Characterization of a novel gene, C21orf6, mapping to a critical region ofchromosome 21q22.1 involved in the monosomy 21 phenotype and of its murineortholog, orf5

Phenotypic and molecular analyses of patients with partial chromosome 21 mo nosomy enabled us to define a region, spanning 2.4 Rib between D21S190 and D21S226, associated with arthrogryposis, mental retardation, hypertonia, an d several facial anomalies. The markers of the region were used to screen a total human PAC library (Ioannou, RZPD). We isolated 57 PACs, which formed primary contigs. EST clusters (UNIGENE collection) located in a 6-Mb inter val, between D21S260 and D21S263, were mapped in individual bacterial clone s. We mapped the WI-17843 cluster to the PAC clone J12100, which contains t he two anchor markers LB10T and LA329. The open reading frame extends over 960 bp, with three putative start codons. The 1695-bp cDNA containing a pol yadenylation signal should correspond to the full-length cDNA. From the gen omic sequence, we deduced that the gene contained five exons and that there was a putative promoter sequence upstream from exon 1. In silico screening of DNA databases revealed similarity with a murine EST. The corresponding cDNA (1757 bp) sequence was very similar (>85%) to the human cDNA and had a n open reading frame of 876 nucleotides. Somatic hybrid mapping localized t he cDNA to mouse chromosome 16. EST analyses and RT-PCR indicated that the third exon in the human gene (exon 2 in the mouse) undergoes alternative sp licing. Northern blot hybridization showed that the gene was ubiquitously e xpressed in humans and mice. The longest mouse clone was used to generate r iboprobes, which were hybridized to murine embryos at stages E-9.5, E-10.5, E-12.5, E-13.5, and E-14.5-15, to study the pattern of expression during d evelopment. Ubiquitous labeling was observed, with strong signals restricte d to limited areas of the telencephalon, the mesencephalon, and the interrh ombomeric regions in the central nervous system, and other regions of the b ody such as the limb buds, branchial arches, and somites. (C) 2000 Academic Press.