The homoebox-containing genes of the Hox-5 complex are expressed in different but overlapping domains in limbs during murine development. The more 5' the position of these genes in the complex, the later and more distal is their expression. Antero-posterior differences are also observed. A model is proposed that accounts for the establishment of these expression domains in relation to the existence of a morphogen released by the zone of polarizing activity. Comparison of these observations with the expression patterns of the genes of Hox complexes in the early embryo suggests that similar molecular mechanisms are involved in the positional signalling along the axes of both the embryonic trunk and the fetal limbs.

The cloning, characterization and developmental expression patterns of two novel murine Hox genes, Hox4.6 and Hox4.7, are reported. Structural data allow us to classify the four Hox4 genes located in the most upstream (5') position in the HOX4 complex as members of a large family of homeogenes related to the Drosophila homeotic gene Abdominal B (AbdB). It therefore appears that these vertebrate genes are derived from a selective amplification of an ancestral gene which gave rise, during evolution, to the most posterior of the insect homeotic genes so far described. In agreement with the structural colinearity, these genes have very posteriorly restricted expression profiles. In addition, their developmental expression is temporally regulated according to a cranio-caudal sequence which parallels the physical ordering of these genes along the chromosome. We discuss the phylogenetic alternative in the evolution of genetic complexity by amplifying either genes or regulatory sequences, as exemplified by this system in the mouse and Drosophila. Furthermore, the possible role of 'temporal colinearity' in the ontogeny of all coelomic (metamerized) metazoans showing a temporal anteroposterior morphogenetic progression is addressed.

To cure the fireblight pathogen Erwinia amylovora of a 29 kb plasmid (pEA29) that is common in strains of these bacteria, restriction fragments of this plasmid were inserted into plasmids based on replication functions of bacteriophage fd, which cannot replicate in bacteria without expression of viral gene 2-protein. A 4.4 kb PstI fragment including the unique BamHI site of pEA29 allowed pfd plasmids to be propagated in E. amylovora. Furthermore, selection for these plasmids removed the natural 29 kb plasmid, apparently because of plasmid incompatibility. The pfd plasmids with the 4.4 kb insertion had a high tendency to segregate in E. amylovora when grown without selective pressure. Plasmid-free E. amylovora strains were virulent in standard tests on slices of immature pears, but symptom development was delayed compared to the wild-type. When assayed on pear seedlings a deficiency in pathogenicity was observed. Furthermore, strains without pEA29 spread more slowly on a lawn of pear cells than did wildtype strains. Plasmid-free strains of E. amylovora were not auxotrophic, but synthesis of extracellular polysaccharide was altered under certain growth conditions.

All strains of Erwinia amylovora characterized carry a medium-size plasmid of 29 kilobases (pEA29). We mapped this plasmid with various restriction enzymes, cloned the whole DNA into an Escherichia coli plasmid, and subcloned restriction fragments. These DNA species were used for identification of E. amylovora after handling of strains in the laboratory and also in field isolates. About 70 strains of E. amylovora and 24 strains from nine other species, mainly found in plant habitats, were checked in a colony hybridization test. Virulent and avirulent E. amylovora strains reacted positively, whereas the other species were negative. Apart from the hybridization assay, the positive strains were additionally tested for ooze production on rich agar with 5% sucrose and on immature-pear slices. Unspecific background hybridization of nonE. amylovora strains found for hybridization with the whole E. amylovora plasmid was almost eliminated when a 5-kilobase Sail fragment from pEA29 was used as a probe and when the washes after the hybridization procedure were done with high stringency. Under these conditions, E. amylovora could be readily identified from field isolates.

We report the cloning, genomic localization, primary structure and developmental expression pattern of the novel mouse Hox-4.3 gene. This gene is located within the HOX-4(5) complex, at a position which classifies it as a member of the Hox-3.1 and -2.4 subfamily, the DNA and predicted protein sequences further confirmed this classification. Hox-4.3 has a primary structure characteristic of a Hox gene but, in addition, contains several monotonic stretches of amino acids, one of the ‘paired’-like type. As expected from its presence and position within the complex. Hox-4.3 is developmentally expressed in structures of either mesodermal or neurectodermal origin located or derived from below a precise craniocaudal level. However, a very important offset between anteroposterior boundaries within neuroectoderm versus mesoderm derivatives is observed. Like other genes of the HOX-4(5) complex, Hox-4.3 is expressed in developing limbs and gonads, suggesting that ‘cluster specificity’ could be a feature of the HOX network.