Expression of PROKR1 and PROKR2 in human enteric neural precursor cells and identification of sequence variants suggest a role in HSCR

Background

The enteric nervous system (ENS) is entirely derived from neural crest and its normal development is regulated by specific molecular pathways. Failure in complete ENS formation results in aganglionic gut conditions such as Hirschsprung''s disease (HSCR). Recently, PROKR1 expression has been demonstrated in mouse enteric neural crest derived cells and Prok-1 was shown to work coordinately with GDNF in the development of the ENS.

Principal Findings

In the present report, ENS progenitors were isolated and characterized from the ganglionic gut from children diagnosed with and without HSCR, and the expression of prokineticin receptors was examined. Immunocytochemical analysis of neurosphere-forming cells demonstrated that both PROKR1 and PROKR2 were present in human enteric neural crest cells. In addition, we also performed a mutational analysis of PROKR1, PROKR2, PROK1 and PROK2 genes in a cohort of HSCR patients, evaluating them for the first time as susceptibility genes for the disease. Several missense variants were detected, most of them affecting highly conserved amino acid residues of the protein and located in functional domains of both receptors, which suggests a possible deleterious effect in their biological function.

Conclusions

Our results suggest that not only PROKR1, but also PROKR2 might mediate a complementary signalling to the RET/GFRα1/GDNF pathway supporting proliferation/survival and differentiation of precursor cells during ENS development. These findings, together with the detection of sequence variants in PROKR1, PROK1 and PROKR2 genes associated to HSCR and, in some cases in combination with RET or GDNF mutations, provide the first evidence to consider them as susceptibility genes for HSCR.     

Exome-Wide Association Study Identified New Risk Loci for Hirschsprung’s Disease

Hirschsprung disease (HSCR) is a rare congenital disease caused by impaired proliferation and migration of neural crest cells. In this study, we aimed to investigate the genetic loci involved in the pathogenesis of HSCR. The exome-wide scan was performed to screen the genetic variants with minor allele frequency (MAF)?<?0.05 in exonic regions. Candidate mutation type and the wild type were overexpressed to investigate the affection on cell proliferation and migration. We found that ten variants were associated with HSCR at P?<?10^?4 in the single-variant analysis while ten genes were also associated with HSCR at P?<?10^?4 in the optimized sequence kernel association test (SKAT-O) test analysis. Among these SNPs, the missense variants catechol-O-methyltransferase (COMT) (rs6267) and armadillo repeat gene deleted in velocardiofacial syndrome (ARVCF) (rs80068543) indicated an ectopic expression in colon tissues of HSCR patients. The Ala72Ser variant in COMT induced proliferation suppression through NOTCH signal pathway, while the ARVCF affected cell migration via the downregulating of RHOA and ROC. In conclusion, this exome array study identified the COMT and ARVCF missense coding variants as candidate loci for HSCR. The finding implies the abnormal variant of COMT and ARVCF may account for the pathogenesis of HSCR.     

Molecular genetics of Hirschsprung disease: a model of multigenic neurocristopathy

Hirschsprung's disease (HSCR, aganglionic megacolon) is a frequent congenital malformation regarded as a multigenic neurocristopathy. Three susceptibility genes have been recently identified in HSCR, namely the RET proto-oncogene, the endothelin B receptor (EDNRB) gene, and the endothelin 3 (EDN3) gene. RET gene mutations were found in significant proportions of familial (50%) and sporadic (15-20%) HSCR, while homozygosity for EDNRB or EDN3 mutations accounted for the rare HSCR-Waardenburg syndrome (WS) association. More recently, heterozygous EDNRB an EDN3 missense mutations have been reported in isolated HSCR patients. Some of these results were obtained after the identification of mouse genes whose natural or site-directed mutations resulted in megacolon and coat color spotting. There is also conclusive evidence for the involvement of other independent loci in HSCR. In particular, the recent identification of neurotrophic factors acting as RET ligands (GDNF and Neurturin) provide additional candidate genes for HSCR. The dissection of the genetic etiology of HSCR disease may then provide a unique opportunity to distinguish between a polygenic and a genetically heterogeneous disease, thereby helping to understand other complex disorders and congenital malformations hitherto considered as multifactorial in origin. Finally, the study of the molecular bases of HSCR is also a step towards the understanding of developmental genetics of the enteric nervous system giving support to the role of the tyrosine kinase and endothelin-signaling pathways in the development of neural crest-derived enteric neurons in human.     

Fine Structure Mapping and Deletion Analysis of the Murine Piebald Locus

piebald (s) is a recessive mutation that affects the development of two cell types of neural crest origin: the melanocytes, responsible for pigment synthesis in the skin, and enteric ganglia, which innervate the lower bowel. As a result, mice carrying piebald mutations exhibit white spotting in the coat and aganglionic megacolon. Previously the gene had been localized to the distal half of mouse chromosome 14. To determine its precise location relative to molecular markers, an intersubspecific backcross was generated. Two anchor loci of chromosome 14, slaty and hypogonadal, in addition to simple sequence length repeat markers, were used to localize s to a 2-cM interval defined by the markers D14Mit38 and D14Mit42. The molecular markers were also used to characterize nine induced s alleles. Three of these mutations exhibited no deletions or rearrangements of the flanking markers, whereas the other six had two or more of these markers deleted. The extent of the deletions was found to be consistent with the severity of the homozygous phenotype. The location of deletion breakpoints in the induced alleles, coupled with the recombination breakpoints in the backcross progeny, provide useful molecular landmarks to define the location of the piebald gene.     

RET and NRG1 interplay in Hirschsprung disease

Hirschsprung disease (HSCR, aganglionic megacolon) is a complex genetic disorder of the enteric nervous system (ENS) characterized by the absence of enteric neurons along a variable length of the intestine. While rare variants (RVs) in the coding sequence (CDS) of several genes involved in ENS development lead to disease, the association of common variants (CVs) with HSCR has only been reported for RET (the major HSCR gene) and NRG1. Importantly, RVs in the CDS of these two genes are also associated with the disorder. To assess independent and joint effects between the different types of RET and NRG1 variants identified in HSCR patients, we used 254 Chinese sporadic HSCR patients and 143 ethnically matched controls for whom the RET and/or NRG1 variants genotypes (rare and common) were available. Four genetic risk factors were defined and interaction effects were modeled using conditional logistic regression analyses and pair-wise Kendall correlations. Our analysis revealed a joint effect of RET CVs with RET RVs, NRG1 CVs or NRG1 RVs. To assess whether the genetic interaction translated into functional interaction, mouse neural crest cells (NCCs; enteric neuron precursors) isolated from embryonic guts were treated with NRG1 (ErbB2 ligand) or/and GDNF (Ret ligand) and monitored during the subsequent neural differentiation process. Nrg1 inhibited the Gdnf-induced neuronal differentiation and Gdnf negatively regulated Nrg1-signaling by down-regulating the expression of its receptor, ErbB2. This preliminary data suggest that the balance neurogenesis/gliogenesis is critical for ENS development.     

A new role for the Endothelin-1/Endothelin-A receptor signaling during early neural crest specification

The neural crest is induced at the border of the neural plate in a multistep process by signals emanated from the epidermis, neural plate and mesoderm. In this work we show for the first time the existence of a neural crest maintenance step which is dependent on signals released from the mesoderm. We identified Endothelin-1 (Edn1) and its receptor (Ednra) as key players of this signal and we show that Edn1/Ednra signaling is required for maintenance of the neural crest by a dual mechanism of cell specification and cell survival. We show that: (i) Ednra is expressed in prospective neural crest; (ii) loss-of-function experiments with antisense morpholino or with specific chemical inhibitor suppress the expression of early neural crest markers; (iii) gain-of-function experiments expand the neural crest territory; (iv) epistatic experiments show that Ednra/Edn1 is downstream of the early neural crest gene Msx1 and upstream of the late genes Sox9 and Sox10; and (v) Edn1/Ednra signaling inhibits apoptosis and controls cell specification of the neural crest. Together, our results provide insight on a new role of Edn1/Ednra cell signaling pathway during early neural crest development.     

EDNRB/EDN3 and Hirschsprung Disease Type II

The study of vertebrate pigmentary anomalies has greatly improved our understanding of melanocyte biology. One such disorder, Waardenburg syndrome (WS), is a mendelian trait characterized by hypopigmentation and sensorineural deafness. It is commonly subdivided into four types (WS1–4), defined by the presence or absence of additional symptoms. WS type 4 (WS4), or Shah‐Waardenburg syndrome, is also known as Hirschsprung disease Type II (HSCR II) and is characterized by an absence of epidermal melanocytes and enteric ganglia. Mutations in the genes encoding the endothelin type‐B receptor (EDNRB) and its physiological ligand endothelin 3 (EDN3) are now known to account for the majority of HSCR II patients. Null mutations in the mouse genes Ednrb and Edn3 have identified a key role for this pathway in the normal development of melanocytes and other neural crest‐derived lineages. The pleiotropic effects of genes in this pathway, on melanocyte and enteric neuron development, have been clarified by the embryologic identification of their common neural crest (NC) ancestry. EDNRB and EDN3 are transiently expressed in crest‐derived melanoblast and neuroblast precursors, and in the surrounding mesenchymal cells, respectively. The influence of EDNRB‐mediated signaling on the emigration, migration, proliferation, and differentiation of melanocyte and enteric neuron precursors, in vivo and in vitro has recently been the subject of great scrutiny. A major emergent theme is that EDN3‐induced signaling prevents the premature differentiation of melanocyte and enteric nervous system precursors and is essential between 10 and 12.5 days post‐coitum. We review the present understanding of pigment cell development in the context of EDNRB/EDN3 – a receptor‐mediated pathway with pleiotropic effects.     

Endothelin regulates neural crest deployment and fate to form great vessels through Dlx5/Dlx6-independent mechanisms

Endothelin-1 (Edn1), originally identified as a vasoconstrictor peptide, is involved in the development of cranial/cardiac neural crest-derived tissues and organs. In craniofacial development, Edn1 binds to Endothelin type-A receptor (Ednra) to induce homeobox genes Dlx5/Dlx6 and determines the mandibular identity in the first pharyngeal arch. However, it remains unsolved whether this pathway is also critical for pharyngeal arch artery development to form thoracic arteries. Here, we show that the Edn1/Ednra signaling is involved in pharyngeal artery development by controlling the fate of neural crest cells through a Dlx5/Dlx6-independent mechanism. Edn1 and Ednra knock-out mice demonstrate abnormalities in pharyngeal arch artery patterning, which include persistent first and second pharyngeal arteries, resulting in additional branches from common carotid arteries. Neural crest cell labeling with Wnt1-Cre transgene and immunostaining for smooth muscle cell markers revealed that neural crest cells abnormally differentiate into smooth muscle cells at the first and second pharyngeal arteries of Ednra knock-out embryos. By contrast, Dlx5/Dlx6 knockout little affect the development of pharyngeal arch arteries and coronary arteries, the latter of which is also contributed by neural crest cells through an Edn-dependent mechanism. These findings indicate that the Edn1/Ednra signaling regulates neural crest differentiation to ensure the proper patterning of pharyngeal arch arteries, which is independent of the regional identification of the pharyngeal arches along the dorsoventral axis mediated by Dlx5/Dlx6.     

The Down-Regulation of Neuroligin-2 and the Correlative Clinical Significance of Serum GABA Over-Expression in Hirschsprung’s Disease

The goal of this study was to investigate the expression level of neuroligin-2 in different colon tissue segments of children with Hirschsprung’s disease (HSCR) and the correlative clinical significance of serum Gamma-Aminobutyric Acid (serum GABA) in HSCR. Neuroligin-2 was assessed by Immunohistochemistry staining method on routine paraffin section from different colon tissue segments of HSCR (ganglionic colonic segment, transitional colonic segment and aganglionic colonic segment). Western-blot analysis and real-time fluorescence quantitative PCR(qRT-PCR) were applied to compare and evaluate the expression levels of neuroligin-2 from three segments of HSCR, and we used Enzyme-linked Immunosorbent Assay (ELISA) method to detect and compare the serum GABA between HSCR and non-HSCR. Immunohistochemistry staining demonstrated that intensive neuroligin-2 staining was detected in the ganglion cells in the ganglionic colonic and transitional colonic segments from the HSCR children; however, neuroligin-2 staining was down-regulated significantly in the aganglionic colonic segments. The expression levels of neuroligin-2 mRNA and protein in the aganglionic colonic segment were decreased compared to the ganglionic colonic segment and transitional colonic segment (P < 0.05). And the level of serum GABA was significantly higher in HSCR than that in non-HSCR. The expression of neuroligin-2 varies from different segments of HSCR. The down-regulation of neuroligin-2 in aganglionic colonic segments may be correlated with the excessive intestine contraction and further result in HSCR. The over-expression of serum GABA may be considered as a new diagnostic method of HSCR.     

A detailed linkage map of subtelomeric murine Chromosome 12 region including the situs inversus mutation locus IV

A mouse model is an invaluable tool to tackle genesis of human congenital diseases that have so far eluded human studies. Homozygote for the iv mutation, the murine Si/Col strain presents a left-right lateralization defect of thoracic and abdominal organs and heart defects very similar to human ones. This iv mutation has been mapped to the region between the Aat and Igh-C loci, suggesting the presence of an equivalent human gene in the human syntenic 14q3 region. A precise linkage map of the region is, there-fore, of great interest since it will contribute to the genetic approach of the iv gene. Analysis of 242 backcross progeny from Mus musculus (MAI) or spretus strains of mice and SI/Col mice has allowed mapping of the iv gene to a linkage group of eight markers. It includes four genes: Aat (1-antitrypsin), Ckb (creatine kinase, brain form), Crip (cysteine-rich intestinal protein), and Igh-C (immunoglobulin heavy chain constant region complex); three murine microsatellites: D12Mit6, D12Mit7, and D12Mit8; and one new marker, D12Mtpl, defined by a minisatellite human probe, pYNZ2. After analysis of the data by the LINKAGE program, the following multilocus map has been constructed: centromere-D12Mit6-6.9 cM-D12Mit7-1.7 cM-D12Mtp1-2.6 cM-Aat-5.0 cM-(Ckb, Igh-C)-0.4 cM-D12Mit8-0.4 cM-Crip-11.2 cM-iv-telomere. This map differs from the previous map in placing iv locus telomeric to Igh-C. D12Mit6 and D12Mit7 are now precisely mapped centromeric to the locus Aat. In addition, a new locus D12Mtp1 is located between Aat and D12Mit7.     

HumanGFRA1: Cloning, Mapping, Genomic Structure, and Evaluation as a Candidate Gene for Hirschsprung Disease Susceptibility

Congenital aganglionic megacolon, commonly known as Hirschsprung disease (HSCR), is the most frequent cause of congenital bowel obstruction. Germline mutations in theRETreceptor tyrosine kinase have been shown to cause HSCR. Knockout mice forRETand for its ligand, glial cell line-derived neurotrophic factor (GDNF), exhibit both complete intestinal aganglionosis and renal defects. Recently, GDNF and GFRA1 (GDNF family receptor, also known as GDNFR-α), its GPI-linked coreceptor, were demonstrated to be components of a functional ligand for RET. Moreover,GDNFhas been implicated in rare cases of HSCR. We have mappedGFRA1to human chromosome 10q25, isolated human and mouse genomic clones, determined the gene's intron–exon boundaries, isolated a highly polymorphic microsatellite marker adjacent to exon 7, and scanned forGFRA1mutations in a large panel of HSCR patients. No evidence of linkage was detected in HSCR kindreds, and no sequence variants were found to be in significant excess in patients. These data suggest thatGFRA1's role in enteric neurogenesis in humans remains to be elucidated and that RET signaling in the gut may take place via alternate pathways, such as the recently described GDNF-related molecule neurturin and its GFRA1-like coreceptor, GFRA2.     

Treatment of Aganglionic Megacolon Mice via Neural Stem Cell Transplantation

To explore a potential methodology for treating aganglionic megacolon, neural stem cells (NSCs) expressing engineered endothelin receptor type B (EDNRB) and glial cell-derived neurotrophic factor (GDNF) genes were transplanted into the aganglionic megacolon mice. After transplantation, the regeneration of neurons in the colon tissue was observed, and expression levels of differentiation-related genes were determined. Primary culture of NSCs was obtained from the cortex of postnatal mouse brain and infected with recombinant adenovirus expressing EDNRB and GDNF genes. The mouse model of aganglionic megacolon was developed by treating the colon tissue with 0.5 % benzalkonium chloride (BAC) to selectively remove the myenteric nerve plexus that resembles the pathological changes in the human congenital megacolon. The NSCs stably expressing the EDNRB and GDNF genes were transplanted into the benzalkonium chloride-induced mouse aganglionic colon. Survival and differentiation of the implanted stem cells were assessed after transplantation. Results showed that the EDNRB and GDNF genes were able to be expressed in primary culture of NSCs by adenovirus infection. One week after implantation, grafted NSCs survived and differentiated into neurons. Compared to the controls, elevated expression of EDNRB and GDNF was determined in BAC-induced aganglionic megacolon mice with partially improved intestinal function. Those founding indicated that the genes transfected into NSCs were expressed in vivo after transplantation. Also, this study provided favorable support for the therapeutic potential of multiple gene-modified NSC transplantation to treat Hirschsprung’s disease, a congenital disorder of the colon in which ganglion cells are absent.     

High-resolution linkage map in the vicinity of the Lp locus

Looptail (Lp) is a mutation that profoundly affects neurulation in mouse and is characterized by craniorachischisis, an open neural tube extending from the midbrain to the tail in embryos homozygous for the mutation. Lp maps to the distal portion of mouse chromosome 1, and as part of a positional cloning approach, we have generated a high-resolution linkage map of the Lp chromosomal region. For this, we have carried out extensive segregation analysis in a total of 706 backcross mice informative for Lp and derived from two crosses, (Lp/ + X SJL/J)F1 X SJL/J and (Lp/ + X SWR/J)F1 X SWR/J. In addition, 269 mice from a (Mus spretus X C57BL/6J)F1 X C57BL/6J interspecific backcross were also used to order marker loci and calculate intergene distances for this region. With these mice, a total of 28 DNA markers corresponding to either cloned genes or anonymous markers of the SSLP or SSCP-types were mapped within a 5-cM interval overlapping the Lp region, with the following locus order and interlocus distances (in cM): centromere-D1Mit110 / Atp1β1 / Cd3ζ / Cd3η / D1Mit145 — D1Hun14 / D1Mit15 — D1Mit111 / D1Mit112 — D1Mit114 — D1Mit148 / D1Mit205/ D1Mit36 / D1Mit146 / D1Mit147 / D1Mit270 / D1Hun13 — Fcgr2 — Mpp — Apoa2/Fcer1γ - Lp - D1Mit149 / Spna1/Fcer1α-Eph1-Hlx1/D1Mit62. These studies have allowed the delineation of a maximum genetic interval for Lp of 0.5 cM, a size amenable to physical mapping techniques.     

Mutational Spectrum of Semaphorin 3A and Semaphorin 3D Genes in Spanish Hirschsprung patients

Hirschsprung disease (HSCR, OMIM 142623) is a developmental disorder characterized by the absence of ganglion cells along variable lengths of the distal gastrointestinal tract, which results in tonic contraction of the aganglionic colon segment and functional intestinal obstruction. The RET proto-oncogene is the major gene associated to HSCR with differential contributions of its rare and common, coding and noncoding mutations to the multifactorial nature of this pathology. In addition, many other genes have been described to be associated with this pathology, including the semaphorins class III genes SEMA3A (7p12.1) and SEMA3D (7q21.11) through SNP array analyses and by next-generation sequencing technologies. Semaphorins are guidance cues for developing neurons implicated in the axonal projections and in the determination of the migratory pathway for neural-crest derived neural precursors during enteric nervous system development. In addition, it has been described that increased SEMA3A expression may be a risk factor for HSCR through the upregulation of the gene in the aganglionic smooth muscle layer of the colon in HSCR patients. Here we present the results of a comprehensive analysis of SEMA3A and SEMA3D in a series of 200 Spanish HSCR patients by the mutational screening of its coding sequence, which has led to find a number of potentially deleterious variants. RET mutations have been also detected in some of those patients carrying SEMAs variants. We have evaluated the A131T-SEMA3A, S598G-SEMA3A and E198K-SEMA3D mutations using colon tissue sections of these patients by immunohistochemistry. All mutants presented increased protein expression in smooth muscle layer of ganglionic segments. Moreover, A131T-SEMA3A also maintained higher protein levels in the aganglionic muscle layers. These findings strongly suggest that these mutants have a pathogenic effect on the disease. Furthermore, because of their coexistence with RET mutations, our data substantiate the additive genetic model proposed for this rare disorder and further support the association of SEMAs genes with HSCR.     

Molecular markers near two mouse Chromosome 13 genes,muted and pearl,which cause platelet storage pool deficiency (SPD)

The recessive muted (mu) and pearl (pe) mutations on Chromosome (Chr) 13 cause pigment dilution and platelet storage pool deficiency (SPD) in mice. In addition, mu causes inner ear abnormalities and pe has symptoms associated with night blindness. Using an interspecific backcross involving the wild-derived Mus musculus musculus (PWK) stock, we have mapped 33 microsatellite markers and four cDNAs relative to mu, pe, and another recessive mutation, satin (sa). Analyzing a total of 528 backcross offspring, we found tight linkage between the pigment loci and several microsatellite markers (D13Mit87, D13Mit88, D13Mit137 with mu; and D13Mit104, D13Mit160, D13Mit161, and D13Mit169 with pe). These markers should aid the eventual molecular identification of these specific SPD genes.     

Expression and Significance of Neuroligins in Myenteric Cells of Cajal in Hirschsprung's Disease

Purpose

The aim of this study was to investigate the expression and significance of neuroligins in myenteric cells of Cajal (ICC-MY) in Hirschsprung’s disease (HSCR).

Methods

Longitudinal muscle with adherent myenteric plexus (LMMP) from surgical excision waste colon of HSCR children were prepared by peeling off the mucous layer, sub-mucosal layer and circular muscle. Neuroligins, c-Kit (c-Kit-immunoreactivity representing ICC) and their relationship were assessed by double labeling immunofluorescence staining. ICC-MY were dissociated and cultured from LMMP by enzymolysis method, and were purified and analyzed using a combination of magnetic-activated cell sorting (MACS) and flow cytometry (FCM). Western-blot analysis was applied to compare and evaluate the expression levels of neuroligins in ICC-MY which were dissociated from different segments of HSCR (ganglionic colonic segment, transitional colonic segment and aganglionic colonic segment).

Results

Neuroligins and c-Kit were expressed on the same cells (ICC-MY); ICC-MY were dissociated, cultured and purified. For HSCR, neuroligins were expressed significantly in ICC-MY from ganglionic colonic segments, moderately in those from transitional colonic segments and down-regulated significantly in those from aganglionic colonic segments.

Conclusions

Neuroligins were expressed in ICC-MY of human beings, and the expression varies from different segments of HSCR. This abnormal expression might play an important role in the pathogenesis of this disease through affecting the synaptic function of ICC-MY.     

Long non‐coding RNA FAL1 functions as a ceRNA to antagonize the effect of miR‐637 on the down‐regulation of AKT1 in Hirschsprung's disease

Objectives

Emerged evidence demonstrates that long non‐coding RNAs (lncRNAs) may play quintessential regulatory roles in the cellular processes, tumourigenesis and the development of disease. Though focally amplified lncRNA on chromosome 1 (FAL1) has been identified to have crucial functions in many diseases, its biological mechanism in the development of Hirschsprung's disease (HSCR) still remains unknown.

Materials and methods

The expression levels of FAL1 in HSCR aganglionic tissues and matched normal specimens were detected by quantitative real‐time PCR (qRT‐PCR). Cell proliferation and migration were detected by Cell Counting Kit‐8 (CCK‐8) assay, Ethynyl‐deoxyuridine (EdU) assay and transwell assay relatively. Cell cycle and apoptosis were assessed using flow cytometer analysis. Moreover, the novel targets of FAL1 were confirmed with the help of bioinformatics analysis and dual‐luciferase reporter assay. Western blot assay as well as RNA immunoprecipitation (RIP) assay was conducted to investigate the potential mechanism.

Results

FAL1 expression was markedly down‐regulated in HSCR aganglionic tissues and decreased FAL1 expression was associated with the diagnosis of HSCR. Cell functional analyses indicated that FAL1 overexpressing notably promoted cell proliferation and migration, while down‐regulation of FAL1 suppressed cell proliferation and migration. Additionally, Flow cytometry assay demonstrated that knockdown of FAL1 induced markedly cell cycle stalled in the G0/G1 phase. Furthermore, FAL1 could positively regulate AKT1 expression by competitively binding to miR‐637.

Conclusions

These results illuminated that FAL1 may work as a ceRNA to modulate AKT1 expression via competitively binding to miR‐637 in HSCR, suggesting that it may be clinically valuable as a biomarker of HSCR.

     

Phactr4

The enteric nervous system (ENS) is critically important for many intestinal functions such as peristalsis and secretion. Defects in the embryonic formation of the ENS cause Hirschsprung disease (HSCR) or megacolon, a severe birth defect that affects approximately 1 in 5,000 newborns. One of the least understood aspects of ENS development are the cellular and molecular mechanisms that control chain migration of the ENS cells during their migration into and along the embryonic gut. We recently reported a mouse model of HSCR in which mutant embryos carrying a hypomorphic allele of the Phactr4 gene show an embryonic gastrointestinal defect due to loss of enteric neurons in the colon. We found that Phactr4 modulates integrin signaling and cofilin activity to coordinate the forces that drive enteric neural crest cell (ENCC) migration in the mammalian embryo. In this extra view, we briefly summarize the current knowledge on integrin signaling in ENCC migration and introduce the Phactr protein family. Employing the ENS as a model, we shed some light on the mechanisms by which Phactr4 regulates integrin signaling and controls the cell polarity required for directional ENCC migration in the mouse developing gut.