Chromosome evolution

(supported by grant Ha1374/5-2 from the Deutsche Forschungsgemeinschaft)

Evolutionary chromosome rearrangements have played a predominant role in shaping the genomes of existing vertebrates. Comparative (Zoo-)FISH mapping of complex human DNA probes on chromosomes of heterologous mammalian species can visualize interspecific homologous chromosome segments and determine ancestral chromosomal segments or gene arrangements (Grützner et al. 1999a; Sharon et al. 1999). We perform cytogenetic and molecular searches for evolutionary chromosome breakpoints which have occurred during primate, in particular hominoid, evolution. Large-insert (YAC, BAC, PAC and cosmid) clones spanning a particular breakpoint are easily identified by FISH because they produce "split" hybridization signals on the rearranged ape or monkey chromosome(s). When the breakpoint has been narrowed down to a relatively small intervall (of approximately 100 kb), this region will be analyzed in detail by exon trapping, hybridization to cDNA libraries, automated sequencing, and computer-assisted gene searches, to study whether evolutionary breaks affect the structure of genes. It is possible that evolutionary chromosome rearrangements interfere with genome function either directly, by disrupting a gene(s), or indirectly, by (in)activating closely juxtaposed genes (position effects). Evolutionary rearrangements could also disrupt imprinted chromosome regions and thus change the imprinting pattern of specific genes. Examination of the structural and functional features of evolutionary breakpoints and comparison with pathological human chromosome rearrangements will provide new insights as to whether particular DNA sequences or structural features are predisposed to chromosome breakage and whether chromosome pathology provides a momentary snapshot in the ongoing process of evolution.

Comparative mapping of individual gene loci (instead of complex human DNA probes) allows the generation of homology maps in distantly related species, such as birds and fishes. Thus far, we have found conserved synteny between the chicken Z sex chromosome and human chromosome 9pter-q22 (Nanda et al. 1999, 2000). Based on this observation, we have identified the chicken orthologue of a human DM-domain gene expressed in the testis, DMRT1, on the chicken Z (Shan et al. 2000a). We are studying whether DMRT1, which has been implicated in XY sex reversal in humans (Shan et al. 2000b), plays a role in male sexual differentiation in birds. It is conceivable to assume that two copies of DMRT1 are required for testis formation, whereas a single copy along with the W chromosome leads to female sexual differentiation. Although sex determination has undergone considerable evolutionary changes, regulatory genes such as DMRT1 appear to be functionally highly conserved.

Because of its highly compact genome, the pufferfish has become an important animal model in genome research. Despite the small size of their chromosomes, we have established both classical and molecular cytogenetic techniques in two pufferfish species, Fugu rubripes and Tetraodon nigroviridis (Grützner et al. 1999b, Brunner et al. 2000). In order to find linkage groups which are conserved between pufferfish and humans, we hybridize pufferfish BACs, which share orthologous gene sequences with humans (collaboration with H. Roest-Crollius and J. Weissenbach, Genoscope, Evry, France), on pufferfish chromosomes. In a pilot study focusing on genes from human chromosome 9 and X, we found a surprisingly high degree of conserved chromosomal synteny between pufferfish and humans, which diverged more than 400 million years ago. Evolutionary conservation appears to be an intrinsic chromosomal property and can dramatically differ between chromosomes or chromosome regions. By mapping several hundred of these pufferfish clones with homologs distributed throughout the entire human genome, we will establish a first-generation homology map between pufferfish and humans.