RESUMEN
Chromosome 19 has the highest gene density of all human chromosomes, more than double the genome-wide average. The large clustered gene families, corresponding high G + C content, CpG islands and density of repetitive DNA indicate a chromosome rich in biological and evolutionary significance. Here we describe 55.8 million base pairs of highly accurate finished sequence representing 99.9% of the euchromatin portion of the chromosome. Manual curation of gene loci reveals 1,461 protein-coding genes and 321 pseudogenes. Among these are genes directly implicated in mendelian disorders, including familial hypercholesterolaemia and insulin-resistant diabetes. Nearly one-quarter of these genes belong to tandemly arranged families, encompassing more than 25% of the chromosome. Comparative analyses show a fascinating picture of conservation and divergence, revealing large blocks of gene orthology with rodents, scattered regions with more recent gene family expansions and deletions, and segments of coding and non-coding conservation with the distant fish species Takifugu.
Asunto(s)
Cromosomas Humanos Par 19/genética , Genes/genética , Mapeo Físico de Cromosoma , Empalme Alternativo/genética , Animales , Composición de Base , Secuencia Conservada/genética , Islas de CpG/genética , Evolución Molecular , Duplicación de Gen , Genética Médica , Humanos , Ratones , Datos de Secuencia Molecular , Familia de Multigenes/genética , Seudogenes/genética , Análisis de Secuencia de ADNRESUMEN
Allelic losses of the q13.3 region of chromosome 19 have been documented in malignant gliomas, neuroblastomas, and ovarian carcinomas, strongly suggesting the presence of a 19q13.3 tumor suppressor gene. Deletion mapping in tumors over the past decade has narrowed the candidate region considerably but has produced partially conflicting results, with some small candidate regions defined only by isolated tumors with deletions. Mutation and expression screening of genes from the most likely candidate regions has failed to identify the gene of interest, perhaps because of the conflicting deletion mapping data. The recently increased public availability of human genomic sequence, combined with improved bioinformatics capabilities, has now made it possible to map much larger candidate regions in considerable detail. We have manually generated a transcript map that spans most of the 19q13.3 tumor suppressor candidate region, from D19S219 to D19S246, with a resolution and quality superior to that of computer-generated maps. These results are presented in the hope that an improved map of the candidate region will facilitate further widespread screening and eventual identification of the gene or genes deleted in human gliomas, neuroblastomas, and ovarian cancers.
Asunto(s)
Cromosomas Humanos Par 19/genética , Genes Supresores de Tumor , Genoma Humano , Humanos , Mapeo Físico de Cromosoma , Transcripción GenéticaRESUMEN
Adaptins are major structural components of heterotetrameric protein complexes called adaptors, which are involved in intracellular receptor transport via clathrin-coated vesicles. In mice, one of these adaptins has been shown to be encoded by two genes, alpha(A)-adaptin and alpha(C)-adaptin, the former of which is expressed as two alternatively spliced transcripts. Using positional cloning gene approaches, we were able to identify the human alpha(A)-adaptin gene, which consists of 24 exons spanning over 40 kb on chromosome 19q13.3 between the loci of the R-ras gene and the polynucleotide kinase phosphatase gene. The novel gene encodes a 977 amino acid, 107.6 kDa protein with 98% amino acid sequence identity to its murine ortholog. Human alpha(A)-adaptin is expressed as a full-length transcript in forebrain, skeletal muscle, spinal cord, cerebellum, salivary gland, heart and colon. It is also ubiquitously expressed in tissues and in ZR-75-1 breast cancer cells and LNCaP prostate carcinoma cells as a smaller variant generated by splicing out of an exon encoding 22 amino acids in the hinge region of the protein.
Asunto(s)
Complejo 2 de Proteína Adaptadora , Proteínas Portadoras/genética , Proteínas de la Membrana/genética , Subunidades alfa de Complejo de Proteína Adaptadora , Empalme Alternativo , Secuencia de Aminoácidos , Secuencia de Bases , Cromosomas Humanos Par 19/genética , Clonación Molecular , ADN/química , ADN/genética , Exones , Femenino , Expresión Génica , Genes/genética , Humanos , Intrones , Masculino , Datos de Secuencia Molecular , Mapeo Físico de Cromosoma , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Células Tumorales CultivadasRESUMEN
Defining regulatory elements governing cell-restricted gene expression can be difficult because cis-elements may reside tens of kilobases away from start site(s) of transcription. Artificial chromosomes, which harbor hundreds of kilobases of genomic DNA, preserve a large sequence landscape containing most, if not all, regulatory elements controlling the expression of a particular gene. Here, we report on the use of a bacterial artificial chromosome (BAC) to begin understanding the in vivo regulation of smooth muscle calponin (SM-Calp). Long and accurate polymerase chain reaction, sequencing, and in silico analyses facilitated the complete sequence annotation of a BAC harboring human SM-Calp (hSM-Calp). RNase protection, in situ hybridization, Western blotting, and immunohistochemistry assays showed the BAC clone faithfully expressed hSM-Calp in both cultured cells and transgenic mice. Moreover, expression of hSM-Calp mirrored that of endogenous mouse SM-Calp suggesting that all cis-regulatory elements governing hSM-Calp expression in vivo were contained within the BAC. These BAC mice represent a new model system in which to systematically assess regulatory elements governing SM-Calp transcription in vivo.