4]Identification and functional analysis of mutations in the human PAX6 gene

Abstract

This chapter discusses the identification and functional analysis of mutations in the human PAX6 gene. One class of human disorders only recently subjected to mutation analysis at the molecular level includes those responsible for congenital malformation syndromes. In several cases, these syndromes are because of mutations in transcription factors. The chapter presents a review on the methodologies employed to detect and to analyze the functional consequences of mutations in one such transcription factor, which is encoded by the PAX6 developmental control gene. Mutations in PAX6 are responsible for a dominantly inherited defect of human ocular and central nervous system (CNS) development, called “aniridia.” PAX6 belongs to a family of evolutionarily conserved Pax genes, unified by the presence of a paired box, which encodes a 128-amino acid bipartite DNA-binding domain. Because the loss of function mutations in Pax genes give rise to semidominant phenotypes, PAX mutations have been identified in a number of human and mouse developmental syndromes, such as aniridia ( PAX6 ), Waardenburg's syndrome ( PAX3 ), and renal agenesis with coloboma ( PAX2 ). Thus, the identification of PAX gene mutations has a direct clinical relevance. In addition, human PAX gene mutations provide valuable insight into the molecular function of PAX gene products. The chapter presents a review on a polymerase chain reaction (PCR)-based single-strand conformational polymorphism (SSCP) assay for detecting mutations in the human PAX6 gene and discusses methods pertinent to the functional analysis of PAX6 mutations.