As mentioned previously, the 100-amino acid forkhead-binding domain is well conserved across species and families. This domain is often exclusively used for phylogenetic analysis. Figure shows a neighbour-joining dendrogram of human and mouse forkhead domains. Nineteen subfamilies, denoted by different letters (A, B, C, etc.) can be distinguished based on evolutionary divergence. Note that FOXN proteins are split into two distinct subgroups, at the top and bottom of the dendrogram, and that the FOXL proteins are also split into different branches. Figure shows the same alignment using the full FOX protein sequence. Due to high divergence on either side of the forkhead domains, the full sequence is difficult to align between subfamilies, but has been helpful in defining each subfamily [7
]. In Figure , however, one again can see the splitting of the FOXN proteins and the FOXL proteins into distinctly separate branches. This global alignment also divides the FOXJ members into far different branches. One can conclude that alignment of the forkhead domain only provides a better assessment of evolutionary divergence.
Figure 1 Neighbour-joining dendrogram of the forkhead domains of all human and mouse FOX proteins. Protein sequences were aligned using ClustalW. Labels at branches indicate subclasses. Sequence information was downloaded from UniProt through the Hugo Gene (more ...)
Neighbour-joining dendrogram of the full sequences of all human and mouse FOX proteins. Sequences were aligned using ClustalW. Labels at branches indicate subfamilies.
Where possible, the nomenclature committee gave the same name to orthologues across species. Based on analysis of full sequences, mice have orthologues of all human FOX
genes with high sequence similarity, with one exception. The murine FOXD4 protein clusters together with seven human proteins -- FOXD4, and the FOXD4-like FOXD4L1 to FOXD4L6, and shares the most identity with FOXD4L1. The duplications that gave rise to the FOXD4 group appear to be relatively recent -- that is, during hominid evolution. Very little information is available about these proteins, but researchers have shown that at least two FOXD4L
genes are transcriptionally active; furthermore, evidence of purifying selection in the forkhead domain of these proteins suggests that they may play a physiological role [24
Many authors have performed detailed phylogenetic analyses, but the first analysis and naming scheme has been generally upheld,[4
] with a few criticisms illustrated in Figures and . In analysis of the forkhead domain, FOXR falls within the FOXN subclass and some authors have proposed combining these two groups. In analysis of the full sequence, however, FOXR1 and FOXR2 are associated more closely with FOXN1 and FOXN4, but not with FOXN2 and FOXN3. Thus, some researchers have proposed splitting these into three subclasses. Finally, in many analyses, FOXL1 and FOXL2 do not cluster together.
In an analysis of the origin and expansion of early transcription factors, it was found that FOXJ1 was probably the oldest family member, present in the opisthokont last common ancestor of the fungi and metazoans [25
]. Expansion occurred early, with bilaterians having 19 FOX
genes and most mammals more than 40.