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Hox genes have been shown to play a key role in the acquisition of positional identity by precursors of embryonic axial, paraxial and limb structures. This function is thought to depend on the sequential, concerted expression of these genes in time and space. However the underlying molecular mechanisms of this collinear expression are still largely unknown. So far we had identified proximal regulatory elements driving expression of Hoxb-8/LacZ transgenes in Hox-like expression patterns with rostral boundaries more posterior than those of the endogenous gene. In this work we have analyzed 30 kb of 3' genomic sequences for Hoxb-8 regulatory activity in transgenic mice. We have identified a control region in the Hoxb-5/b-4 intergenic region that rostrally extends the Hoxb-8/LacZ expression domain into the posterior hindbrain. In combination with the Hoxb-8 minimal promoter, the 3' control region drives transgene expression with boundaries more anterior than those of Hoxb-8 in the neural tube. When combined with a 4.5 kb Hoxb-8 upstream sequence, where essential proximal regulatory sequences are located, the 3' control region drives transgene expression in a domain which seems to correspond to that of the endogenous Hoxb-8. By deletion analysis we have narrowed down to 550bp the regulatory activity interacting with the Hoxb-8 minimal promoter. We discuss the possibility that this remote 3' enhancer, which is the closest regulatory region found in the cluster to rostrally extend Hoxb-8/LacZ expression, could be involved in the regulation of Hoxb-8 and interact with the proximal control elements.

A 3' remote control region is a candidate to modulate Hoxb-8 expression boundaries.