I have a big soft spot for Hox genes, or rather, Hox proteins. Thanks to some of my earlier work, I also have a soft spot for all their secret little sequence motifs that help them interact with other proteins and help us classify them (e. g. Balavoine et al., 2002). Probably the best-known such motif is the hexapeptide. (“Hexa-” would kind of imply that it’s made of six amino acids, but people only ever seem to talk about four. Don’t ask me why they call it a hexapeptide…)
This motif is very widespread, occurring not just in Hox proteins but also in many others in the larger class of homeodomain proteins that Hoxes belong to. For many years, the hexapeptide has been regarded as the key to the interaction of Hox proteins with another homeodomain-bearing protein, called Extradenticle in flies and Pbx plus a number (we have 3 of them) in vertebrates*. (Above is a cartoon version of DNA with the homeodomains – the purple curls – of Exd and the fly Hox protein Ubx bound to it, from the Protein Data Bank.) Hox proteins bind DNA to regulate various genes, and are absolutely vital for an embryo to develop the right organs in the right places. This interaction changes their DNA binding behaviour, making the hexapeptide possibly the most important four amino acids in animal development.
And now we’re supposed to scrap that?
I’ve just skimmed through Hudry et al. (2012), and died a little inside.
The study claims – on what seems to be good evidence – that the hexapeptide is not all it’s cracked up to be. Out of six fruit fly Hox proteins examined, only two stop interacting with Exd when the hexapeptide is mutated beyond recognition, and even then one of them is kind of half-hearted about it. The team also tested a few mouse Hox genes in cultured cells and – for whatever reason – chick embryos, and got largely the same results.
I rather like their approach, though. I think the method for detecting interaction is incredibly clever, though it’s clearly not something they invented. The idea is based on fluorescent proteins. These are very commonly used to track the levels and whereabouts of other proteins. Since they are pretty small and innocuous, the gene encoding them can be tacked onto the gene of interest, and the resulting protein chimaera will do whatever the target protein would do without its fluorescent companion. The only difference is now it glows wherever it goes. The more protein, the brighter the glow.
The nice thing about fluorescent proteins is that you can cut them in half, and if the two parts get close enough, they’ll still glow. Therefore, if you glue one half of the DNA for a fluorescent protein to gene 1, the other half to gene 2, and let them loose in the same cell, you can tell whether the protein products of gene 1 and gene 2 interact just by looking for the telltale fluorescence. And the tale it’s telling is that all these Hox proteins are getting snug with Exd despite the loss of the motif supposedly necessary for the interaction.
I’ll just go away and quietly get over that now.
*Vertebrate geneticists have no imagination. Okay, they did come up with lunatic fringe and Sonic hedgehog. After the fly people named fringe and hedgehog.
Balavoine G et al. (2002) Hox clusters and bilaterian phylogeny. Molecular Phylogenetics and Evolution 24:366-373
Hudry B et al. (2012) Hox proteins display a common and ancestral ability to diversify their interaction mode with the PBC class cofactors. PLoS Biology 10:e1001351