A tyrosine-rich domain within homeodomain transcription factor Nkx2-5 is an essential element in the early cardiac transcriptional regulatory machinery

Abstract

Homeodomain factor Nkx2-5 is a central component of the transcription factor network that guides cardiac development; in humans, mutations in NKX2.5 lead to congenital heart disease (CHD). We have genetically defined a novel conserved tyrosine-rich domain (YRD) within Nkx2-5 that has co-evolved with its homeodomain. Mutation of the YRD did not affect DNA binding and only slightly diminished transcriptional activity of Nkx2-5 in a context-specific manner in vitro. However, the YRD was absolutely essential for the function of Nkx2-5 in cardiogenesis during ES cell differentiation and in the developing embryo. Furthermore, heterozygous mutation of all nine tyrosines to alanine created an allele with a strong dominant-negative-like activity in vivo: ES cell}embryo chimaeras bearing the heterozygous mutation died before term with cardiac malformations similar to the more severe anomalies seen in NKX2.5 mutant families. These studies suggest a functional interdependence between the NK2 class homeodomain and YRD in cardiac development and evolution, and establish a new model for analysis of Nkx2-5 function in CHD.

Somatic muscle development in Sepia officinalis (cephalopoda ‐ mollusca): A new role for NK4

“…After stage 20, SoNK4 ex-pression has been detected neither in branchial or systemic hearts nor gills. In a late stage of development (probably stage 26, personal communication), Elliott et al (2006) reported an expression of SoNK4 in the systemic heart but not in branchial hearts of Loligo paelii. These results confirm that expression of SoNK4 in gills and branchial hearts prospective areas is transient and restricted to the earlier stages of development in cephalopods.…”

“…Among them, NK4 is particularly interesting. This gene belongs to the NK gene cluster, which is probably the most ancient homeobox gene cluster, dating at least from the base of Metazoa (Larroux et al, 2007), and NK4 gene homologues have been identified in numerous species (Harvey, 1996;Elliott et al, 2006). Moreover, the main function of these genes seems to be highly conserved within Metazoa.…”

“…In Caenorhabditis elegans, the tinman homologue ceh-22 is involved in pharyngeal muscle differentiation (Okkema and Fire, 1994;Okkema et al, 1997); these muscles are responsible for intrinsic rhythmic contractions of the pharynx, and thus considered as analogs of vertebrate cardiac muscle (Pilon and Morck, 2005). In cephalopods, NK4 homologue expression in the systemic heart and the adjacent muscular ink sac of Loligo paelii has been pointed out in one late embryonic stage (Elliott et al, 2006). No data are available regarding the expression in earlier stages when muscle determination and differentiation occur.…”

Ciona intestinalis as a model for cardiac development

“…Sequence alignments demonstrate that the Ci-Nk4 homeodomain is highly conserved with those of vertebrate Nkx2 family genes (>90%). These alignments also indicate that Ci-Nk4 contains a N-terminal TN domain, linker and NK domains [42] along with a potential C-terminal tyrosine enriched domain or YRD [43]. Ci-Nk4 does not contain an Nkx2.5 box, but this domain is apparently specific to vertebrate orthologs (it is also not found in the Nkx2.5 ortholog from the invertebrate chordate, amphioxus, Amphi-Nk2Tin [26]).…”

“…More significantly, Ci-Nk4 lacks the highly conserved GIRAW domain that appears to mediate interaction with co-repressors in all other chordate Nkx2.5 orthologs. This domain is also absent in Drosophila tinman and other non-chordate orthologs [43].…”