Paralogous Vitamin D Receptors in Teleosts: Transition of Nuclear Receptor Function

Abstract

The availability of multiple teleost (bony fish) genomes is providing unprecedented opportunities to understand the diversity and function of gene duplication events using comparative genomics. Here we describe the cloning and functional characterization of two novel vitamin D receptor (VDR) paralogs from the freshwater teleost medaka (Oryzias latipes). VDR sequences were identified through mining of the medaka genome database in which gene organization and structure was determined. Two distinct VDR genes were identified in the medaka genome and mapped to defined loci. Each VDR sequence exhibits unique intronic organization and dissimilar 5' untranslated regions, suggesting they are not isoforms of the same gene locus. Phylogenetic comparison with additional teleosts and mammalian VDR sequences illustrate that two distinct clusters are formed separating aquatic and terrestrial species. Nested within the teleost cluster are two separate clades for VDRalpha and VDRbeta. The topology of teleost VDR sequences is consistent with the notion of paralogous genes arising from a whole genome duplication event prior to teleost radiation. Functional characterization was conducted through the development of VDR expression vectors including Gal4 chimeras containing the yeast Gal4 DNA binding domain fused to the medaka VDR ligand binding domain and full-length protein. The common VDR ligand 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)(2)D(3)] resulted in significant transactivation activity with both the Gal4 and full-length constructs of medaka (m) VDRbeta. Comparatively, transactivation of mVDRalpha with 1alpha,25(OH)(2)D(3) was highly attenuated, suggesting a functional divergence between these two nuclear receptor paralogs. We additionally demonstrate through coactivator studies that mVDRalpha is still functional; however, it exhibits a different sensitivity to 1alpha,25(OH)(2)D(3), compared with VDRbeta. These results suggest that in mVDRalpha and VDRbeta have undergone a functional divergence through a process of sub- and/or neofunctionalization of VDR nuclear receptor gene pairs.

Effects of dietary vitamin D3 administration on innate immune parameters of seabream (Sparus aurata L.)

“…Since fish stand between the invertebrates and vertebrates in evolution they serve as a unique model for the study of the evolutionary and physiological significance of vD 3 . In spite of this, knowledge on the role of vD 3 in fish is poor because comparative physiological studies on marine teleosts are few [11], and the role of vD 3 in Ca regulation in teleosts remains equivocal [27][28][29].…”

Vitamin D: calcium and bone homeostasis during evolution

“…In late or advanced teleost fish, VDR is also present as demonstrated in the freshwater medaka (Ozyrias latipes) 41 and green spotted pufferfish (Tetraodon nigrividis). 48 These teleosts have an acellular postcranial bone (lack of osteocytes) and no multinucleated osteoclasts 49 (see discussion below).…”

“…The teleosts display two paralogous VDRs (a and b) with different functionality in different fish. [39][40][41] These fish VDRs have about 70% homology with VDRs from higher vertebrates. The zebrafish VDR is structurally and functionally very similar to that of mammals and in fact the zebrafish VDR-ligand binding domain was used to cocrystalize with 1,25(OH) 2 D 3 and define its ligand binding pocket.…”

In Summary

“…In instances in which NR functions between paralogs have been examined, putative functional differences have been noted. This is particularly evident with trout glucocorticoid receptors, where GR paralogs exhibit differential transactivation and transrepression activities [31]. Similarly, functional differences have been observed with retinoic acid receptors, which play a major role in chordate embryonic development; fushi tarazu gene (transcription factor)/ steroidogenic transcription factor 1 (FTX/SF-1), a critical receptor in reproductive processes; and VDRs, which are thought to play critical roles in ion homeostasis, immune function, and neurodevelopment [31].…”

“…In the second model, duplicates are retained through the acquisition of novel functions (i.e., neofunctionalization) [30]. The precedent for subfunctionalization of tissue-specific NR expression after gene duplication has been established for several teleost NR subclasses, including NR3C, NR1B, and NR5A [31]. These examples illustrate the necessity of NR duplicates to retain defined ancestral expression patterns in conjunction with balanced signaling and regulatory pathways to maintain regulatory homeostasis.…”