Association between a common missense variant in LOXL3 gene and the risk of non-syndromic cleft palate

To investigate possible association between functional common variants in the lysyl oxidase like 3 gene and non-syndromic cleft palate we selected a common missense variant p.Ile615Phe (rs17010021), which was predicted to have a probably damaging effect on the lysyl oxidase like 3 enzyme. We genotyped 258 non-syndromic cleft palate case-parent triads of European origin and tested genetic association using the transmission disequilibrium test and log-linear regression analyses of genotypic relative risks and of parent-of-origin effects. The observed genotype frequency in parents was in Hardy-Weinberg equilibrium. Compared with wild-type Ile/Ile homozygotes, the relative risks for Phe/Phe homozygote infants was 6.87 (P value 3.0 × 10-3 ), while that for Ile/Phe heterozygotes was not significant. Assuming an autosomal recessive model, the relative risks for Phe/Phe genotype resulted 10.54 (P value 2.9 × 10-5 ), with a 3.6% population attributable risk. No parent-of-origin effect was observed. The identification in lysyl oxidase like 3 of a missense variant which under a recessive model associates with 10-fold increased risk of non-syndromic cleft palate supports the hypothesis that the genetic etiology of this congenital anomaly includes relatively uncommon recessive variants with moderate penetrance and located in genes which are also involved in syndromes that include cleft palate as part of the phenotype. Our findings require functional validation and replication in a larger independent genetic association study

LINE-1 methylation in cleft lip tissues:influence of infant MTHFR c.677C>T genotype

Objective: To investigate the influence of MTHFR c.677C>T genotype on LINE-1 methylation in lateral and medial tissues from cleft lip (CL). Methods: Forty-five consecutive non-syndromic cleft lip with or without cleft palate (nsCL/P) cases were included in the study. Genomic DNA was extracted from tissues at both sides of cleft lip, and LINE-1 methylation was detected by bisulfite conversion and pyrosequencing. MTHFR c.677C>T genotyping was carried out using the TaqMan genotyping assay. Results: LINE-1 methylation level was significantly higher on medial side of cleft lip compared with lateral side (p = 0.001). This difference was not significantly influenced by the case's sex or cleft type. However, MTHFR c.677C>T genotyping revealed that the difference in LINE-1 methylation across cleft lip was restricted to carriers of C allele of MTHFR c.677C>T and was not apparent in TT homozygous cases (p = 0.027). Conclusion: This integrated analysis supports the previous finding of differences in DNA methylation across the two sides of cleft lip and further suggests a possible role of MTHFR c.677C>T genotype in establishing this difference

Meta-analysis Reveals Genome-Wide Significance at 15q13 for Nonsyndromic Clefting of Both the Lip and the Palate, and Functional Analyses Implicate GREM1 As a Plausible Causative Gene

Nonsyndromic orofacial clefts are common birth defects with multifactorial etiology. The most common type is cleft lip, which occurs with or without cleft palate (nsCLP and nsCLO, respectively). Although genetic components play an important role in nsCLP, the genetic factors that predispose to palate involvement are largely unknown. In this study, we carried out a meta-analysis on genetic and clinical data from three large cohorts and identified strong association between a region on chromosome 15q13 and nsCLP (P = 8.13×10−14 for rs1258763; relative risk (RR): 1.46, 95% confidence interval (CI): 1.32–1.61)) but not nsCLO (P = 0.27; RR: 1.09 (0.94–1.27)). The 5 kb region of strongest association maps downstream of Gremlin-1 (GREM1), which encodes a secreted antagonist of the BMP4 pathway. We show during mouse embryogenesis, Grem1 is expressed in the developing lip and soft palate but not in the hard palate. This is consistent with genotype-phenotype correlations between rs1258763 and a specific nsCLP subphenotype, since a more than two-fold increase in risk was observed in patients displaying clefts of both the lip and soft palate but who had an intact hard palate (RR: 3.76, CI: 1.47–9.61, Pdiff<0.05). While we did not find lip or palate defects in Grem1-deficient mice, wild type embryonic palatal shelves developed divergent shapes when cultured in the presence of ectopic Grem1 protein (P = 0.0014). The present study identified a non-coding region at 15q13 as the second, genome-wide significant locus specific for nsCLP, after 13q31. Moreover, our data suggest that the closely located GREM1 gene contributes to a rare clinical nsCLP entity. This entity specifically involves abnormalities of the lip and soft palate, which develop at different time-points and in separate anatomical regions.Clefts of the lip and palate are common birth defects, and require long-term multidisciplinary management. Their etiology involves genetic factors and environmental influences and/or a combination of both, however, these interactions are poorly defined. Moreover, although clefts of the lip may or may not involve the palate, the determinants predisposing to specific subphenotypes are largely unknown. Here we demonstrate that variations in the non-coding region near the GREM1 gene show a highly significant association with a particular phenotype in which cleft lip and cleft palate co-occ

Chromogenic in situ hybridization to detect EGFR gene copy number in cell blocks from fine-needle aspirates of non small cell lung carcinomas and lung metastases from colo-rectal cancer

<p>Abstract</p> <p>Background</p> <p>Several studies demonstrated that epidermal growth factor receptor (EGFR) gene copy number (GCN) correlates to the response to tyrosine kinase inhibitors in non small cell lung cancer (NSCLC) and to anti-EGFR monoclonal antibodies (MoAbs) in metastatic colorectal cancer (CRC). In the presence of lung nodules, cytology is often the only possible diagnostic approach. Chromogenic <it>in situ </it>hybridization (CISH) is an alternative technique to fluorescence <it>in situ </it>hybridization (FISH), but its feasibility in detecting EGFR GCN in cell blocks from fine-needle aspiration cytology (FNAC) of lung nodules has not yet been established.</p> <p>Methods</p> <p>We evaluated the feasibility of CISH on 33 FNAC from 20 primary NSCLC (5 squamous carcinomas, 8 large cell carcinomas and 7 adenocarcinomas) and 13 lung metastases from CRC.</p> <p>Results</p> <p>Of the 33 FNAC analyzed by CISH, 27 (82%) presented a balanced increase in EGFR gene and chromosome 7 number: 10 cases (30%) showed a low polysomy, 15 (45%) a high polysomy and 2 (6%) NSCLC were amplified. No significant differences between NSCLC and CRC lung metastases were found in relation to disomic or polysomic status. In addition, no correlation between EGFR GCN and EGFR immunohistochemical overexpression was found. Furthermore, we compared CISH results with those obtained by FISH on the same samples and we found 97% overall agreement between the two assays (k = 0.78, p < 0.0001). Two cases were amplified with both assays, whereas 1 case of NSCLC was amplified by FISH only. CISH sensitivity was 67%, the specificity and positive predictive value (PPV) was 100%, and the negative predictive value (NPV) was 97%.</p> <p>Conclusions</p> <p>Our study shows that CISH is a valid method to detect EGFR GCN in cell blocks from FNAC of primary NSCLC or metastatic CRC to the lung.</p

Périgord black truffle genome uncovers evolutionary origins and mechanisms of symbiosis

LetterInternational audienceThe Périgord black truffle ($Tuber\ melanosporum$ Vittad.) and the Piedmont white truffle dominate today's truffle market. The hypogeous fruiting body of $T.\ melanosporum$ is a gastronomic delicacy produced by an ectomycorrhizal symbiont endemic to calcareous soils in southern Europe. The worldwide demand for this truffle has fuelled intense efforts at cultivation. Identification of processes that condition and trigger fruit body and symbiosis formation, ultimately leading to efficient crop production, will be facilitated by a thorough analysis of truffle genomic traits. In the ectomycorrhizal $Laccaria\ bicolor$, the expansion of gene families may have acted as a 'symbiosis toolbox'. This feature may however reflect evolution of this particular taxon and not a general trait shared by all ectomycorrhizal species. To get a better understanding of the biology and evolution of the ectomycorrhizal symbiosis, we report here the sequence of the haploid genome of $T.\ melanosporum$, which at $\sim$125 megabases is the largest and most complex fungal genome sequenced so far. This expansion results from a proliferation of transposable elements accounting for $\sim$58% of the genome. In contrast, this genome only contains $\sim$7,500 protein-coding genes with very rare multigene families. It lacks large sets of carbohydrate cleaving enzymes, but a few of them involved in degradation of plant cell walls are induced in symbiotic tissues. The latter feature and the upregulation of genes encoding for lipases and multicopper oxidases suggest that $T.\ melanosporum$ degrades its host cell walls during colonization. Symbiosis induces an increased expression of carbohydrate and amino acid transporters in both $L.\ bicolor$ and $T.\ melanosporum$, but the comparison of genomic traits in the two ectomycorrhizal fungi showed that genetic predispositions for symbiosis $-$'the symbiosis toolbox'$-$ evolved along different ways in ascomycetes and basidiomycete

The Imaging X-ray Polarimetry Explorer (IXPE): Technical Overview

The Imaging X-ray Polarimetry Explorer (IXPE) will expand the information space for study of cosmic sources, by adding linear polarization to the properties (time, energy, and position) observed in x-ray astronomy. Selected in 2017 January as a NASA Astrophysics Small Explorer (SMEX) mission, IXPE will be launched into an equatorial orbit in 2021. The IXPE mission will provide scientifically meaningful measurements of the x-ray polarization of a few dozen sources in the 2-8 keV band, including polarization maps of several x-ray-bright extended sources and phase-resolved polarimetry of many bright pulsating x-ray sources

Identification of signalling components in tyrosine kinase cascades using phosphopeptide affinity chromatography

Various methods are now available to identify the molecular partners of the component of a signal transduction pathway. Some interactions, however, can be technically difficult to detect because they depend upon transient tyrosine phosphorylation. Here, we present a simple affinity chromatography approach based on synthetic phosphopeptides to purify potential partners of phosphotyrosine-containing proteins. With this approach, we confirm the previously characterized interaction between Grb2 and the EGF receptor, and we identify novel partners of the IGF-1 receptor and of the JAK proteins. Methenyltetrahydrofolate synthetase (MTHFS) was identified as a potential mediator of IGF-1R dependent transformation. P85alpha, the regulatory subunit of PI3 kinase, was identified as one of four proteins recruited by a phosphopeptide mimicking a motif conserved in all JAK family members

Characterization of an antibody that can detect an activated IGF-I receptor in human cancers

The type 1 insulin-like growth factor receptor (IGF-IR) plays an important role in malignant transformation and in apoptosis. Its role in human cancer has now been firmly established. IGF-IR signaling occurs only when the receptor is activated by its ligands, which induce autophosphorylation of the receptor at several tyrosine residues. Although the IGF-IR (phosphorylated or not) can be detected in human cancers with conventional antibodies, it would be desirable to obtain antibodies that can detect the IGF-IR only when activated by its ligands. We describe and characterize in this paper such an antibody and show that it can be used in sections of human cancers to detect an autophosphorylated IGF-IR. This antibody will be useful in detecting autocrine or paracrine influences on normal and tumor cells and could eventually be also useful in diagnostic and prognostic studies of human primary and metastatic cancer

The human multidrug resistance gene (MDR-1): immunocytochemical detection of its expression in oral SCC.

A large number of oral cancer patients show poor or partial response to chemotherapy and the mechanisms are poorly understood. At present, an MDR-1 product, the P-170 glycoprotein, is the best known of the P-170 family and is involved in resistance to natural product-based chemotherapeutics, including taxanes, anthracyclines, vinca alkaloids, podophyllotoxins and camptothecins. Although several reports suggest that P-170 is clinically relevant in haematological malignancies, its role in solid tumours is not well understood. Its overexpression has been found to be correlated with the poor outcome observed in patients treated with chemotherapy and presenting drug resistance. The aim of this study was to detect the protein expression patterns of MDR-1 product by immunohistochemistry in formalin-fixed-paraffin-embedded tissues. For these reasons, 30 oral SCC and 6 healthy oral mucosa specimens were tested with anti-P-170 antibodies using standard streptavidin-biotin-peroxide technique. Immunohistochemistry demonstrated that 4 cases (66.6%) of normal oral mucosa and 24 cases (80%) of oral SCC showed positivity. Four cases (13.4%) showed strong positivity in tumour areas and complete negativity in normal epithelial cells adjacent to the tumour. No staining was observed in stromal structures, with the exception of the lymphocytic compartment that showed a strong staining as reported in literature for CD56+ and CD8+ cells. Four G1 tumours (33%) and 2 G3 tumour (33%) showed strong positivity in areas with a higher degree of differentiation. P-170 positivity in normal epithelial cells of smoker patients, in differentiated area of neoplasia and negativity or zonal positivity in undifferentiated area of tumour suggested that activation of the MDR-1 gene or selection of intrinsically multidrug resistance neoplastic cells may occur at early stages of tumorigenesis of oral cancers, before the real evidence of cellular transformation. Thus the contact with possible chemical carcinogens, such as those of tobacco smoke, may induce activation of MDR-1 gene. This study was conducted only on untreated carcinomas so for this reason it cannot indicate the real incidence of acquired multidrug resistance. The data of MDR-1 product expression by immunohistochemistry in oral SCC might suggest that an overexpression of this protein could constitute a hallmark of potential more aggressive phenotype for this type of neoplasia and a rapid method for pre-screening tumours for a constitutive multidrug resistance in order to orientate the cancer treatment