PROXIMAL CIS-ACTING ELEMENTS COOPERATE TO SET HOXB-7 (HOX-2.3) EXPRESSION BOUNDARIES IN TRANSGENIC MICE

The Hox genes have been proved to be instrumental in establishing the positional identity of cells along the embryonic anteroposterior (A-P) axis. Studying the regulation of these genes is a first step toward e lucidating the molecular basis of regionalization during embryogenesis . We report here on the identification of cis-acting elements controll ing the expression of Hoxb-7 (Hox-2.3). We show that elements driving A-P restricted gene expression are located within the 3.5 kb proximal upstream sequences of the Hoxb-7 gene. A deletion analysis provides ev idence for at least three cis-acting control elements upstream from Ho xb-7, and for cooperative interactions between some of these elements in generating the A-P restricted transgenic pattern. One element, conf erring by itself Hox-like expression boundaries to the transgene, has been studied in more detail and found to act in an orientation-and pro moter-dependent manner. Together the 3.5 kb sequences proximal to Hoxb -7 mediate A-P restricted Hoxb-7/lacZ gene expression in a domain show ing rostral boundaries more posterior than those of Hoxb-7. The evolut ion throughout embryogenesis of the expression pattern of a transgene carrying these sequences has been analysed and shown to mimick that of the endogenous gene, except for a slight delay in the initial express ion. We conclude that the transgenes that we tested, spanning a total of 27 kb genomic sequences, do not reproduce all the features of the H oxb-7 expression pattern. The differences in expression between Hoxb-7 and the transgenes may reveal an aspect of the Hox regulation for whi ch either remote cis-acting control elements and/or gene clustering is required. Additional features that may have favoured maintenance of c lustered organisation during evolution are partial overlap of transcri ption units with the regulatory regions of the neighbouring genes, and cis-regulatory interactions between multiple Hox genes: not only do c is-acting control elements of the Hoxb-7 gene map in the 3' untranslat ed sequences of the Hoxb-8 (Hox-2.4) gene, but our experiments suggest that Hoxb-7 control sequences modulate expression of the Hoxb-8 gene as well.