Hox genes working in association with segmentation genes
How transcription factors achieve their in vivo specificities is a fundamental question in biology. Regulation of Distalless (dll) gene in insects served as an excellent model to study mechanisms of Hox genes and this is the topic of our classic paper today. Richard mann’s lab published some stunning stuff on role played by segmentation genes in regulation of distalless gene in 2004 Nature issue. This article is a real classic providing some finer details in repression of distalless in abdomen of insects. Ever since the first paper by Vachon et al in Cell ,1992 describing the direct role played by Hox genes in turning off distalless activation in abdomen ,many labs provided finer details concerning the exact mechanism and Richard mann’s lab contributed a significant deal. In fact it was this work proved for the first time that that Hox factors directly interact with segmentation proteins such as En to control gene expression against previous notion that that anterior–posterior compartmentalization (by Hox genes) and segment identity specification are independent processes.
Distalless (Dll) is limb primodia throughout animal kingdom ,which is a bonafide Hox target gene and its role in drosophila leg development is quite well studied. Dll is expressed in three thoracic segments but repressed in abdomen. 
A cis regulatory element called DMX derived from dll gene drives accurate dll likes expression in thorax. The abdominal Hox genes Ultrabithorax (Ubx) and abdominalA (abdA) directly repress Dll , thereby blocking leg development in the abdomen.
DMX is composed of a large activator element (DMXact) and a 57-base-pair (bp) repressor element . It was again the work from mann’s lab that shown previously that that Ubx and AbdA bind to DMX in a co operative manner with some help from two homeodomain cofactors, Extradenticle (Exd) and Homothorax (Hth). In contrast, the thoracic Hox protein Antennapedia (Antp) does not repress Dll and does not bind DMX-R with high affinity in the presence or absence of Exd and Hth, that’s the reason for presence of legs in thorax and not in abdomen.
One view in regulation of distalless depends partly on the ability of these cofactors(exd -hth) to selectively enhance the binding of the abdominal Hox proteins to DMX-R. However these two classical cofactors are involved in both activation and repression of terget genes by Hox. So it can be said that these complexes do not directly recruit co-activators or co-repressors, but instead are required for target gene selection and hence there might be some other sequences adjacent to Hox-exd-hth sequences ,which determines whether a gene will be activated or repressed.Richard mann’s lab then looked into the DMX-R region from 6 drosophila species and found some nice conservation outside the known Hox-exd-hth suggesting these sequences might be essential for regulation.
To gain more insights into the role played by these sequences, Brian Gebelein et al, performed a
thorough mutagenesis of DMX-R and each mutant was tested for activity in a standard reporter gene assay in transgenic embryos. Surprisingly some mutants show specific de repression only in anterior or posterior segments only. Engrailed is well known segmentation gene which gives posterior identity to the cells ,forms nice complexes with Abd A Hox gene in vitro and de repression of DMX-R enhancer can be seen in posterior specific cells by mutating the engrailed consensus sequence. The transgene which shows anterior specific de repression bears mutation in the sequence which resembles consensus binding site for Forkhead (Fkh) domain proteins. Sloppy paired 1 and 2 are examples of a Fkh domain factor and they are expressed in in anterior compartment cells adjacent and anterior to En-expressing posterior compartment cells. The transgenic fly with mutations of both engrailed and sloppy paired shows maximum de repression and spans both compartments.
After analysis of various genetic and gel shifts experiments authors came up with model of dll regulation,where two bithorax genes ubx and abdA bind directly to dll regulatory region and brings in the repression in a compartment wise manner in abdomen.
Well known hox co factors exd and hth are crucial part of this complex . The compartment wise repression of dll is possible by involvement of two segmentation genes engrailed and sloppy paired.
Ubx in association with slp ,exd and hth binds to their respective binding sites on DMXR and thus shuts off dll in anterior compartment and Engrailed the posterior specific segmentation gene along with exd and hth helps out abdA to repress dll in posterior compartment. In this way ubx and adbA repress dll in abdomen.
This is an excellent article in the field of hox function elucidating direct involvement of segmentation genes in hox regulation of target genes.
Reference:
Homeotic genes of the Bithorax complex repress limb development in the abdomen of the Drosophila embryo through the target gene Distal-less.
Vachon G, Cohen B, Pfeifle C, McGuffin ME, Botas J, Cohen SM.
Cell. 1992 Oct 30;71(3):437-50.
Specificity of Distalless repression and limb primordia development by abdominal Hox proteins.
Gebelein B, Culi J, Ryoo HD, Zhang W, Mann RS.
Dev Cell. 2002 Oct;3(4):487-98.
Direct integration of Hox and segmentation gene inputs during Drosophila development.
Gebelein B, McKay DJ, Mann RS.
Nature. 2004 Oct 7;431(7009):653-9.
Link to Richard mann’s lab
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