Alternative titles; symbols
HGNC Approved Gene Symbol: ALX4
Cytogenetic location: 11p11.2 Genomic coordinates (GRCh38) : 11:44,260,440-44,310,139 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 11p11.2 | {Craniosynostosis 5, susceptibility to} | 615529 | Autosomal dominant | 3 |
| Frontonasal dysplasia 2 | 613451 | Autosomal recessive | 3 | |
| Parietal foramina 2 | 609597 | Autosomal dominant | 3 |
The ALX4 gene encodes a homeodomain transcription factor important for many developmental processes (summary by Bertola et al., 2013).
Wu et al. (2000) identified a human BAC clone mapping to chromosome 11p11.2 that contained a region homologous to the MSX2 gene (123101), which is mutated in parietal foramina-1 (PFM1; 168500). Further sequence analysis demonstrated that this region contained the human ortholog of the mouse Aristaless-like-4 gene (Alx4). Wu et al. (2000) assembled a full-length ALX4 cDNA encoding a deduced 410-amino acid protein that shares 80% sequence identity with the mouse Alx4 protein. Northern blot analysis demonstrated that expression of both the human and mouse genes is restricted to bone.
The ALX4 coding region comprises 4 exons (Wu et al., 2000; Wuyts et al., 2000).
Wu et al. (2000) identified the ALX4 gene within the critical region of Potocki-Shaffer syndrome (601224), also known as the proximal 11p deletion syndrome (P11pDS), which includes parietal foramina as a feature.
Qu et al. (1998) mapped the mouse Alx4 gene of a region of chromosome 2 that shares homology of synteny with human chromosome 11p12-q12.
McGaughran et al. (1995) described a patient with the combination of multiple exostoses (133701), features of Potocki-Shaffer syndrome (601224), and WAGR syndrome (Wilms tumor, aniridia, genitourinary anomalies, and mental retardation; 194072) associated with a del(11)(p14.2p11.2). The ALX4 gene contains 4 exons and maps centromeric to D11S2095 in reverse orientation to the EXT2 gene (608210); hence ALX4 lies just outside the P11pDS critical interval proposed by Wuyts et al. (1996). In the patient defining this centromeric boundary (McGaughran et al., 1995), ALX4 expression may have been abrogated by a position effect.
Parietal Foramina 2
Using FISH, Wu et al. (2000) tested for the presence or heterozygous deletion of the region corresponding to the BAC clone containing ALX4 on 11p in 2 patients with the Potocki-Shaffer syndrome and found that this region was deleted in these patients. The authors stated that the involvement of Alx4 in murine skull development (Qu et al., 1997), its bone-specific expression pattern, the finding that Alx4 is a dosage-sensitive gene in the mouse, and the localization of a human genomic clone containing ALX4 to 11p11.2, with hemizygosity in patients with deletion of 11p11.2 who have biparietal foramina, supported the contention that ALX4 is a candidate gene for parietal foramina in the Potocki-Shaffer syndrome.
Wuyts et al. (2000) identified heterozygous mutations in the ALX4 gene in affected members of 2 families segregating parietal foramina (PFM2; 609597). One mutation resulted in a premature stop codon (605420.0004) and the second was a missense mutation in a conserved region of the homeobox domain (R272P; 605420.0005), which Wuyts et al. (2000) concluded would disrupt DNA binding. Wuyts et al. (2000) did not find a mutation in either ALX4 or MSX2 in a third family with PFM.
In 4 families segregating PFM, Mavrogiannis et al. (2001) identified 3 distinct heterozygous ALX4 mutations in 19 affected individuals (605420.0001-605420.0003). The cranial phenotype associated with the ALX4 mutations ranged from severe deficiency in mineralization of the skull vault in infancy (cranium bifidum) to apparent nonpenetrance; most individuals had classic PFM. Mavrogiannis et al. (2001) found no significant difference in the size of PFM between ALX4 and MSX2 mutations, but cranium bifidum had not been described in association with mutations of MSX2. It remained to be determined whether MSX2 and ALX4 act hierarchically or in parallel developmental pathways. PFM and craniosynostosis (premature fusion of the cranial sutures) may result from opposite perturbations in osteogenic differentiation in the skull vault. Both occur with different mutations in the MSX2 gene, suggesting that ALX4 is a candidate gene for craniosynostosis as well as for PFM.
Bertola et al. (2013) and Altunoglu et al. (2014) independently identified heterozygous ALX4 mutations (605420.0012 and 605420.0013) in affected members of 2 unrelated families with autosomal dominant PFM associated with mild features of frontonasal dysplasia. The reports expanded the phenotype associated with ALX4 heterozygous mutations. The authors postulated that a dominant-negative effect may cause frontonasal abnormalities in addition to PFM, whereas haploinsufficiency may result in isolated PFM. Altunoglu et al. (2014) discussed the implications for genetic counseling.
Frontonasal Dysplasia 2
In affected members of 2 consanguineous Turkish families segregating frontonasal dysplasia mapping to chromosome 11p11.2-q12.3 (FND2; 613451), Kayserili et al. (2009) identified homozygosity for a nonsense mutation in the ALX4 gene (605420.0008). Affected individuals exhibited a large skull defect, coronal craniosynostosis, hypertelorism, severely depressed nasal bridge and ridge, bifid nasal tip, cryptorchidism, agenesis of the corpus callosum, total alopecia, and mental retardation. Skin from a biopsy of an affected individual demonstrated a hypomorphic interfollicular epidermis with reduced suprabasal layers associated with impaired interfollicular epidermal differentiation. Hair follicle-like structures were present but showed altered differentiation. Kayserili et al. (2009) concluded that ALX4 plays a critical role in craniofacial development as well as in skin and hair follicle development.
In a boy, born of consanguineous Turkish parents, with a mild form of FND2, Kayserili et al. (2012) identified a homozygous missense mutation in the ALX4 gene (Q225E; 605420.0011). Functional studies were not performed. The parents, who were heterozygous for the mutation, had uncovered bilateral parietal foramina of small size. The patient had mild nasal malformations, parietal foramina, and a hypoplastic and kinked corpus callosum, but normal psychomotor development. Kayserili et al. (2012) postulated that the less severe phenotype observed in this patient compared to the patients reported by Kayserili et al. (2009) was due to the missense mutation retaining some functionality.
Susceptibility to Craniosynostosis 5
Yagnik et al. (2012) studied 203 patients with nonsyndromic craniosynostosis (see CRS5, 615529), of whom 197 had single sagittal suture fusion and 6 had multiple-suture fusion with sagittal involvement. In 111 of the 203 probands with single-suture fusion and all 6 probands with multiple-suture fusion, Boyadjiev (2007) had previously excluded mutations in hotspot areas of the FGFR1 (136350), FGFR2 (176943), and FGFR3 (134934) genes, and the entire TWIST1 gene (601622). Yagnik et al. (2012) sequenced the entire coding region of the candidate gene ALX4 (605420) in the 203 CRS probands and identified heterozygous missense variants, V7F (605420.0009) and K211E (605420.0010), in 2 of the probands. Functional analysis showed a gain of function for both variants, which were also present in an unaffected parent from each family. Yagnik et al. (2012) concluded that the variants represent low-penetrance mutations that predispose to CRS but are not causative by themselves.
The zone of polarizing activity (ZPA) is a group of mesenchymal cells localized to the posterior margin of the limb bud mesoderm that express Shh (600725) and function as a signaling center for patterning the anterior-posterior axis. Mice with the 'Strong luxoid' (lst) phenotype exhibit preaxial polydactyly that is preceded during development by formation of an ectopic ZPA and expression of Shh in the anterior mesenchyme of the limb bud. Qu et al. (1998) identified 3 inactivating mutations in the Alx4 gene in 3 independent strains of mice with the Strong luxoid phenotype, in addition to other strain-specific phenotypes. The 3 Alx4 mutations included a large deletion encompassing the Alx4 gene, a 16-bp deletion within the region encoding the paired-type homeodomain, resulting in a frameshift, and a missense mutation, arg206 to gln (R206Q), affecting a critical conserved arginine within the paired-type homeodomain. Qu et al. (1998) showed that wildtype Alx4 bound as a dimer to the high-affinity P3 palindromic DNA sequence, whereas the R206Q homeodomain failed to bind P3 as either a dimer or monomer. When expressed in human 293 cells, wildtype Alx4 efficiently activated transcription of a reporter construct containing the P3 sequence, but the R206Q mutant did not. Qu et al. (1998) concluded that the Alx4 mutations caused the polydactyly and abnormal ZPA signaling in the Strong luxoid mice, while the variable phenotypes present in heterozygous and homozygous mice, including hemimelia, ventral body wall defects, and alopecia, resulted from strain-specific rather than allele-specific effects.
Independently, Takahashi et al. (1998) identified the semidominant 16-bp deletion in the Alx4 gene in Strong luxoid mice. Using chicken embryos, they found that Alx4 expression originated from the anterior half of the apical ectodermal ridge. Alx4 acted downstream of early events that established anterior-posterior gene asymmetries, and Alx4 expression was downregulated prior to Shh upregulation. Local application of Shh and fibroblast growth factor (see 131220), or removal of the apical ectodermal ridge, suggested that Alx4 and Shh interact in a negative-feedback loop during limb outgrowth. The interaction of Alx4 and Shh in mice was independent of the negative Shh regulator Gli3 (165240). Takahashi et al. (1998) concluded that ALX4 is required for correct ZPA positioning.
In affected members of a family segregating parietal foramina-2 (PFM2; 609597), Mavrogiannis et al. (2001) identified a heterozygous c.418C-T transition in the ALX4 gene, resulting in a gln140-to-ter (Q140X) substitution. The mutation was predicted to completely or partially eliminate the DNA-binding homeodomain.
In affected members of a family segregating parietal foramina-2 (PFM2; 609597), Mavrogiannis et al. (2001) identified a heterozygous c.736C-T transition in the ALX4 gene, resulting in a gln246-to-ter (Q246X) substitution. The mutation was predicted to completely or partially eliminate the DNA-binding homeodomain.
In affected members of 2 unrelated families with parietal foramina-2 (PFM2; 609597), Mavrogiannis et al. (2001) identified a heterozygous c.653G-A transition in the ALX4 gene, resulting in an arg218-to-gln (R218Q) substitution. The mutation substituted a highly conserved residue in the N-terminal arm of the homeodomain that contacts the minor groove of DNA. In mouse, the identical mutation underlies the Strong luxoid allele, Alx4(lst), and abolishes DNA binding and transcriptional activation in vitro, with no detectable dominant-negative effect.
In a review of 7 patients from 2 unrelated families with parietal foramina due to the R218Q mutation, Mavrogiannis et al. (2006) found that the patients had disproportionately wide defects compared to patients with other mutations. The authors postulated a dominant-negative effect.
In a family segregating parietal foramina-2 (PFM2; 609597), Wuyts et al. (2000) identified a 1-bp deletion at position 504 (c.504delT) of the cDNA (adenosine of start codon +1) resulting in a premature stop codon after 179 amino acids.
In affected members of a family segregating parietal foramina-2 (PFM2; 609597), Wuyts et al. (2000) identified a G-to-C substitution at position 815 resulting in the replacement of an arginine by a proline residue (arg272 to pro; R272P). Arginine-272 is in a highly conserved region of the homeobox, and the authors concluded that this mutation was likely to disrupt DNA binding.
In 5 affected members of a family with parietal foramina-2 (PFM2; 609597), Mavrogiannis et al. (2006) identified a heterozygous c.620C-A transversion in the ALX4 gene, resulting in a ser207-to-ter (S207X) substitution. Three sibs also had mild dysmorphic facial features, high palate, and short, broad thumbs. The affected father and paternal grandfather had parietal foramina and broad thumbs. All had normal mental development.
In a female with parietal foramina-2 (PFM2; 609597), Mavrogiannis et al. (2006) identified a heterozygous 10-bp deletion from nucleotides 385 to 394 in the ALX4 gene, resulting in a frameshift and premature termination.
In affected members of 2 consanguineous Turkish families segregating frontonasal dysplasia-2 (FND2; 613451), Kayserili et al. (2009) identified homozygosity for a 793C-T transition in the ALX4 gene, which resulted in premature termination at codon 265 (R265X). Affected individuals exhibited a large skull defect, coronal craniosynostosis, hypertelorism, severely depressed nasal bridge and ridge, bifid nasal tip, cryptorchidism, agenesis of the corpus callosum, total alopecia, and mental retardation.
In a patient with isolated sagittal craniosynostosis (CRS5; 615529), Yagnik et al. (2012) identified heterozygosity for a c.19G-T transversion in the ALX4 gene, resulting in a val7-to-phe (V7F) substitution at a highly conserved residue. Dual-luciferase assay in transfected human calvarial osteoblasts demonstrated that the V7F mutant resulted in a 5.8-fold increase in luminescence compared to wildtype. Postsurgical evaluation of the proband confirmed age-appropriate development and the absence of associated anomalies. The mutation was also present in an unaffected parent; Yagnik et al. (2012) concluded that the V7F variant represents a low-penetrance gain-of-function mutation that predisposes to CRS but is not causative by itself.
In a patient with isolated craniosynostosis involving the metopic, right squamous, and sagittal sutures (CRS5; 615529), Yagnik et al. (2012) identified heterozygosity for a c.631A-G transition in the ALX4 gene, resulting in a lys211-to-glu (K211E) substitution at a highly conserved residue. Dual-luciferase assay in transfected human calvarial osteoblasts demonstrated that the K211E mutant resulted in a 5-fold increase in luminescence compared to wildtype. Postsurgical evaluation of the proband confirmed age-appropriate development and the absence of associated anomalies. The mutation was also present in an unaffected parent who had been evaluated by a clinical geneticist; Yagnik et al. (2012) concluded that the K211E variant represents a low-penetrance gain-of-function mutation that predisposes to CRS but is not causative by itself.
In a boy, born of consanguineous Turkish parents, with a mild form of frontonasal dysplasia-2 (FND2; 613451), Kayserili et al. (2012) identified a homozygous c.673C-G transversion in exon 2 of the ALX4 gene, resulting in a gln225-to-glu (Q225E) substitution at a highly conserved residue in the first helix motif of the homeodomain. The mutation was not detected in 100 ethnically matched control chromosomes; functional studies were not performed. The parents, who were heterozygous for the mutation, had uncovered bilateral parietal foramina of small size. The boy had hypertelorism, upslanting palpebral fissures, broad nasal bridge and ridge, bifid nasal tip, broad columella, cleft alae nasi, and an upper labiogingival sulcus. Skull and brain imaging showed bilateral parietal foramina and a mildly hypoplastic, kinked body of the corpus callosum and underdevelopment of the vermis. He had normal psychomotor development.
In a son and his mother with parietal foramina-2 (PFM2; 609597) and frontonasal abnormalities, Bertola et al. (2013) identified a heterozygous deletion/insertion mutation in the last exon of the ALX4 gene (c.1080-1089del/ins), resulting in premature termination (Asp326fsTer21). The mutation was predicted to result in loss of the conserved OAR domain. Bertola et al. (2013) postulated that the mutation would escape nonsense-mediated mRNA decay and could interfere with the normal ALX4 allele in a dominant-negative manner.
In 4 members of a consanguineous Turkish family with parietal foramina-2 (PFM2; 609597), Altunoglu et al. (2014) identified a heterozygous c.646C-G transversion in exon 2 of the ALX4 gene, resulting in an arg216-to-gly (R216G) substitution at a conserved residue in the homeodomain. The mutation was not found in publicly available SNP databases; functional studies were not performed. In addition to minute parietal foramina confirmed by imaging, the patients showed a phenotypic spectrum ranging from mild nasal clefting and broad columella to subtle changes in nasal configuration.
In a girl, born of consanguineous Iranian parents, with frontonasal dysplasia-2 (FND2; 613451), Kariminejad et al. (2014) identified a homozygous 1-bp deletion (c.503delC, NM_021926.3) in exon 2 of the ALX4 gene, resulting in a frameshift and premature termination (Pro168LeufsTer12). Each parent, who showed mild features, was heterozygous for the mutation, which was predicted to result in a complete loss of protein function.
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