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

Adam Davis,Edmund Stellwag,Michael Reubens

doi:10.3390/jdb4020015

Abstract

Hoxa2 is an evolutionarily conserved developmental regulatory gene that functions to specify rhombomere (r) and pharyngeal arch (PA) identities throughout the Osteichthyes. Japanese medaka (Oryzias latipes) hoxa2a, like orthologous Hoxa2 genes from other osteichthyans, is expressed during embryogenesis in r2–7 and PA2-7, whereas the paralogous medaka pseudogene, ψhoxa2b, is expressed in noncanonical Hoxa2 domains, including the pectoral fin buds. To understand the evolution of cis-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 hoxa2a and ψhoxa2b in transient and stable-line transgenic medaka embryos. The CREs tested were previously shown to contribute to directing mouse Hoxa2 gene expression in r3, r5, and PA2-4. Our results reveal the presence of sequence elements embedded in the medaka hoxa2a and ψhoxa2b upstream enhancer regions (UERs) that mediate expression in r4 and the PAs (hoxa2a r4/CNCC element) or in r3–7 and the PAs ψhoxa2b 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 Hoxa2 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.

Highlights

Clustered Hox genes are a family of evolutionarily-related developmental regulatory genes that function to pattern regional tissue identities along the anterior-posterior (A-P) axis of animal species [1]
These results suggested that sequence elements downstream of RE4 within the of RE4 within the medaka hoxa2a upstream enhancer region (UER)(K20-RE5), RE3, RE2 and RE5, are necessary for medaka hoxa2a UER(K20-RE5), RE3, RE2 and RE5, are necessary for directing medaka directing medaka hoxa2a expression in r4 and the cranial neural crest cells (CNCCs) of PA2 and the posterior arches
BoxA, RE4, and RE3 were retained, did not show any enhanced green fluorescent protein (eGFP) expression in the hindbrain (0%) or in the pharyngeal arches (0%) (Table 2). These results showed that the sequence containing the elements corresponding to RE3 and RE2 of the medaka hoxa2a UER(K20-RE5) is required for r4 expression but the surrounding elements corresponding to Krox20, BoxA, RE4 and RE5 are not

Summary

Introduction

Clustered Hox genes are a family of evolutionarily-related developmental regulatory genes that function to pattern regional tissue identities along the anterior-posterior (A-P) axis of animal species [1]. Multiple genome level duplications have expanded the total number of Hox clusters from one in chordates to four in tetrapods and at least seven or eight in most teleost fishes [5,6,7,8,9,10,11]. Post-genome duplication independent gene loss has generated clustered paralog groups (PGs) that differ in gene number depending on the historical timing of gene losses relative to genome duplications [6,12,13,14,15].

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Discussion

Conclusion