Functional and Comparative Genomics of Hoxa2 Gene cis-Regulatory Elements: Evidence for Evolutionary Modification of Ancestral Core Element Activity.

is an evolutionarily conserved developmental regulatory gene that functions to specify rhombomere (r) and pharyngeal arch (PA) identities throughout the Osteichthyes. Japanese medaka () like orthologous genes from other osteichthyans, is expressed during embryogenesis in r2-7 and PA2-7, whereas the paralogous medaka pseudogene, , is expressed in noncanonical domains, including the pectoral fin buds. To understand the evolution of -regulatory element (CRE) control of gene expression, we conducted eGFP reporter gene expression studies with extensive functional mapping of several conserved CREs upstream of medaka and in transient and stable-line transgenic medaka embryos. The CREs tested were previously shown to contribute to directing mouse gene expression in r3, r5, and PA2-4. Our results reveal the presence of sequence elements embedded in the medaka and upstream enhancer regions (UERs) that mediate expression in r4 and the PAs ( r4/CNCC element) or in r3-7 and the PAs r3-7/CNCC element), respectively. Further, these elements were shown to be highly conserved among osteichthyans, which suggests that the r4 specifying element embedded in the UER of is a deeply rooted rhombomere specifying element and the activity of this element has been modified by the evolution of flanking sequences that redirect its activity to alternative developmental compartments.