Sequencing Study Highlights Genetic Diversity in Southern Africa

26/04/2010 às 13:30 (Genotipagem de SNPs, GWAS, hapmap, Sequenciamento)

By a GenomeWeb staff reporter

NEW YORK (GenomeWeb News) – An international research team reported in Nature today that it has characterized five human genomes from southern Africa, identifying millions of SNPs never before found in the human population.

The American, African, and Australian researchers sequenced the full genomes of two African individuals: a member of a hunter-gatherer population in the Kalahari desert known as the Bushmen, San, or Khoisan, and a Bantu individual from South Africa — Nobel peace prize winner Archbishop Desmond Tutu. After sequencing the exomes of three other Khoisan men, the team compared all five genomes, identifying more than 1.3 million previously undetected SNPs.

During their subsequent analyses, they not only found genetic differences between southern African populations and populations from other parts of Africa and the world but also within the Khoisan population — findings that may eventually inform everything from studies of human population history and adaptations to agriculture to personalized medicine strategies in southern Africa.

“On average, there are more genetic differences between any two Bushmen in our study than between a European and an Asian,” co-lead author Stephan Schuster, a biochemistry and molecular biology researcher with Pennsylvania State University’s Center for Comparative Genomics and Bioinformatics, said in a statement.

Southern Africa is believed to be the source of modern humans and, subsequently, is home to a great deal of human genetic diversity. But despite the decades-long effort to characterize the human genome and human population genetics, most studies have lacked representatives from this region, Schuster explained during a telephone briefing with reporters this morning.

In an effort to get a better sense of the genetic variation within humans, he and his team set out to characterize the genomes of individuals from the Khoisan population — thought to be the oldest modern human population. Schuster described the project at the American Society for Human Genetics meeting last fall, though this paper marks the first publication from the sequencing effort.

Archbishop Tutu, who has ancestry from Sotho-Tswana and Nguni language groups, which represent roughly 90 percent of southern Africans, also participated in the study. Tutu was a good candidate not only because of his ancestry but also because he is known to have survived polio, tuberculosis, and prostate cancer and because he is a voice for southern Africa and indigenous populations, senior author Vanessa Hayes, a cancer genetics researcher at the University of New South Wales, told reporters.

The four Khoisan men who participated in the study all came from different communities in Namibia’s Kalahari Desert. Each was the most elder member of his community.

The team sequenced the genome of a Khoisan man named named !Gubi to 10.2 times coverage using paired-end sequencing with the Roche 454 GS FLX Titanium platform. !Gubi is believed to be around 86 years old and lives in the southern Kalahari on the Namibia-Botswana border. They also used a similar approach to sequence the genome of a second Khoisan man named G/aq’o, from a community in the northern Kalahari, to about two times coverage.

Meanwhile, the researchers sequenced Tutu’s genome using the Applied Biosystems SOLiD 3.0 platform, generating sequence covering the genome about 12.3 times.

For the exome sequencing portion of the study, the team captured protein-coding sequences for each of the five individuals with the NimbleGen 2.1 M array and sequenced them by Roche 454 Titanium sequencing.

The genomic and exomic sequences were verified using a range of approaches, including genotyping and whole-genome and exome sequencing with the Illumina platform, which was used to sequence !Gubi’s genome to 23.2 times coverage and Tutu’s genome to 7.2 times coverage.

When the team compared the genomes and exomes to version 18 of the human reference genome and eight personal genomes sequenced, they found 1.3 million previously undetected SNPs, including 13,146 new SNPs that alter the amino acid sequence of 7,720 genes.

!Gubi’s genome contained more SNPs than Tutu’s, though both contained more SNPs overall — and more novel SNPs — than any other individual genome sequenced so far.

And from their population level analyses, the researchers detected as many or more genetic differences between the Khoisan and West African populations than between West African and European populations.

The team’s preliminary peek at the functional role of genes affected by new SNPs in the Khoisan population suggests that these variants tend to fall in genes involved in immune response, reproduction, and sensory perception.

“We believed that because of their extremely long lineage, their genome would be very different,” co-lead author Webb Miller, a researcher at Penn State’s Center for Comparative Genomics and Bioinformatics, told reporters. And, he said, the findings so far support that hypothesis.

This type of genetic diversity within the human genome is believed to have helped humans thrive over thousands of years, Schuster said, though he emphasized that modern human genomes from all around the world still share far more similarities than differences. “We are genetically one healthy species,” he said.

The team believes understanding human genetic diversity in southern Africa will likely be medically important, both for developing personalized medicine in this region and for identifying and understanding the roles of rare variants in human health and disease in general.

“Adding the described variants to current databases will facilitate the inclusion of southern Africans in medical researchers’ efforts, particularly when family and medical histories can be correlated with genome-wide data,” the researchers wrote.

The researchers have already started developing microarrays incorporating the newly identified southern African SNPs. For the next phase of the study, they plan to use these microarrays to genotype hundreds of individuals from southern Africa.

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A Novel Method for SNP Identification

26/04/2010 às 13:28 (Diagnóstico Molecular, Genotipagem de SNPs, GWAS)

In an advance online publication of Genome Research,

research out of the Scripps Translational Science Institute describes a novel method for SNP identification, “SNIP-Seq.” SNIP-Seq utilizes population sequence data to detect SNPs and assign genotypes to individuals. The team used data from a region on chromosome 9p21 of the human genome (sequenced in 48 individuals, with five sequenced in duplicate) and found that many of the novel SNPs identified by SNIP-Seq were validated by pooled sequencing data; they were also confirmed by Sanger sequencing. “Collectively, these results suggest that analysis of population sequencing data is a powerful approach for the accurate detection of SNPs and the assignment of genotypes to individual samples,” the team writes.

Also published in advance online this week, Wilfried Haerty and G. Brian Golding of McMaster University, Ontario, describe their discovery of genome-wide evidence for selection acting on single amino acid repeats. Haerty and Golding tested the effect of splicing on the structure of homopolymer sequences. The team discerned a “relationship between alternative splicing and homopolymer sequences with alternatively spliced genes being enriched in number and length of homopolymer sequences.” They also found lower codon density and longer homocodons, which they say suggests a balance connected with the pressures imposed by selection.

This week in Genome Research, researchers at Harvard and MIT propose an improved method for identifying gene interactions using high-dimensional single-cell morphological data from genetic screens, applied in a systematic computational model to RhoGAP/GTPase regulation in Drosophila melanogaster. The team writes that while their model appears to create only mediocre predictions, it represents a vast improvement from alternative methods. “This work demonstrates the fundamental fact that high-throughput morphological data can be used in a systematic, successful fashion to identify genetic interactions and, using additional elementary knowledge of network structure, to infer signaling relations,” they write.

In a methods paper, Andrew Young of the National Human Genome Research Institute and his colleagues describe a novel strategy for de novo genomic assemblies using short sequence reads and reduced representation libraries. Young et al. developed a method to partition the genome prior to assembly by using two independent restriction enzymes to create overlapping fragment libraries ― each containing a manageable subset of the genome. “Together, these libraries allow us to reassemble the entire genome without the need of a reference sequence,” the team writes. In a proof-of-concept study, the team applied their method in assembling the Drosophila genome, and when compared with the reference genome, they deemed their version significantly comparable.

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Q&A: JCVI’s New Rockville Campus Director Karen Nelson on Current and Future Sequencing Projects

08/01/2010 às 19:17 (Expressão Gênica, GWAS, Medicina, Sequenciamento, Solid, whole transcriptome)

Name: Karen Nelson
Age: 45
Position: Director of the J. Craig Venter Institute Rockville campus, since 2009
Experience and Education:
Other positions, JCVI, since 1996
PhD in microbiology, Cornell University, 1996
MS in animal science, University of Florida, Gainesville, 1992
BS in animal science, University of the West Indies, Trinidad and Tobago, 1987

Karen Nelson was recently named head of the J. Craig Venter Institute’s Rockville, Md., campus. She succeeds Robert Strausberg, who is moving to the Ludwig Institute for Cancer Research as director for collaborative sciences.

Nelson has been with JCVI since 1996, when she came on board as a postdoctoral fellow and the center was still known as the Institute for Genomic Research. She was involved with the institute’s early metagenomic projects, including its first metagenomic analysis, published in 2006, of the microbial diversity found in human fecal matter.

Nelson also led the team that sequenced the bacterium Thermotoga maritime, which lives in 175 ºF water. She eventually became the director of human microbiology and metagenomics in the department of human medicine and genomics at JCVI and has played a large role in the human microbiome project, which aims to characterize and analyze the entire human microbiota.

Recently, Nelson spoke with In Sequence about the future of JCVI’s research and how new sequencing technologies are expanding the scope of what researchers there are able to study.

What are your plans and what projects are you involved in as the new head of JCVI’s Rockville campus?

My training is as a microbiologist, so I’m very excited about looking at the areas of metagenomics, and looking at all the microbial species that we have not been able to culture but now we can access with these new sequencing technologies.

We have a very strong infrastructure in terms of sequencing and data analysis. We have a very strong informatics team involved in the annotation and comparative analysis of these large data sets. So I expect that we will continue to play a major role in terms of the metagenomic analyses of not only the human body but a number of other environments.

We have collaborations looking at the metagenomics of domesticated and wild animals, and we have a huge pipeline analysis of viruses, particularly flu, that’s funded by the NIAID. We’re looking at viruses in the environment and how viruses hop from their natural reservoir into humans.

On the environmental side, we were approached by the University of the West Indies to go to Trinidad and start sampling some of the areas where they have oil reservoirs, to use microbial-based approaches and metagenomic-based approaches to help look at the methane cycle in cleaning up the environment.

So, we have a number of very large-scale studies — both single genomes and large metagenomic studies — that I expect will continue to expand and diversify.

Will you be going in any new directions in the coming years?

I’m hoping to take advantage of the new sequencing technologies, and the whole area that is allowing us to generate larger data sets. We have the tools to analyze the data, but now we can actually generate the data at a reduced cost and in much larger quantities than you could imagine before.

Going beyond gene sequencing, I’m also looking at the transcriptome and the whole area of meta-transcriptomics.

And also, partnering more closely with the traditional microbiologists. The thing that is obviously coming out of sequencing now is that we really can’t culture a lot of these organisms, but we can probably figure out ways to pull them out of the environment, for example, using single-cell approaches. We can now start to figure out how to pull out single cells and get the genome of something that hasn’t been cultivated before and, using a reverse of the traditional approach, figure out how to culture these isolates now. So I think that whole area of using sequencing to pull out uncultured isolates and figure out their physiology and what they’re really about is going to be really exciting. We have a group here that is focused on looking at single-cell isolates — pulling these isolates out, generating their genome, and then hopefully culturing some of these guys.

What sequencing technologies are you using?

We work very closely with the companies that produce the instruments and we’re continuously exploring new avenues and keeping ahead of the newest technologies that are out there. We’re working with Illumina, [Roche's] 454 [group], Life Technologies. We are also looking at Pacific Biosciences.

We have 454 on campus, and we also have both [Illumina] and SOLiD systems. So we’re working on all different platforms. And also we’re looking into when you merge the datasets and samples and different platforms together — the different platforms have different benefits. Some are longer, some are shorter, some give you much more data than others. We’re constantly looking into exploring those.

[In the human microbiome project], we’re using a combination of all platforms. We’re even doing a little bit of Sanger. We’re doing a lot of 454 and [Illumina] and we’ll be working towards assembling large data sets based on both platforms. The different platforms have different benefits and we’re aiming to take advantage of those. Just as an example, some of the isolates for our reference genome sequencing are closely related. So, you can [sequence one of the isolates] with 454, and then you can do multiple, closely related cousins on different platforms, and you get deeper coverage at reduced cost. We are being very creative in terms of how we merge different platforms of sequencing — it’s not going to be one single platform.

Where are you in the human microbiome project, and what will you be doing going forward?

We have multiple projects. One component is devoted to the four large-scale centers – Baylor [University], JCVI, [the Broad Institute], [Washington University] — and for that part we are sequencing from 15 to18 different body sites. I think it’s close to 600 people now that we have recruited. So, different samples from the body sites are being sent to the centers and they’re all being collated and then hosted by the data analysis and collation center. So we are creating primarily a huge resource for the community.

In addition to that, we are going to be doing between 200 [and] 300 reference genomes with the hope that we will be able to align a lot of this metagenomic data to these reference genomes. The [human microbiome] consortium is doing between 900 [and] 1,000 reference genomes, but just at JCVI, we’re doing 200 to 300. And, for that we get samples from all over the world.

In addition, we were awarded two demonstration projects. So, I have a project at [New York University] with Zhiheng Pei, where we’re following 80 individuals over the next few years and looking at esophageal cancer. We have preliminary information that would suggest that there is a microbial component to cancer of the esophagus. And, another individual at JCVI who works very heavily on the human microbial initiative is Dr. Barbara Methe. She also has a demonstration project from the National Institutes of Health looking at psoriasis in collaboration with Dr. Martin Blaser [at NYU].

You mentioned earlier being excited about taking advantage of the new sequencing technologies. How are these advancements in sequencing technology impacting the scope of what you’re able to do, and the pace of your research?

The scope became much larger. I always tease my colleagues and say that in 2006 you could get on the cover of Nature with two people. And now, it’s like, if you have a hundred, well maybe that gets you in one of the top journals. So things have really changed in the past few years.

I think by virtue of having access to the new technologies, we can adapt questions beyond just our immediate area. For example, we would like to be able to look at diseases in the developing world. And now we can probably do that because costs have come down so much. You can focus on a population beyond just the populations that have been most traditionally studied. So I think it’s really expanding the concept of which diseases we can look at, which health conditions we can look at, and even for the environment, we can now sequence much deeper in the environment, and access microbial diversity far beyond what we could have done five or ten years ago.

But in parallel with that, we’ve had to obviously keep up the pace on the data analysis side and data storage side because you have so much more data coming out. So that’s also been another exciting area for us. We have people like Shibu Yooseph and Granger Sutton. They work very closely with the data and look at the assembly approaches we have to use and the metagenomics tools that have to be adapted.

One of the things to point out is that of the large sequencing centers, JCVI has been at the forefront of metagenomics and has probably processed the largest number of metagenomic data sets just by virtue of when we got on board in the process. So, we’ve really had to keep apace and learn rapidly the new tools — what works, what doesn’t work, and how we have to grow to adapt to new sequencing technologies as they come on board

It’s actually never been dull, but especially in the last few years with the new sequencing technologies, it’s become particularly exciting.

And I think also, we can now ask this question about the rare species in environments. I think because we were limited before by sequencing depth, we were not able to drill down and see what they call the minor players. And now we can actually start to access these organisms. Just to give you an example, Scott Peterson, who is at the [JCVI] Rockville campus, has been looking at oral plaque and he has been able to show you using subtractive hybridization approaches, that when you pull out the dominant players you can see up to 50 to100 species that you have not seen traditionally before as being present in the oral cavity. And that’s because people have not been able to drill down to that depth previously.

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The Top 10 Everything of 2009

28/12/2009 às 17:23 (GWAS, hapmap)

2. The Human Epigenome, Decoded

By EBEN HARRELL Tuesday, Dec. 08, 2009
Visuals Unlimited / Corbis

The decoding of the human genome nearly a decade ago fueled expectations that an understanding of all human hereditary influences was within sight. But the connections between genes and, say, disease turned out to be far more complicated than imagined. What has since emerged is a new frontier in the study of genetic signaling known as epigenetics, which holds that the behavior of genes can be modified by environmental influences and that those changes can be passed down through generations. So people who smoke cigarettes in their youth, for example, sustain certain epigenetic changes, which may then increase the risk that their children’s children will reach puberty early. In October, a team led by Joseph Ecker at the Salk Institute in La Jolla, Calif., studied human skin and stem cells to produce the first detailed map of the human epigenome. By comparing this with the epigenomes of diseased cells, scientists will be able to work out how glitches in the epigenome may lead to cancers and other diseases. The study, which was published in the journal Nature, is a giant leap in geneticists’ quest to better understand the strange witches’ brew of nature and nurture that makes us who we are.

View the full list for “The Top 10 Everything of 2009″

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Personal Genome Project Sees Whole-Genome Sequencing as ‘Increasingly a Viable Option’

27/12/2009 às 21:29 (GWAS, hapmap)

Personal Genome Project

By Julia Karow

This article was originally published Oct. 14.

Organizers of Harvard Medical School’s Personal Genome Project said that as the cost of DNA sequencing declines, they are considering whole-genome sequencing rather than exome sequencing for the second phase of the study, PGP-100.

The project has already added results from the genome of its founder and principal investigator, George Church, to its website, whose genome was recently sequenced by Complete Genomics.

Launched in 2007 with 10 participants, the PGP aims to sequence the genomes of 100,000 people and to correlate their genotypes with trait information. In April, the project said that it plans to scale up to 100 participants for its second phase (see In Sequence 5/5/2009).

In a newsletter e-mailed last week to individuals interested in the study, PGP organizers said that they have been closely monitoring the decrease in cost of whole human genome sequencing “because it will impact our sequencing strategy for the PGP-100.”

The cost has already fallen to less than $50,000 per genome, PGP said, “with some speculating that the arrival of $5,000 genomes is imminent,” a reference to Complete Genomics’ $5,000 human genome sequencing service, which is scheduled to launch in January.

The project’s initial strategy, according to the organizers, was to focus on the exome, since it is “information rich” and seemed “a more economical alternative” to whole-genome sequencing.

“However, the cost of exome sequencing has not fallen as rapidly as whole-genome sequencing,” they noted, and as a result, for the PGP-100, “the decision to pursue whole genome is increasingly a viable option.”

It will depend, though, on factors such as the project’s ability to raise funding as well as “the willingness of sequencing companies to publicly showcase their technologies through sponsorship of PGP-100 genomes.”

According to its website, the PGP is funded by donations from individuals, Google, Orbimed, the COUQ Foundation, a grant from the Broad Institute, technology development grants from the Department of Energy and NIH, and in-kind support from various organizations. PersonalGenomes.org, a 501(c)3 charitable organization, seeks to raise $1.5 million in donations for the project this year from foundations, private companies, and individuals.

The project has already posted results from an analysis of Church’s genome, which was recently sequenced by Complete Genomics. Church told In Sequence last month that the company has committed to sequencing nine additional PGP genomes, though it was unclear whether it will charge the PGP for its services (see In Sequence 9/15/2009).

For the interpretation of the genomic information, the PGP is using Trait-o-matic, an open-source tool developed in house, which automatically identifies, filters, and annotates genetic variants. The project plans to use the software to generate research reports that contain variants “that may be of potential significance” and has already generated prototypes of such reports for its first 10 participants, based for nine of them on partial exome sequence data.

Future releases of Trait-o-matic will “enable a community of volunteers to annotate and interpret integrated genomic and trait datasets from the PGP.”

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The Technology Pilot

27/12/2009 às 21:25 (Genotipagem de SNPs, GWAS, hapmap)

If you’ve been wondering how the 1,000 Genomes Project is doing, here’s an account from Dan Koboldt at his MassGenomics blog about last week’s meeting at Baylor where participants discussed the “Pilot 3″ phase of the project. “Unlike pilots 1 and 2, which emphasized whole genome sequencing to low or high coverage, respectively, in Pilot 3, the exons of 1,000 genes (~1.5 Mbp total) were selectively targeted for sequencing by capture technologies,” Koboldt writes. The team is also checking data across platforms and pipelines. “Overall, the Pilot 3 variant calls are looking good – dbSNP concordances in the 70-80% range or higher, and transition/transversion ratios of about 3-3.50 – and consistent across 454 and Solexa data from multiple centers,” he writes.

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First genome-wide, single-base-resolution maps

27/12/2009 às 21:16 (Genotipagem de SNPs, GWAS, hapmap)

Joe Ecker is senior author on a paper that provides the first genome-wide, single-base-resolution maps of methylated cytosines in a mammalian genome. Comparing both human embryonic stem cells and fetal fibroblasts, they found “widespread differences” between the two, including almost one-quarter of all methylation in embryonic stem cells was in a non-CG context, suggesting that embryonic stem cells may use different methylation mechanisms to affect gene regulation, they write.

Several opinion pieces appear this week. One checks in with experts and their concerns regarding the stimulus grants, another looks at the open-source Polymath Project, while a third by Cameron Neylon examines the potential of Google’s open-sorce collaboration tool, Google Wave.

A special insight section explores the changing landscape of neuroscience research. Says an editorial, “The experimental landscape has changed markedly over the past few years, given the technological advances in molecular genetics, optogenetics and functional imaging.” Articles cover molecular genetics and imaging technologies for circuit-based neuroanatomy, neuroscience and systems biology, and multimodal techniques for diagnosing Alzheimer’s disease.

Research led by Joel Levine, a neuroscientist from the University of Toronto, has determined that Drosophila melanogaster flies use a single chemical to communicate gender and sibling identity in order to pick the right sex partners. By inserting a transgene into the fly’s genome that killed cells that produced these special hydrocarbon signaling chemicals, they report that hydrocarbon-free male flies attempted copulating with each other, says a story at the BBC. Check out the accompanying video, too.

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Genetics Suggest Population Expansion in Africa Began in Stone Age

29/07/2009 às 17:24 (GWAS, hapmap, Sanger, Sequenciamento)

July 29, 2009
By Andrea Anderson

NEW YORK (GenomeWeb News) – Modern human populations started expanding some 40,000 years ago, according to a paper appearing appeared online today in PLoS ONE.

Researchers from the University of Arizona and the University of California at San Francisco used multi-locus sequence analysis to assess genetic signatures found in nearly 200 individuals from seven populations around the world. Their results suggest human population expansions in Africa started about 40,000 years ago during the Stone Age — a more recent expansion time than that predicted from previous studies.

“[B]oth hunter-gathers (San and Biaka) and food-producers (Mandenka and Yorubans) best fit models with population growth beginning in the Late Pleistocene,” senior author Michael Hammer, a genetics researcher at the University of Arizona, and his co-authors wrote. “These dates are concurrent with the appearance of the Late Stone Age in Africa, supporting the hypothesis that population growth played a significant role in the evolution of Late Pleistocene human cultures.”

Previous studies based on mitochondrial DNA, Y-chromosome data, or autosomal microsatellites provided a broad range of estimates about when modern human population expansion began, dating as far back as about a few hundred thousand years ago. But such estimates often conflict with one another and are based on one or a few sequences that may be under selective pressure, the researchers explained.

In an effort to generate more reliable data for teasing apart human population history, Hammer and his team used Sanger sequencing to re-sequence roughly 6,000 bases of nuclear DNA from each of about 20 autosomal non-coding regions for 184 individuals.

These regions were selected because they were sites with lots of crossing over events but were also far from protein-coding genes and not likely to be under selection. By looking at all of the areas together, Hammer told GenomeWeb Daily News, it’s possible to overcome the noise detected at any single region.

The individuals tested belonged to seven different populations: San, Biaka, Mandenka, Yoruban, French Basque, Han Chinese, and Melanesian.

When the team analyzed their data using multi-locus analysis, they found evidence suggesting that both hunter-gatherer populations (such as the San from Namibia and the Biaka from the Central African Republic) and food-producer populations (such as the Mandenka from Senegal and Yorubans from Nigeria) began expanding roughly 40,000 years ago during the Late Pleistocene period.

That predates the advent of farming in Africa, Hammer noted, and is consistent with archeological evidence suggesting there was a burst of populations interacting and sharing tools and cultural innovations at that time.

Overall, the team concluded that human populations in Africa began a ten-fold expansion some 36,000 years ago. Their data hint that expansion may have been a tad earlier and faster in the hunter-gatherer population — about a 13-fold expansion starting about 41,000 years ago — than in the food-producing populations, which expanded approximately seven-fold starting some 31,000 years ago.

In the future, the team plans to do additional studies looking at more populations from different parts of the world. And, Hammer said, they also hope to employ next-generation sequencing technology to look at even more regions in the genome.

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GWAS and Differences in DNA Between Tissues

21/07/2009 às 15:50 (Genotipagem de SNPs, GWAS, hapmap)

Posted by Bob Grant
[Entry posted at 20th July 2009 04:52 PM GMT]
www.the-scientist.com/blog
Recent findings may spell trouble for genome-wide association studies based on DNA obtained through blood samples: Genetic material may vary between blood cells and other tissues in a single individual, a study in the July issue of Human Mutation reports.

Image: Wikimedia

The study “raises a very interesting question,” Howard Edenberg, director of the Indiana University School of Medicine’s center for medical genomics, told The Scientist. Many genome-wide association studies — especially studies on systemic diseases such as diabetes and atherosclerosis — depend solely upon DNA harvested from blood samples to identify genes associated with medical conditions. But this study “suggests that looking only at blood, you may miss some things.”

Searching for the genes behind a fatal condition called abdominal aortic aneurysm (AAA), researchers from McGill University in Montreal found that complementary DNA from diseased abdominal aortic tissue did not match genomic DNA from leukocytes in blood from the same patient. “We did not expect to find a difference in the tissue [genes] compared to the leukocyte [genes],” said endocrinologist Morris Schweitzer, who led the study.

Schweitzer and his team uncovered three single nucleotide polymorphisms (SNPs) in samples of diseased tissue from 31 AAA patients that were not present in matching blood samples. They also tested five aortic and blood samples from normal individuals and found the same discrepancy. Schweitzer said that the apparent genetic difference between different cells in the body may cast some doubt on genome-wide association studies that only use DNA from blood samples to infer disease states. “I think they may not be accurate because they might not reflect what’s in the tissue,” he said, adding that researchers should look upon such genetic results “very carefully and very trepidatiously”

Edenberg, who was not involved with the study but who conducts genome-wide association studies to explore the genetic roots of alcoholism and bipolar disorder, said that while the findings are interesting, they are very preliminary. “If they’re correct about this, and there are these genomic differences between tissues and blood at certain alleles, then we’re missing some things,” he said. Edenberg explained that experimenters generally take into account that such studies are somewhat “underpowered” in terms of their ability to catch every genetic indicator of disease. Schweitzer’s results, he noted, may add another layer to this consideration, but do not suggest that genome-wide association studies would turn up false positives, or blood-based genes mistakenly attributed to a particular disease.

Sudha Seshadri, a Boston University neurologist who was not involved in the study, told The Scientist that though the McGill group’s results are important, they do not negate genome-wide association data that scientists have already gathered. “I don’t think [the study] says much about the usefulness or validity of genome-wide association studies as they are being done in cohorts around the world.” Genome-wide studies on diabetes, for example, have identified about 16 genes that are related (in varying degrees) to the disease, said Seshadri, who collaborates on the Framingham Heart Study, a six-decade longitudinal study on more than 5,000 people that has more recently included genomic data.

“I think I would have suggested a few more experiments, personally,” Edenberg added. In particular, he pointed to the fact that the McGill researchers were comparing complementary DNA from aortic tissue to genomic DNA from blood. “At the moment,” he said, the discrepancy “seems relatively compatible with RNA editing [rather] than with a genomic issue.” The study should have compared genomic DNA from the aortic tissues with the genomic blood DNA, and cDNA from both cell types, Edenberg said.

Schweitzer said his group is currently working on this experiment and “should have results probably in a couple of weeks.” He noted that differences between tissue and blood DNA may account for the relatively low levels of association turned up by most genome-wide association studies. Of all the genome-wide association studies that have been conducted, he said, “No one has really found that one miracle gene that really points to something.”

Seshadri, however, said it’s hasty to dismiss the value of such studies. “I think [the authors] make some provocative statements that express a viewpoint, but not a widely-accepted viewpoint,” she said. “It’s far too early in the process of genome-wide association studies to conclude that they have not been fruitful.”

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Canadian Initiative Developing Platform to Map Human Interactome, Eyes International Consortium

15/07/2009 às 08:44 (GWAS, Sequenciamento, Solid)

This story originally ran on July 1 and has been updated to include additional comments.

By Tony Fong

A multi-million dollar effort to create a technology platform to map the human interactome is underway in Canada with an eye to making it international.
Last month the Canada Foundation for Innovation awarded C$9.16 million ($7.89 million) to a national initiative to create a technology platform, bringing the total funding for the project to C$22.9 million ($19.7 million).

A total of 12 universities throughout Canada will be working on the interactome project.

Once the national technology platform becomes operational, the plan is to bring in institutions and partners from around the globe in an international push to create a complete set of cellular interaction networks.

In an interview with ProteoMonitor this week, Benoit Coulombe, who is heading the Canadian work and is a professor and director of the Proteomics Discovery Platform at the Institut de Recherches Cliniques de Montreal, said that the national technology platform comprises the 12 universities along with their instruments, methods, workflows, and expertise in elucidating the human interactome.

Much of the funding will be directed at purchasing new equipment and renovating facilities. The C$9.16 funding from CFI, an independent corporation created by the Canadian government, is for infrastructure. The remaining C$13.74 million, which comes from other partners such as the province of Quebec and companies such as Thermo Fisher Scientific, also will be used for infrastructure costs, not operational expenses, Coulombe said.

Among the new equipment that will be purchased are: Thermo Fisher’s Orbitrap mass spectrometers; Illumina’s Genome Analyzer and Applied Biosystems’ SOLiD second-generation DNA sequencing platforms; robotic liquid handlers; confocal microscopes; and other instruments.

While the 12 universities are already mapping the human interactome, the national initiative brings them together in a collaborative mode that can lead to greater efficiency, more reliable results, and generally better science, Coulombe said.

“The idea of this technology platform is that we put together 12 universities across Canada … that already have activities in protein-protein interaction or interactome studies,” he said. In a virtual manner, “these 12 institutions [will now] sit around the same table and plan their activities relating to protein-protein, protein-RNA interaction studies, et cetera. … Now we have a coordinated platform and now we can plan the equipment [and] the technology pipeline that we want to run.”

New methods development, especially in computational approaches, will also be part of the initiative.

The schools involved in the effort are IRCM, which is affiliated with the University of Montreal; Centre for Cellular and Biomolecular Research at the University of Toronto; Samuel Lunenfeld Research Institute at the Mt. Sinai Hospital; the Ottawa Institute of Systems Biology at the University of Ottawa; the Université de Sherbrooke; Dalhousie University; the University of Victoria; the University of British Columbia; the University of Manitoba; the Institut de Recherché en Immunologie et en Cancérologie at the University of Montreal; McGill University; and the Université Laval.

Because each participating institution has its own area of expertise, the initiative will allow researchers to tap into information that they otherwise might not have access to, Coulombe said. In addition, the organizational structure will facilitate interlaboratory work among the participants, which could improve reproducibility, he added.

When different schools perform a similar experiment, it will be important that common standard operational procedures are in place and followed “so that the data that comes out of the many sites…are comparable,” Coulombe said.

“The only way to achieve this is through communication between the sites. So if some of the sites combine their efforts in [a] project, we have to be able to tell the funders that when we do the same type of experiments in different locations, we’re doing it in a way that the data can be compared, is reproducible, [and] is complementary but can be put together,” he said. “So this is one of the important virtues of this type of platform.”

The initiative is currently performing a multi-site pilot project comparing affinity purification techniques. Each site, using similar equipment and analytical methods for the same proteins, is generating data, which will then be analyzed to determine what steps need to be taken to resolve differences between different labs.

In addition, they are investigating methods aside from mass-spec based technologies to monitor protein-protein interactions such as yeast 2-hybrids and luminescence-based mammalian interactome technology, or LUMIER, Coulombe said.

Within six months, most of the new equipment should be installed and the national platform should be “90 percent operational.” In a year, “we plan to have operational funding for at least one big interactome project,” he said.

If that happens, it would be one of the few examples of such a project. While there have been calls in the past for a large-scale human interactome mapping effort, such proposals have failed to take flight and most of the current work has been confined to individual labs. According to Tony Pawson of the Samuel Lunenfeld Research Institute and a participant in the Canadian effort, only about 5 percent of the human interactome has been mapped to date.

The most prominent proponent of a coordinated interlaboratory approach to describing the human interactome has been Marc Vidal, an associate professor of genetics at the Harvard Medical School, who in 2006 published an article in The Scientist advocating for a $100 million investment into a large-scale human interactome mapping effort. While the funding agencies never took him up on his advice, a number of smaller individual efforts have been started since then, he told ProteoMonitor.

The Center for Cancer Systems Biology at the Dana Farber Cancer Institute, of which Vidal is director, has also adopted the Human Interactome Mapping Project as its flagship project.

“We’re not quite there yet … if you were to compare us to the genome sequencing project at its peak, but it’s definitely starting to crystalize a bit,” he said. “People are getting together, people are publishing four, five, six groups together. … I also think that the field as a whole is already past the single lab, single R01 [stage].”

In January, he and a cadre of other collaborators published a series of articles in Nature Methods describing research into the interactomes of various organisms.

The Systems Biology Center New York has also been exploring the idea of a Quantitative Human Interactome Project to “experimentally obtain kinetic constants for cellular interactions between all of the proteins encoded by the human genome and construct a database of these parameters,” according to a report it released in March 2008.

Coulombe said that the Canadian initiative is the only one he knows of that pulls together the resources of so many institutions and directs it at the human interactome.

But at a time when other similar projects, such as mapping the human proteome, have failed to gain any traction, and protein-protein interactions within the human model are still poorly understood, are Coulombe and his peers jumping ahead of themselves with their ambitions to map the human interactome, which looks not only at protein-protein interactions but also at protein-DNA and protein-RNA interactions?

They don’t see it that way. Pawson said that the technology has reached the stage where “it’s really feasible to think about doing these things on a large scale, and also very importantly, people who use different approaches … are starting to talk to each other much more extensively.”

Indeed, while the funding announced last week focuses on building the national technology platform, Coulombe and others in the initiative are already looking ahead to a large-scale effort that would involve researchers from across the globe to map the human interactome. That effort is called the International Interactome Initiative, or I3.

“This is one of the projects that we hope will be supported by the platform,” Coulombe said. “The national platform is the technology platform in Canada that will serve in the international interactome initiative.”

The Canadian initiative and the proposed I3 plan comes out of a project called the Human Proteotheque Initiative that Coulombe has been working on for several years to chart protein interactions that regulate cell growth, differentiation, and disease progression [see PM 08/02/07].

“What you see now [with I3] is the evolution of this initiative,” Coulombe said. “We’re building our way to the interactome.”

He and others involved in trying to get I3 off the ground have created a steering committee “that includes key players in the interactome field from the US, from Europe and from Canada,” that is exploring funding opportunities for the project and setting scientific objectives, Coulombe said, adding that he hopes to have funding for I3 secured next year so that research can begin in early 2011.

“With this international consortium, we feel that if we have appropriate funding, by joining efforts and technologies such as affinity purifications, mass spectrometry, yeast 2-hybrids, protein complementation assays, LUMIER … in five years we [could] have a draft map of the interactome with pretty much full coverage,” Coulombe said.

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