PAX3

Paired box gene 3

The PAX3 (paired box gene 3) gene encodes a member of the paired box or PAX family of transcription factors. The PAX family consists of nine human (PAX1 PAX9) and nine mouse (Pax1 Pax9) members arranged into four subfamilies. Human PAX3 and mouse Pax3 are present in a subfamily along with the highly homologous human PAX7 and mouse Pax7 genes. The human PAX3 gene is located in the 2q36.1 chromosomal region, and contains 10 exons within a 100 kb region.

Alternative splicing and processing generates multiple PAX3 isoforms that have been detected at the mRNA level. PAX3e is the longest isoform and consists of 10 exons that encode a 505 amino acid protein. In other mammalian species, including mouse, the longest mRNAs correspond to the human PAX3c and PAX3d isoforms, which consist of the first 8 or 9 exons of the PAX3 gene, respectively. Shorter PAX3 isoforms include mRNAs that skip exon 8 (PAX3g and PAX3h) and mRNAs containing 4 or 5 exons (PAX3a and PAX3b). In limited studies comparing isoform expression, PAX3d is expressed at the highest levels. From a functional standpoint, PAX3c, PAX3d, and PAX3h stimulate activities such as cell growth whereas PAX3e and PAX3g inhibit these activities, and PAX3a and PAX3b show no activity or inhibit these endpoints. A common alternative splice affecting the PAX3 mRNA involves the sequence CAG at the 5’ end of exon 3. This splice either includes or excludes these three bases, thus resulting in the presence or absence of a glutamine residue in the paired box motif. Limited sequencing studies of full length human cDNAs identified this splicing event as a variant of the PAX3d isoform, and this spliced isoform has been separately termed the PAX3i isoform. The Q+ and Q− isoforms of PAX3 are generally co expressed in cells. At the functional level, the Q+ isoform shows similar or less DNA binding and transcriptional activation than the Q− isoform.

During development, one of the major lineages expressing Pax3 is the skeletal muscle lineage. Pax3 expression is first seen in the pre somitic paraxial mesoderm, and then ultimately becomes restricted to the dermomyotome, which forms from the dorsal region of the somites. To form skeletal muscle in central body segments, PAX3 expressing cells detach from the dermomyotome and then Pax3 expression is turned off as Myf5 and MyoD1 expression is activated. To form other skeletal muscles, Pax3 expressing cells detach from the dermomyotome and migrate to more distant sites, such as the limbs and diaphragm. A subset of these Pax3 expressing dermomyotome derived cells also serves as an ongoing progenitor pool for skeletal muscle growth during fetal development. During later developmental stages, myogenic precursors expressing Pax3 and/or Pax7 form satellite cells within the skeletal muscle, which contribute to postnatal muscle growth and muscle regeneration. These adult satellite cells remain quiescent until injury occurs, and then are stimulated to divide and regenerate the injured muscle. Pax3 is also involved in the development of the nervous system. Expression of Pax3 is first detected in the dorsal region of the neural groove and, as this neural groove deepens to form the neural tube, Pax3 is expressed in the dorsal portion of the neural tube. As the neural tube enlarges, Pax3 expression is localized to proliferative cells in the inner ventricular zone and then this expression is turned off as these cells migrate to more superficial regions. Pax3 is expressed along the length of the neural tube and throughout much of the developing brain, and this expression is subsequently turned off during later developmental stages in a rostral to caudal direction. During early development, Pax3 expression also occurs at the lateral and posterior margins of the neural plate, which is the region from which the neural crest arises. Pax3 is later expressed by various cell types and structures arising from the neural crest, such as melanoblasts, Schwann cell precursors, and dorsal root ganglia. In addition, Pax3 expressing cells derived from the neural crest contribute to the formation of other structures, such as the inner ear, mandible and maxilla. Pax3 controls the location of the nasion (a facial feature between the eyes and at the top of the nose), and is associated with the presence of a unibrow.

Germline mutations of the Pax3 gene cause the splotch phenotype in mice. At the molecular level, this phenotype is caused by point mutations or deletions that alter or abolish Pax3 transcriptional function. In the heterozygous state, the splotch phenotype is characterized by white patches in the belly, tail and feet. These white spots are attributed to localized deficiencies in pigment forming melanocytes resulting from neural crest cell defects. In the homozygous state, these Pax3 mutations cause embryonic lethality, which is associated with prominent neural tube closure defects and abnormalities of neural crest derived structures, such as melanocytes, dorsal root ganglia and enteric ganglia. Heart malformations also result from the loss of cardiac neural crest cells, which normally contribute to the cardiac outflow tract and innervation of the heart. Finally, limb musculature does not develop in the homozygotes and axial musculature demonstrates varying abnormalities. These myogenic effects are caused by increased cell death of myogenic precursors in the dermomyotome and diminished migration from the dermomyotome. Germline mutations of the PAX3 gene occur in the human disease Waardenburg syndrome, which consists of four autosomal dominant genetic disorders (WS1, WS2, WS3 and WS4). Of the four subtypes, WS1 and WS3 are usually caused by PAX3 mutations. All four subtypes are characterized by hearing loss, eye abnormalities and pigmentation disorders. In addition, WS1 is frequently associated with a midfacial alteration called dystopia canthorum, while WS3 (Klein Waardenburg syndrome) is frequently distinguished by musculoskeletal abnormalities affecting the upper limbs. Most WS1 cases are caused by heterozygous PAX3 mutations while WS3 is caused by either partial or total deletion of PAX3 and contiguous genes or by smaller PAX3 mutations in the heterozygous or homozygous state. These PAX3 mutations in WS1 and WS3 include missense, nonsense and splicing mutations; small insertions; and small or gross deletions. Though these changes are usually not recurrent, the mutations generally occur in exons 2 through 6 with exon 2 mutations being most common. As these exons encode the paired box and homeodomain, these mutations often affect DNA binding function.

Alveolar rhabdomyosarcoma (ARMS) is an aggressive soft tissue sarcoma that occurs in children and is usually characterized by a recurrent t(2;13)(q35;q14) chromosomal translocation. This 2;13 translocation breaks and rejoins portions of the PAX3 and FOXO1 genes to generate a PAX3 FOXO1 fusion gene that expresses a PAX3 FOXO1 fusion transcript encoding a PAX3 FOXO1 fusion protein. PAX3 and FOXO1 encode transcription factors, and the translocation results in a fusion transcription factor containing the N terminal PAX3 DNA binding domain and the C terminal FOXO1 transactivation domain. A smaller subset of ARMS cases is associated with less common fusions of PAX7 to FOXO1 or rare fusions of PAX3 to other transcription factors, such as NCOA1. Compared to the wild type PAX3 protein, the PAX3 FOXO1 fusion protein more potently activates PAX3 target genes. In addition, PAX3 FOXO1 binds along with MYOD1, MYOG and MYCN as well as chromatin structural proteins, such as CHD4 and BRD4, to contribute to the formation of super enhancers in the vicinity of a subset of these target genes. These dysregulated target genes contribute to tumorigenesis by altering signaling pathways that affect proliferation, cell death, myogenic differentiation, and migration. A t(2;4)(q35;q31.1) chromosomal translocation that fuses the PAX3 and MAML3 genes occurs in biphenotypic sinonasal sarcoma (BSNS), a low grade adult malignancy associated with both myogenic and neural differentiation. MAML3 encodes a transcriptional coactivator involved in Notch signaling. The PAX3 MAML3 fusion juxtaposes the N terminal PAX3 DNA binding domain with the C terminal MAML3 transactivation domain to create another potent activator of target genes with PAX3 binding sites. Of note, PAX3 is rearranged without MAML3 involvement in a smaller subset of BSNS cases, and some of these variant cases contain a PAX3 NCOA1 or PAX3 FOXO1 fusion. Though PAX3 FOXO1 and PAX3 NCOA1 fusions can be formed in both ARMS and BSNS, there are differences in the pattern of activated downstream target genes suggesting that the cell environment has an important role in modulating the output of these fusion transcription factors. In addition to tumors with PAX3 related fusion genes, there are several other tumor categories that express the wild type PAX3 gene. The presence of PAX3 expression in some tumors can be explained by their derivation from developmental lineages normally expressing wild type PAX3. For example, PAX3 is expressed in cancers associated with neural tube derived lineages, (e. g., glioblastoma), neural crest derived lineages (e. g., melanoma) and myogenic lineages (e. g., embryonal rhabdomyosarcoma). However, PAX3 is also expressed in other cancer types without a clear relationship to a PAX3 expressing developmental lineages, such as breast carcinoma and osteosarcoma. In these wild type PAX3 expressing cancers, PAX3 function impacts on the control of proliferation, apoptosis, differentiation and motility. Therefore, wild type PAX3 exerts a regulatory role in tumorigenesis and tumor progression, which may be related to its role in normal development.