Matera, I., Bachetti, T., Puppo, F., Di Duca, M., Morandi, F., Casiraghi, G. M. et al. PHOX2B mutations and polyalanine expansions correlate with the severity of the respiratory phenotype and associated symptoms in both congenital and late onset central hypoventilation syndrome. J. Med. Genet. 41, 373-380

University of Groningen, Groningen, Groningen, Netherlands
Journal of Medical Genetics (Impact Factor: 6.34). 06/2004; 41(5):373-80. DOI: 10.1136/jmg.2003.015412
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Available from: Francesca Puppo, Sep 26, 2015
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    • "Interestingly, co-transfection of the deleted proteins with the wild-type protein reduced promoter activity in comparison with the full transactivation observed with PHOX2B WT alone only in the case of the PHOX2B Δaa 200–290 mutant (Fig. 7D, compare hatched bars with white bar). These data are in line with the fact that in-frame polyalanine contraction variants (with 7, 13, 14 or 15 repeats in the polyalanine tract) are also found in the control population (Amiel et al., 2003; Hung et al., 2007; Matera et al., 2004; Toyota et al., 2004). Finally, our data support the idea that the portion of the protein downstream of the homeodomain containing the polyalanine tract modulates the DNA binding and transactivation function of the PHOX2B protein, and it is reasonable to hypothesize that the deletion of a large C-terminal region of the protein encompassing the polyalanine stretch can affect the correct orientation, folding and/or protein–protein interaction properties of the remaining portion of the protein generating a mutant variant with dominant-negative effects on a subset of promoters. "
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    ABSTRACT: The PHOX2B transcription factor plays a crucial role in autonomic nervous system development. In humans, heterozygous mutations of the PHOX2B gene lead to Congenital Central Hypoventilation Syndrome (CCHS), a rare disorder characterized by a broad variety of symptoms of autonomic nervous system dysfunction including inadequate control of breathing. The vast majority of patients with CCHS are heterozygous for a polyalanine repeat expansion mutation involving a polyalanine tract of twenty residues in the C-terminus of PHOX2B. Although several lines of evidence support a dominant-negative mechanism for PHOX2B mutations in CCHS, the molecular effects of PHOX2B mutant proteins on the transcriptional activity of the wild-type protein have not yet been elucidated. As one of the targets of PHOX2B is the PHOX2B gene itself, we tested the transcriptional activity of wild-type and mutant proteins on the PHOX2B gene promoter, and found that the transactivation ability of proteins with polyalanine expansions decreased as a function of the length of the expansion, whereas DNA binding was severely affected only in the case of the mutant with the longest polyalanine tract (+13 alanine). Co-transfection experiments using equimolar amounts of PHOX2B wild-type and mutant proteins in order to simulate a heterozygous state in vitro and four different PHOX2B target gene regulatory regions (PHOX2B, PHOX2A, DBH, TLX2) clearly showed that the polyalanine expanded proteins alter the transcriptional activity of wild-type protein in a promoter-specific manner, without any clear correlation with the length of the expansion. Moreover, although reduced transactivation may be caused by retention of the wild-type protein in the cytoplasm or in nuclear aggregates, this mechanism can only be partially responsible for the pathogenesis of CCHS because of the reduction in cytoplasmic and nuclear accumulation when the +13 alanine mutant is co-expressed with wild-type protein, and the fact that the shortest polyalanine expansions do not form visible cytoplasmic aggregates. Deletion of the C-terminal of PHOX2B leads to a protein that correctly localizes in the nucleus but impairs PHOX2B wild-type transcriptional activity, thus suggesting that protein mislocalization is not the only mechanism leading to CCHS. The results of this study provide novel in vitro experimental evidence of a transcriptional dominant-negative effect of PHOX2B polyalanine mutant proteins on wild-type protein on two different PHOX2B target genes.
    Neurobiology of Disease 10/2012; 50(1). DOI:10.1016/j.nbd.2012.10.019 · 5.08 Impact Factor
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    • "Interestingly, genotype-phenotype correlation has been described in CCHS patients with the PHOX2B gene mutation (4). Increasing polyalanine repeat mutation size is associated with a more severe clinical phenotype (4). Trochet et al. (10) described that Haddad syndrome (CCHS + HD type) showed usually expansions of +6 and +7 alanines, and frameshift or missense PHOX2B mutations may predispose to neuroblastoma. "
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    ABSTRACT: Congenital central hypoventilation syndrome with Hirschsprung's disease, also known as Haddad syndrome, is an extremely rare disorder with variable symptoms. Recent studies described that congenital central hypoventilation syndrome had deep relation to the mutation of the PHOX2B gene in its diagnosis and phenotype. We report a newborn male infant with clinical manifestations of recurrent hypoventilation with hypercapnea and bowel obstruction. These clinical manifestations were compatible with congenital central hypoventilation syndrome and Hirschsprung's disease, and polyalanine 26 repeats in the PHOX2B gene supported the diagnosis of congenital central hypoventilation. We described a first case of Haddad syndrome in Korean and its clinical and genetic characteristics were discussed.
    Journal of Korean medical science 02/2011; 26(2):312-5. DOI:10.3346/jkms.2011.26.2.312 · 1.27 Impact Factor
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    • "Physiologic symptoms can include diminished pupillary response, esophageal dysmotility, profound constipation even in the absence of HSCR, breath-holding spells, reduced basal body temperature control, sporadic profuse diaphoresis, lack of perception to dyspnea, altered perception of anxiety and pain, and lack of physiologic responsiveness to the challenges of exercise and environmental stressors (Faure et al., 2002; Goldberg and Ludwig, 1996; Marazita et al., 2001; O'Brien et al., 2005; Paton et al., 1993; Pine et al., 1994; Shea et al., 1993a,b,c; Silvestri et al., 1995; Spengler et al., 1998; Trang et al., 2005; Weese- Mayer et al., 1992, 2001, 2010). Thus, a constellation of features is emerging that defines the PHOX2B-genotype/CCHS-phenotype correlation, including severity of facial dysmorphology, ventilatory dependence, symptoms of autonomic dysregulation, cardiac asystoles , relation to age at presentation, HSCR, and neural crest tumors (Berry-Kravis et al., 2006; Gronli et al., 2008; Matera et al., 2004; Repetto et al., 2009; Todd et al., 2006; Trang et al., 2004; Trochet et al., 2005; Weese-Mayer et al., 2003, 2005). A subgroup of PHOX2B mutation-confirmed CCHS cases have recently been identified outside the newborn period (Weese-Mayer et al., 2009, 2010). "
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    ABSTRACT: The paired-like homeobox 2B gene (PHOX2B) is the disease-defining gene for congenital central hypoventilation syndrome (CCHS). Individuals with CCHS typically present in the newborn period with alveolar hypoventilation during sleep and often during wakefulness, altered respiratory control including reduced or absent ventilatory responses to hypercarbia and hypoxemia, and autonomic nervous system (ANS) dysregulation; however, a subset of individuals present well into adulthood. Thermoregulation is altered and perception of shortness of breath is absent, but voluntary breathing is retained. Structural and functional magnetic resonance imaging (MRI) and limited post-mortem studies in subjects with CCHS reveal abnormalities in both forebrain and brainstem. MRI changes appear in the hypothalamus (responsible for thermal drive to breathing), posterior thalamus and midbrain (mediating O(2) and oscillatory motor patterns), caudal raphé and locus coeruleus (regulating serotonergic and noradrenergic systems), the lateral medulla, parabrachial pons, and cerebellum (coordinating chemoreceptor and somatic afferent activity with breathing), and insular and cingulate cortices (mediating shortness of breath perception). Structural and functional alterations in these sites may result from PHOX2B mutations or be secondary to hypoxia/perfusion alterations from suboptimal management/compliance. The study of CCHS, with collaboration between physician-scientists and basic scientists, offers a rare opportunity to investigate control of breathing within the complex physiological network of the ANS.
    Respiratory Physiology & Neurobiology 10/2010; 173(3):322-35. DOI:10.1016/j.resp.2010.06.013 · 1.97 Impact Factor
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