Novel mutations of FOXC1 and PITX2 in patients with Axenfeld-Rieger malformations

Molecular Genetics Laboratory, University Eye Hospital, Tübingen, Germany.
Investigative Ophthalmology & Visual Science (Impact Factor: 3.4). 10/2006; 47(9):3846-52. DOI: 10.1167/iovs.06-0343
Source: PubMed


To determine the prevalence of FOXC1 and PITX2 mutations and to assess clinical phenotypes in a cohort of German patients with Axenfeld-Rieger malformations.
All coding exons of the FOXC1 and PITX2 genes were amplified by PCR from genomic DNA and subjected to direct DNA sequencing. Analysis of mutations in control subjects was performed by restriction fragment length polymorphism (RFLP) analysis.
Sequence variants were identified by DNA sequencing in 15 of 19 cases. Mutation screening identified four potentially pathogenic FOXC1 mutations causing amino acid substitutions (P79R, Y115S, G149D, and M161V) that were not present in 100 control subjects. In addition, two different 1-bp deletions causing a frameshift and subsequent premature stop codon were identified in two subjects. One patient harbored a FOXC1 nonsense mutation (S48X). Mutation screening also identified two potentially pathogenic PITX2 mutations (P64L and P64R) in two index patients that were excluded in 100 healthy control subjects.
The findings in the present study clearly demonstrate that FOXC1 and PITX2 mutations are responsible for a significant proportion of Axenfeld-Rieger malformations in Germany.

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    • "The zebrafish foxc1a gene encodes a 476 amino acid protein, which is 63% identical to the human and mouse FoxC1 homologs. FoxC1 proteins belong to the forkhead family of transcription factors characterized by the forkhead domain consisting of two DNA-binding wing helix domains, crucial for FoxC1 function (Murphy et al., 2004; Nishimura et al., 2001; Saleem et al., 2004; Weisschuh et al., 2006). The p162 mutation W118* truncates the open reading frame in the forkhead domain (amino acids 72-163). "
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    ABSTRACT: Neural connectivity between the spinal cord and paired appendages is key to the superior locomotion of tetrapods and aquatic vertebrates. In contrast to nerves that innervate axial muscles, those innervating appendages converge at a specialized structure, the plexus, where they topographically reorganize before navigating towards their muscle targets. Despite its importance for providing appendage mobility, the genetic program that drives nerve convergence at the plexus, as well as the functional role of this convergence, are not well understood. Here, we show that in zebrafish the transcription factor foxc1a is dispensable for trunk motor nerve guidance but is required to guide spinal nerves innervating the pectoral fins, equivalent to the tetrapod forelimbs. In foxc1a null mutants, instead of converging with other nerves at the plexus, pectoral fin nerves frequently bypass the plexus. We demonstrate that foxc1a expression in muscle cells delineating the nerve path between the spinal cord and the plexus region restores convergence at the plexus. By labeling individual fin nerves, we show that mutant nerves bypassing the plexus enter the fin at ectopic positions, yet innervate their designated target areas, suggesting that motor axons can select their appropriate fin target area independently of their migration through the plexus. Although foxc1a mutants display topographically correct fin innervation, mutant fin muscles exhibit a reduction in the levels of pre- and postsynaptic structures, concomitant with reduced pectoral fin function. Combined, our results reveal foxc1a as a key player in the development of connectivity between the spinal cord and paired appendages, which is crucial for appendage mobility. © 2015. Published by The Company of Biologists Ltd.
    Full-text · Article · Feb 2015 · Development
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    • "GH deficiency (Feingold et al. 1969, Sadeghi-Nejad & Senior 1974) and short stature (Brooks et al. 1989) have also been reported in several cases, with an enlarged sella turcica and combined GH/TSH or GH/ACTH deficiency (Polomeno et al. 1980). A broad range of PITX2 mutations have been described, with missense mutations usually affecting the homeodomain and leading, in some cases, to a dominantnegative mutant form (Semina et al. 1996, Alward et al. 1998, Kulak et al. 1998, Perveen et al. 2000, Priston et al. 2001, Phillips 2002, Xia et al. 2004, Idrees et al. 2006, Kniestedt et al. 2006, Vieira et al. 2006, Weisschuh et al. 2006). Splice-site (Semina et al. 1996, Doward et al. 1999, Perveen et al. 2000, Maciolek et al. 2006), frameshift, and nonsense mutations have been found throughout the gene and may result in truncated proteins (Semina et al. 1996, Perveen et al. 2000, Priston et al. 2001, Wang et al. 2003, Brooks et al. 2004, Lines et al. 2004, Saadi et al. 2006, Vieira et al. 2006). "
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    ABSTRACT: The bicoid-like transcription factor PITX2 has been previously described to interact with the pituitary-specific POU homeodomain factor POU1F1 (human ortholog of PIT-1) to achieve cell-specific expression of prolactin (PRL) and GH in pituitary somatolactotroph cells. In this work, we have investigated the functional properties of three PITX2 mutants reported in Axenfeld-Rieger syndrome patients relative to the regulation of these genes, using reporter genes under the control of human PRL (hPRL), hGH, or POU1F1 promoters transfected in nonpituitary and pituitary cell lines. Among the three mutations studied, Y167X and E101X introduce a premature stop codon, and F104L leads to an amino acid substitution. While PITX2(E101X) is not expressed in the cells following transfection, and PITX2(F104L) is functionally inactive, the PITX2(Y167X) mutant keeps its DNA-binding capacity and displays a markedly enhanced activation of the hPRL and POU1F1 promoters, but not of the hGH promoter. Y167X is the first mutation of PITX2 described to result in a differential effect on the activation of its different physiological targets, hPRL and POU1F1 on one hand and hGH on the other hand. The differential effect of the Y167X mutation might be linked to an interaction of PITX2 with different transcription factors or cofactors when bound to the hPRL and POU1F1 or the hGH promoters. These results might form the basis for the identification of the PITX2 protein complex necessary for the differential GH or PRL expression.
    Full-text · Article · Oct 2010 · Journal of Molecular Endocrinology
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    • "Pitx2 encodes a homeodomain transcription factor which plays an essential role during ocular anterior segment patterning and development (Gage et al., 1999; Hjalt et al., 2000; Gage et al., 2005). Heterozygous mutations in human PITX2 results in Axenfeld-Rieger syndrome characterized by anterior segment dysgenesis and high risk of developing glaucoma (Semina et al., 1996; Mears et al., 1998; Kozlowski and Walter, 2000; Vieira et al., 2006; Weisschuh et al., 2006). Global and neural crest specific Pitx2 knockout mice exhibit a similar eye phenotype including abnormal anterior segment differentiation, vasculogenesis, eyelid defects and coloboma (Gage et al., 1999; Kitamura et al., 1999; Lu et al., 1999; Evans and Gage, 2005). "
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    ABSTRACT: Morphogenesis during eye development requires retinoic acid (RA) receptors plus RA-synthesizing enzymes, and loss of RA signaling leads to ocular disorders associated with loss of Pitx2 expression in perioptic mesenchyme. Several Wnt signaling components are expressed in ocular tissues during eye development including Dkk2, encoding an inhibitor of Wnt/beta-catenin signaling, which was previously shown to be induced by Pitx2 in the perioptic mesenchyme. Here, we investigated potential cross-talk between RA and Wnt signaling during ocular development. Genetic studies using Raldh1/Raldh3 double null mice deficient for ocular RA synthesis demonstrated that Pitx2 and Dkk2 were both down-regulated in perioptic mesenchyme. Chromatin immunoprecipitation and gel mobility shift studies demonstrated the existence of a DR5 RA response element upstream of Pitx2 that binds all three RA receptors in embryonic eye. Axin2, an endogenous readout of Wnt/beta-catenin signaling, was up-regulated in cornea and perioptic mesenchyme of RA deficient embryos. Also, expression of Wnt5a was expanded in perioptic mesenchyme of RA deficient eyes. Our findings demonstrate excessive activation of Wnt signaling in the perioptic mesenchyme of RA deficient mice which may be responsible for abnormal development leading to defective optic cup, cornea, and eyelid morphogenesis.
    Full-text · Article · Apr 2010 · Developmental Biology
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