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Three common diseases, isolated cleft lip and cleft palate (CLP), hypothyroidism and thyroid cancer all map to the FOXE1 locus, but causative variants have yet to be identified. In patients with CLP, the frequency of coding mutations in FOXE1 fails to account for the risk attributable to this locus, suggesting that the common risk alleles reside in nearby regulatory elements. Using a combination of zebrafish and mouse transgenesis, we screened 15 conserved non-coding sequences for enhancer activity, identifying three that regulate expression in a tissue specific pattern consistent with endogenous foxe1 expression. These three, located -82.4, -67.7 and +22.6 kb from the FOXE1 start codon, are all active in the oral epithelium or branchial arches. The -67.7 and +22.6 kb elements are also active in the developing heart, and the -67.7 kb element uniquely directs expression in the developing thyroid. Within the -67.7 kb element is the SNP rs7850258 that is associated with all three diseases. Quantitative reporter assays in oral epithelial and thyroid cell lines show that the rs7850258 allele (G) associated with CLP and hypothyroidism has significantly greater enhancer activity than the allele associated with thyroid cancer (A). Moreover, consistent with predicted transcription factor binding differences, the -67.7 kb element containing rs7850258 allele G is significantly more responsive to both MYC and ARNT than allele A. By demonstrating that this common non-coding variant alters FOXE1 expression, we have identified at least in part the functional basis for the genetic risk of these seemingly disparate disorders.

MSX1 is one of the homeoproteins with the homeodomain (HD) sequence, which regulates proliferation and differentiation of mesenchymal cells. In this study, we investigated the nuclear localization signal (NLS) in the MSX1 HD by deletion and amino acid substitution analyses. The web-based tool NLStradamus predicted 2 putative basic motifs in the N- and C-termini of the MSX1 HD. Green fluorescent protein (GFP) chimera studies revealed that NLS1 ([.sub.161][RKHKTNRKPR.sub.170]) and NLS2 ([.sub.216][NRRAKAKR.sub.223]) were independently insufficient for robust nuclear localization. However, they can work cooperatively to promote nuclear localization of MSX1, as was shown by the 2 tandem NLS motifs partially restoring functional NLS, leading to a significant nuclear accumulation of the GFP chimera. These results demonstrate a unique NLS motif in MSX1, which consists of an essential single core motif in helix-I, with weak potency, and an auxiliary subdomain in helix-III, which alone does not have nuclear localization potency. Additionally, other peptide sequences, other than predicted 2 motifs in the spacer, may be necessary for complete nuclear localization in MSX1 HD. Key words: MSX1, homeodomain (HD), nuclear localization signal (NLS). MSX1 est une des homeoproteines a sequence homeodomaine (HD) qui regule la proliferation et la differenciation des cellules mesenchymateuses. Dans cette etude, les auteurs ont examine le signal de localisation nucleaire (SLN) de l'HD de MSX1 au moyen d'analyses de deletion et de substitution d'acides amines. L'outil NLStradamus disponible sur le web predisait deux motifs basiques presumes aux extremites N- et C-terminales de l'HD de MSX1. Des etudes realisees avec des chimeres comportant la proteine fluorescente verte GFP ont revele que SLN1 ([.sub.161][RKHKTNRKPR.sub.170]) and NLS2 ([.sub.216][NRRAKAKR.sub.223]) etaient insuffisants independamment pour assurer une localisation nucleaire forte. Cependant, ils peuvent travailler de maniere cooperative pour favoriser la localisation nucleaire de MSX1, comme le demontrent les deux motifs SLN en tandem qui retablissaient partiellement un SLN fonctionnel, conduisant a une accumulation nucleaire significative des chimeres GFP. Ces resultats demontrent qu'il existe un motif SLN unique chez MSX1, qui consiste en un motif cceur unique essentiel dans l'helice-I, a faible potentiel, et un sous-domaine auxiliaire dans l'helice-III qui, seul, ne possede pas d'activite de localisation nucleaire. En outre, d'autres sequences peptidiques differentes des deux motifs predits dans l'espaceur, peuvent etre necessaires pour assurer une localisation nucleaire complete de l'HD de MSX1. [Traduit par la Redaction] Mots-cles: MSX1, homeodomaine (DH), signal de localisation nucleaire., Introduction Homeoproteins are transcription factors that regulate cell proliferation and differentiation (Ladam and Sagerstrom 2014). These proteins contain a highly conserved polypeptide, termed the homeodomain (HD). This domain enables them [...]

The study of 10 populations (comprising a total of 1968 families) with cleft lip or palate showed highly significant transmission disequilibrium for the V2741 variant of the IRF gene. The contribution of variants in single gene to cleft lip or palate is an important consideration in genetic counseling.

Nonsyndromic cleft lip with or without cleft palate (CL/P) and cleft palate only (CPO) are representative of complex genetic traits. Disease loci of CP/P and CPO can be hypothesized to be identified by a candidate-gene linkage-disequilibrium (LD) strategy. Candidate genes for clefting have been screened for LD with either CL/P or CPO in a population mainly Caucasian. Both case-control- and nuclear-family-based approaches were used. No common mutations were found in the coding regions in a mutation search in certain genes in 69 CPO patients and in a group within the larger group of CL/P patients. Several rare variants of MSX1 and TGFB3 were found. Results are a foundation for further research.

Background: We performed a randomized controlled chemoprevention trial of oral leukoplakia by administrating a low dose of beta-carotene and vitamin C supplements. 17% of subjects in the experimental arm (4/23) demonstrated clinical remission (complete or partial response) at completion of the trial. The objective of this study was to determine whether baseline expression of p53 and ki67 demonstrated any differences between those responding or not responding to our intervention. A secondary objective was to elucidate any relationship between dietary factors and clinical responses.Methods: For this biomarker study, we included all subjects in the experimental group (n = 23) who were non-smokers. Among 16 who completed the trial for 1 year of supplementation, there were four responders and 12 non-responders at 1-year follow-up. Following immuno-staining for p53 and ki67, the percentage of positive cell nuclei were analyzed as labeling index (LI).Results: Expression of p53 was greater in basal layers than in para-basal layers. Mean para-basal LI of p53 was higher in non-responding (26.0) than in responding subjects (11.2) (P = 0.028). ki67 LIs were not significantly different in the two groups.Conclusions: Expression of p53 was inversely related to clinical response to the supplements. Other biomarkers that may recognize subject's responsiveness to chemoprevention require further study. [ABSTRACT FROM AUTHOR]

Cleidocranial dysplasia (CCD; MIM 119600) is an autosomal dominant skeletal dysplasia characterised by hypopalstic and/or aplastic clavicles, midface hypoplasia, absent or delayed closure of cranial sutures, moderately short stature, delayed eruption of permanent dentition and supernumerary teeth. The molecular pathogenesis can be explained in about two-thirds of CCD patients by haploinsufficiency of the RUNX2 gene. In our current study, we identified a novel and rare variant of the RUNX2 gene (c.181_189dupGCGGCGGCT) in a Japanese patient with phenotypic features of CCD. The insertion led an alanine tripeptide expansion (+3Ala) in the polyalanine tract. To date, a RUNX2 variant with alanine decapeptide expansion (+10Ala) is the only example of a causative variant of RUNX2 with polyalanine tract expansion to be reported, whilst RUNX2 (+1Ala) has been isolated from the healthy population. Thus, precise analyses of the RUNX2 (+3Ala) variant were needed to clarify whether the tripeptide expanded RUNX2 is a second disease-causing mutant with alanine tract expansion. We therefore investigated the biochemical properties of the mutant RUNX2 (+3Ala), which contains 20 alanine residues in the polyalanine tract. When transfected in COS7 cells, RUNX2 (+3Ala) formed intracellular ubiquitinated aggregates after 24h, and exerted a dominant negative effect in vitro. At 24h after gene transfection, whereas slight reduction was observed in RUNX2 (+10Ala), all of these mutants significantly activated osteoblast-specific element-2, a cis-acting sequence in the promoter of the RUNX2 target gene osteocalcin. The aggregation growth of RUNX2 (+3Ala) was clearly lower and slower than that of RUNX2 (+10Ala). Furthermore, we investigated several other RUNX2 variants with various alanine tract lengths, and found that the threshold for aggregation may be RUNX2 (+3Ala). We conclude that RUNX2 (+3Ala) is the cause of CCD in our current case, and that the accumulation of intracellular aggregates in vitro is related to the length of the alanine tract., (© The Author 2015. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)