapy, including one patient with compound heterozygous NPHS1 and dominant ACTN4 mutation, respectively. Ten patients entered partial remission (28%, all FSGS), including two with NPHS2 mutations. Seven patients (six FSGS, one MCNS) did not respond to treatment. In 15 of 19 responders to CSA, treatment was stopped after a median of 3.1 years (range, 0.5-14) and no further relapses occurred in 11/15 (73%) patients with median follow-up of 9.7 years. CONCLUSIONS: CSA monotherapy is effective in SRNS. Discontinuation of CSA is possible in many patients with complete remission.

Detecting copy-number variations in whole-exome sequencing data using the eXome Hidden Markov Model: an 'exome-first' approach.

Whole-exome sequencing (WES) is becoming a standard tool for detecting nucleotide changes, and determining whether WES data can be used for the detection of copy-number variations (CNVs) is of interest. To date, several algorithms have been developed for such analyses, although verification is needed to establish if they fit well for the appropriate purpose, depending on the characteristics of each algorithm. Here, we performed WES CNV analysis using the eXome Hidden Markov Model (XHMM). We validated its performance using 27 rare CNVs previously identified by microarray as positive controls, finding that the detection rate was 59%, or higher (89%) with three or more targets. XHMM can be effectively used, especially for the detection of >200 kb CNVs. XHMM may be useful for deletion breakpoint detection. Next, we applied XHMM to genetically unsolved patients, demonstrating successful identification of pathogenic CNVs: 1.5-1.9-Mb deletions involving NSD1 in patients with unknown overgrowth syndrome leading to the diagnosis of Sotos syndrome, and 6.4-Mb duplication involving MECP2 in affected brothers with late-onset spasm and progressive cerebral/cerebellar atrophy confirming the clinical suspect of MECP2 duplication syndrome. The possibility of an 'exome-first' approach for clinical genetic investigation may be considered to save the cost of multiple investigations.

A cost savings approach to SPRED1 mutational analysis in individuals at risk for neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is a clinically diagnosed autosomal dominant disorder requiring routine clinical management, particularly during the pediatric years. An overlapping disorder, Legius syndrome, at times is clinically indistinguishable from NF1 and results in a small percentage of individuals being mischaracterized. Distinguishing these two entities is increasingly important for prognosis, reproductive planning, and clinical management. The goal of our study was to evaluate the cost impact of genetic testing for patients with solely pigmentary findings. The costs of genetic testing in patients aged 1.5-18 years were modeled using a simulated population, assuming the clinical management approach of a single NF1 clinic. Two genetic testing algorithms (SPRED1 testing alone, and NF1 mutation analysis with reflex to SPRED1) were compared against a baseline of no genetic testing. The cost for SPRED1 mutation analysis for each individual meeting NF1 diagnostic criteria without neoplastic or boney manifestation, when compared to the no-testing approach with routine follow-up mutations between the ages of 10 and 14 years, was minimal (range of $4-$16). Based on the clinical practice of one NF1 clinic, we found that the cost difference to perform SPRED1 mutation analysis on individuals who meet diagnostic criteria for NF1 without neoplastic or boney manifestation were minimal. Therefore it is important that "when to test decisions" remain a physician/patient discussion, as individual benefits may be greatest at a different age than when it is most cost efficient.

Monovalent and multivalent ligation of the B cell receptor exhibit differential dependence upon Syk and Src family kinases.

The Src and Syk families of kinases are two distinct sets of kinases that play critical roles in initiating membrane-proximal B cell receptor (BCR) signaling. However, unlike in other lymphocytes, such as T cells, the "division of labor" between Src family kinases (SFKs) and Syk in B cells is not well separated because both Syk and SFKs can phosphorylate immunoreceptor tyrosine-based activation motifs (ITAMs) present in proteins comprising the BCR. To understand why B cells require both SFKs and Syk for activation, we investigated the roles of both families of kinases in BCR signaling with computational modeling and in vitro experiments. Our computational model suggested that positive feedback enabled Syk to substantially compensate for the absence of SFKs when spatial clustering of BCRs was induced by multimeric ligands. We confirmed this prediction experimentally. In contrast, when B cells were stimulated by monomeric ligands that failed to produce BCR clustering, both Syk and SFKs were required for complete and rapid BCR activation. Our data suggest that SFKs could play a pivotal role in increasing BCR sensitivity to monomeric antigens of pathogens and in mediating a rapid response to soluble multimeric antigens of pathogens that can induce spatial BCR clustering.

The role of memory-related gene polymorphisms, KIBRA and CLSTN2, on replicate memory assessment in the elderly.

The role of the CLSTN2 (rs6439886) and KIBRA (rs17070145) SNPs in cognitive impairment was analysed in a 75-76 years old group. Various memory assessment tests were carried out on individuals at baseline and during follow-up investigations, and biallelic genotyping was performed. No influence of the allele status of either SNPs was observed on any memory test. No increased risk of any type of late development, and cognitive impairment was associated with rs6439886 or rs17070145.