RESUMO
We present the analysis of twenty human genomes to evaluate the prospects for identifying rare functional variants that contribute to a phenotype of interest. We sequenced at high coverage ten "case" genomes from individuals with severe hemophilia A and ten "control" genomes. We summarize the number of genetic variants emerging from a study of this magnitude, and provide a proof of concept for the identification of rare and highly-penetrant functional variants by confirming that the cause of hemophilia A is easily recognizable in this data set. We also show that the number of novel single nucleotide variants (SNVs) discovered per genome seems to stabilize at about 144,000 new variants per genome, after the first 15 individuals have been sequenced. Finally, we find that, on average, each genome carries 165 homozygous protein-truncating or stop loss variants in genes representing a diverse set of pathways.
Assuntos
Genoma Humano/genética , Análise de Sequência de DNA , Sequência de Bases , Estudos de Casos e Controles , Variações do Número de Cópias de DNA/genética , Bases de Dados Genéticas , Éxons/genética , Fator VIII/genética , Duplicação Gênica/genética , Técnicas de Inativação de Genes , Genética Populacional , Genótipo , Hemofilia A/genética , Humanos , Mutação INDEL/genética , Análise de Sequência com Séries de Oligonucleotídeos , Fases de Leitura Aberta/genética , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
The renal outer medullary potassium channel (ROMK) is expressed in the kidney tubule and critically regulates sodium and potassium balance. The physiological functions of other inward rectifying K(+) (Kir) channels expressed in the nephron, such as Kir7.1, are less well understood in part due to the lack of selective pharmacological probes targeting inward rectifiers. In an effort to identify Kir channel probes, we performed a fluorescence-based, high-throughput screen (HTS) of 126,009 small molecules for modulators of ROMK function. Several antagonists were identified in the screen. One compound, termed VU590, inhibits ROMK with submicromolar affinity, but has no effect on Kir2.1 or Kir4.1. Low micromolar concentrations inhibit Kir7.1, making VU590 the first small-molecule inhibitor of Kir7.1. Structure-activity relationships of VU590 were defined using small-scale parallel synthesis. Electrophysiological analysis indicates that VU590 is an intracellular pore blocker. VU590 and other compounds identified by HTS will be instrumental in defining Kir channel structure, physiology, and therapeutic potential.