Identification and functional validation of a 5' upstream regulatory sequence in the human tyrosinase gene homologous to the locus control region of the mouse tyrosinase gene

Comparison analysis of the sequences of the mouse and human genomes has proven a powerful approach in identifying functional regulatory elements within the non-coding regions that are conserved through evolution between homologous mammalian loci. Here, we applied computational analysis to identify regions of homology in the 5' upstream sequences of the human tyrosinase gene, similar to the locus control region (LCR) of the mouse tyrosinase gene, located at -15 kb. We detected several stretches of homology within the first 30 kb 5' tyrosinase gene upstream sequences of both species that include the proximal promoter sequences, the genomic region surrounding the mouse LCR, and further upstream segments. We cloned and sequenced a 5' upstream regulatory sequence found between -8 and -10 kb of the human tyrosinase locus (termed h5'URS) homologous to the mouse LCR sequences, and confirmed the presence of putative binding sites at -9 kb, homologous to those described in the mouse tyrosinase LCR core. Finally, we functionally validated the presence of a tissue-specific enhancer in the h5'URS by transient transfection analysis in human and mouse cells, as compared with homologous DNA sequences from the mouse tyrosinase locus. Future experiments in cells and transgenic animals will help us to understand the in vivo relevance of this newly described h5'URS sequence as a potentially important regulatory element for the correct expression of the human tyrosinase gene.     

Rapid evolution of human pseudoautosomal genes and their mouse homologs

Comparative studies of genes in the pseudoautosomal region (PAR) of human and mouse sex chromosomes have thus far been very limited. The only comparisons that can presently be made indicate that the PARs of humans and mice are not identical in terms of gene content. Here we describe additional comparative studies of human pseudoautosomal genes and their mouse homologs. Using a somatic cell hybrid mapping panel, we have assigned the mouse homolog of the human pseudoautosomal interleukin 3 receptor alpha subunit (IL3RA) gene to mouse Chromosome (Chr) 14. Attempts to clone the mouse homolog of the human pseudoautosomal adenine nucleotide translocase-3 (ANT3) gene resulted in the isolation of the murine homologs of the human ANT1 and ANT2 genes. The mouse Ant1 and Ant2 genes are very similar in sequence to their human homologs, and we have mapped them to mouse Chromosomes (Chrs) (8 and X respectively) that exhibit conserved synteny with the chromosomes on which the human genes are located. In contrast, the homolog of ANT3 appears to be either very divergent or absent from the mouse genome. Southern blot analysis of DNA from a variety of mammalian species shows restricted conservation of human pseudoautosomal genes, a trend that also applies to the two cloned mouse homologs of these genes and to neighboring human genes in distal Xp22.3. Our observations combined with those of other workers lead us to propose a model for the evolution of the PAR that includes both rapid sequence evolution and the incremental reduction in size of the region during mammalian evolution. Received: 4 May 1995 / Accepted: 21 August 1995     

Cloning, characterization, and tissue distribution of mouse phosphodiesterase 7A1

We have cloned a cDNA representing mouse phosphodiesterases (PDE) 7A1. The open reading frame encodes a protein of 482 amino acids with a predicted molecular mass of 55417. Like human PDE7A variants, mouse PDE7A1 and A2 are 5' splice variants from a common gene. The distinct N-terminal sequence of mouse PDE7A1 is highly homologous to the corresponding sequence of human PDE7A1 with a similarity of 98% but not to that of mouse PDE7A2 (with a similarity of 12%), and is more hydrophilic than that of mouse PDE7A2. Mouse PDE7A1 expressed in SF9 cells has been compared with human PDE7A1 under identical conditions. Mouse PDE7A1 has a Km for cAMP of 0.2 microM, an optimal pH of 7.5, an IC(50) value of 14 microM for 3-isobutyl-1-methylxanthine (IBMX), and is dependent on Mg(2+) for activity. All these characteristics are very similar to those of human PDE7A1. In mice, PDE7A1 is expressed in tissues of the immune system (lymph node, thymus, spleen, and blood leukocyte), testis, brain, kidney and lung but not in skeletal muscle, heart, embryo, or liver, while PDE7A2 is expressed in skeletal muscle, heart, embryo, and kidney, but not in the other tissues. This tissue distribution profile is very similar to that in humans, and hence suggests that PDE7A1 and 7A2 might play a similar role in different species.