Molecular cloning and characterization of the Fugu rubripes MEST/COPG2 imprinting cluster and chromosomal localization in Fugu and Tetraodon nigroviridis

We isolated Fugu genomic clones using the human MEST (<(Me)under bar>soderm-(S) under bar pecific (T) under bar ranscript) cDNA as probe. Sequence analysis revealed the presence of MEST and three additional genes which show homology to plant DNBP (<(DN)under bar>A-(B) under bar inding (P) under bar rotein), vertebrate COPG2 (<(Co)under bar>at (P) under bar rotein (G) under bar amma (2) under bar), as well as to human and mouse UCN ((U) under bar ro (c) under bar orti (n) under bar). Structures of Fugu and human MEST, COPG2 and UCN genes are very similar. Since MEST and COPG2 are neighboring genes on human chromosome 7q32, we can conclude that we identified their orthologs and that linkage of these genes is evolutionarily conserved in vertebrates. Unlike human MEST which underlies isoform-specific imprinting and is methylated in a parent-of-origin-specific fashion, the CpG island of the Fugu ortholog is completely methylated. The translation start of Fugu MEST is identical to the non-imprinted human isoform which is in good agreement with the assumption that genomic imprinting is restricted to mammals. Comparative mapping of these genes by fluorescence in-situ hybridization to metaphase chromosomes of Fugu rubripes and Tetraodon nigroviridis showed clear signals on one of the smallest acrocentric chromosomal pairs, which in Fugu, can be easily classified by its unique triangular shape.