That Was Then, This Is Now

Charles Parker, MD

2010-07-01

Dr. Parker indicated no relevant conflicts of interest.

Roach JC, Glusman G, Smit AF, et al. Analysis of genetic inheritance in a family quartet by whole-genome sequencing. Science. 2010;328:636-639.

In the not-so-distant future, clinicians may order whole-genome sequencing of patient DNA the way we now order an MRI. In the run-up to the routine clinical use of whole-genome sequencing, investigators are taking advantage of this remarkable technical achievement to characterize the human genome and to solve challenging genetic and pathophysiological problems.1,2 In a recent publication in Science, the power and beauty of whole-genome sequencing was illustrated by the ingenious studies of a collaborative group of scientists led by David Galas and Leroy Hood from the Institute for Systems Biology in Seattle and Lynn Jorde from the University of Utah. The problem to be solved by the study (to identify the genetic basis of a rare autosomal recessive disorder) was straightforward, but the experimental design was inspired and the studies were made possible by the generosity and selflessness of the involved family. Roach, Glusman, Smit, and Huff, who shared first authorship on the paper, and colleagues identified a pedigree that consisted of a mother, a father, and two siblings, both of whom were affected by two distinct autosomal recessive disorders, Miller syndrome (postaxial acrofacial dysostosis) and primary ciliary dyskinesia. Sequencing the genome of the parents allowed the investigators to delineate precisely sites of recombination, and the results illustrate the remarkable genetic diversity that exists between siblings (Figure). The sibling pair was found to be only ~20 percent genetically identical, and the investigators took advantage of this finding to narrow their search for the disease-causing mutations (Figure). Reasoning that rare phenotypes would likely be encoded by rare nucleotide sequence variants allowed the investigators to further narrow the search for the disease-causing mutations. Data analysis identified sequence variants that fit the model of either simple recessive (a gene mutated at the same position in both alleles) or compound heterozygous (mutation of the same gene at different nucleotide positions in each of the two alleles) in four candidate genes. By comparing core exome sequencing of two unrelated individuals with Miller syndrome, the disease-causing mutation for Miller syndrome in the sibling pair was confirmed to be the consequence of a compound heterozygous state affecting dihydroorotate dehydrogenase (DHODH), a gene that encodes a key enzyme in pyrimidine biosynthesis. The gene (DNAH5 that encodes a dynein protein that functions as a force-generating protein with ATPase activity) that is mutated in primary ciliary dyskinesia was previously known, and the studies by Roach et al. confirmed that the gene was mutated in a compound heterozygous state in the two affected siblings. As such, primary ciliary dyskinesia served as a positive control for the studies. In the process of discovering the genetic mutation that causes Miller syndrome and confirming the genetic basis of primary ciliary dyskinesia, Roach and colleagues reported a number of other interesting, basic observations including the following:

  • Recombination in maternal meioses occurs ~1.7 times more frequently than in paternal meioses (98 maternal events compared to 57 paternal events).
  • 92 of the 155 recombinations took place in a known genetic hotspot.
  • There are ~70 mutations/diploid genome, resulting in a mutation rate of ~1.1 x 10-8 per position per haploid genome.
  • The mutational frequency at CpG sites (sites of genomic methylation) is ~11 times greater than at other sites.
  • 323,255 novel single nucleotide polymorphisms were identified.

Deep sequencing of the pedigree reveals the extensive nature of genetic recombination - The illustrated chromosome is vertically split to depict the inheritance state from the father (P, left half) and the mother (M, right half) based on nucleotide sequencing of the genomes of the parents and the two children in the pedigree. Mendelian inheritance patterns can be grouped into four states for each polymorphic nucleotide position with children receiving one of the following: a) the same allele from both the father and the mother (identical); b) the same allele from the mother but the opposite allele from the father (haploidentical maternal); c) the same allele from the father but opposite from the mother (haploidentical paternal); and d) opposites from both parents (non-identical). In the illustration, the pattern of inheritance is shown in blocks, with the dark blocks representing regions of recombination that occurred in the parental germ line during meiosis. In areas where a light blue segment abuts a dark blue segment, the siblings are haploidentical (paternal or maternal). In areas where a dark blue segment abuts a dark blue segment, the siblings are non-identical. Only in those regions in which light blue portions of the schematic karyotype are adjacent are the siblings effectively twins. Overall, 22 percent of the siblings’ genomes were found to be identical. This finding markedly narrowed the area that needed to be searched for the disease-causing mutations (red horizontal lines), as a recessive inheritance pattern was suggested to account for both Miller syndrome and primary ciliary dyskinesia.
Click image to enlarge

Recalling the progress that has ensued since the National Center for Human Genome Research was established in 1990 with a goal of sequencing the human genome in 15 years, the studies of Roach and colleagues bring to mind a quote from Arthur C. Clarke: “Any sufficiently advanced technology is indistinguishable from magic.”

  1. Lupski JR, Reid JG, Gonzaga-Jauregui C, et al. Whole-genome sequencing in a patient with Charcot-Marie-Tooth neuropathy. N Engl J Med. 2010;362:1181-1191.
  2. Mardis ER, Ding L, Dooling DJ, et al. Recurring mutations found by sequencing an acute myeloid leukemia genome. N Engl J Med. 2009;361:1058-1066.

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