Closing the gap between genotype and phenotype
2
NEWS AND VIEWS in ALL pathogenesis, but many important questions remain regarding the mechanisms by which these alleles contribute to leuke- mogenesis. Papaemmanuil et al. report that the risk haplotype at IKZF is associated with reduced expression of IKZF in Epstein-Barr virus (EBV)-transformed lymphocytes, suggesting that an allele-specific reduction in IKZF expression might impair normal lymphoid development and increase the risk of lymphoid transformation. In con- trast, Papaemmanuil et al. did not see allele- specific differential expression of ARID5B and CEBPE in EBV-transformed lym- phocytes, although Treviño et al. did find ARID5B genotype–specific effects on expres- sion in leukemic cells. As is the case for most germline variants identified in GWAS, the mechanisms by which these alleles predis- pose to leukemia is not fully understood. However, the relatively short latency of pediatric ALL, the detailed understanding of the molecular pathogenesis of ALL and the existence of well-established model systems should facilitate the extensive genetic and functional studies needed to unravel how these germline variants increase ALL risk. In addition, these studies demonstrate the importance of integrating data from disease- wide GWAS with the clinical and molecular het- erogeneity of human malignancies (Table 1). Although some germline alleles might increase the general risk of a particular tumor type, it is equally likely that inherited variants might confer an increased risk for developing specific disease subtypes, as was demonstrated in the current studies for ARID5B and B-cell hyper- diploid ALL. In addition, some germline alleles might instead serve as modifier loci that do not increase the risk of transformation but rather influence the malignant phenotype. Moreover, some germline risk alleles might increase the propensity to acquire specific somatic muta- tions, as has been demonstrated in specific instances 11,12 . Future studies should assess whether the germline IKZF risk alleles increase the risk of acquiring somatic alterations at this locus 9 and whether there are additional ger- mline alleles that increase the risk of acquiring other somatic alterations that contribute to the pathogenesis of ALL. It is likely that subsequent GWAS efforts in adult and pediatric ALL and in acute myeloid leukemia (AML) will identify additional loci associated with increased risk of these diseases. In some cases, these loci might fall within genes encoding known members of hematopoietic pathways, while in other cases, these studies might implicate new genes and provide insight into new pathways relevant to leukemogen- esis. The current studies illustrate that only by incorporating clinical, pathological and molecular annotation into future GWAS will investigators elucidate the full complement of germline alleles that contribute to the patho- genesis of human malignancies. 1. Mullighan, C.G. et al. Nature 446, 758–764 (2007). 2. Papaemmanuil, E. et al. Nat. Genet. 41, 1006–1010 (2009). 3. Treviño, L.R. et al. Nat. Genet. 41, 1001–1005 (2009). 4. Hasle, H., Clemmensen, I.H. & Mikkelsen, M. Lancet 355, 165–169 (2000). 5. Morrell, D., Cromartie, E. & Swift, M. J. Natl. Cancer Inst. 77, 89–92 (1986). 6. Ford, A.M. et al. Nature 363, 358–360 (1993). 7. Lichtenstein, P. et al. N. Engl. J. Med. 343, 78–85 (2000). 8. Hemminki, K. & Jiang, Y. Leukemia 16, 297–298 (2002). 9. Mullighan, C.G. et al. Nature 453, 110–114 (2008). 10. Mullighan, C.G. et al. N. Engl. J. Med. 360, 470–480 (2009). 11. Laken, S.J. et al. Nat. Genet. 17, 79–83 (1997). 12. Campbell, P.J. Nat. Genet. 41, 385–386 (2009). 958 VOLUME 41 | NUMBER 9 | SEPTEMBER 2009 | NATURE GENETICS Closing the gap between genotype and phenotype Peter K Gregersen Making causative connections between genotypic and phenotypic variation is a major challenge for geneticists engaged in the study of human disease. A study drawing this connection for a type 1 diabetes risk locus now demonstrates the importance of focusing on specific quantitative traits and studying them in normal subjects. Peter K. Gregersen is at the Robert S. Boas Center for Genomics and Human Genetics, The Feinstein Institute for Medical Research, North Shore Long Island Jewish Health System, Manhasset, New York, USA. e-mail: [email protected] Over the last few years, genome-wide asso- ciation studies (GWAS) have reported an extraordinary harvest of new genetic associa- tions. This productivity has been particularly rich in the area of human autoimmune dis- orders, where in the aggregate well over 100 confirmed associations have been reported in diseases including type 1 diabetes 1 , inflamma- tory bowel disease 2 and rheumatoid arthritis, among many others 3 . This has presented the scientific community with a tremendous chal- lenge because, with few exceptions, neither the causative variants nor the exact mechanisms underlying these associations are known. On page 1011 of this issue, Linda Wicker and colleagues 4 report a gene-to-phenotype association for IL2RA (or CD25) a locus pre- viously associated with type 1 diabetes 5 . In doing so, they demonstrate an approach to GWAS follow-up studies for exploring the functional basis for a disease association. This study depended on the availability of a population of normal subjects willing to donate blood samples repeatedly for immu- nological analysis, as part of the Cambridge BioResource, who were selected for this study on the basis of IL2RA genotype. Dendrou et al. 4 used flow cytometry to survey the dif- ferences in cell surface expression of the IL2RA protein on several immune cell types and then examined the correlation between those data and IL2RA haplotypes. The results suggest that IL2RA haplotype–specific CD25 expression patterns are directly involved in the risk for autoimmunity. Immunological phenotypes The genetic architecture of most diseases is extremely complex, and therefore, following reports of associated genetic variants, we are usually reduced to handwaving about plau- sible pathways of pathogenesis. One way to get around this dilemma is to focus on less complex phenotypic variation. In the field of immunology, fine-grained analyses of phe- notypic variation have traditionally been carried out in murine systems; however, recent commentaries 6 —including one from a prominent mouse immunologist 7 —have emphasized the importance of examining the full range of functional and phenotypic diversity in the human immune system. The study by Dendrou et al. 4 illustrates the utility of selecting discrete immunologically related phenotypes for study in humans in order to begin to understand the functional significance of disease-associated alleles. © 2009 Nature America, Inc. All rights reserved.
Transcript of Closing the gap between genotype and phenotype
news and v iews
in ALL pathogenesis, but many important questions remain regarding the mechanisms by which these alleles contribute to leuke-mogenesis. Papaemmanuil et al. report that the risk haplotype at IKZF is associated with reduced expression of IKZF in Epstein-Barr virus (EBV)-transformed lymphocytes, suggesting that an allele-specific reduction in IKZF expression might impair normal lymphoid development and increase the risk of lymphoid transformation. In con-trast, Papaemmanuil et al. did not see allele- specific differential expression of ARID5B and CEBPE in EBV-transformed lym-phocytes, although Treviño et al. did find ARID5B genotype–specific effects on expres-sion in leukemic cells. As is the case for most germline variants identified in GWAS, the mechanisms by which these alleles predis-pose to leukemia is not fully understood. However, the relatively short latency of pediatric ALL, the detailed understanding of the molecular pathogenesis of ALL and the existence of well-established model systems should facilitate the extensive genetic and functional studies needed to unravel how these germline variants increase ALL risk.
In addition, these studies demonstrate the importance of integrating data from disease-wide GWAS with the clinical and molecular het-erogeneity of human malignancies (Table 1). Although some germline alleles might increase the general risk of a particular tumor type, it is equally likely that inherited variants might confer an increased risk for developing specific disease subtypes, as was demonstrated in the current studies for ARID5B and B-cell hyper-diploid ALL. In addition, some germline alleles might instead serve as modifier loci that do not increase the risk of transformation but rather influence the malignant phenotype. Moreover, some germline risk alleles might increase the propensity to acquire specific somatic muta-tions, as has been demonstrated in specific instances11,12. Future studies should assess whether the germline IKZF risk alleles increase the risk of acquiring somatic alterations at this locus9 and whether there are additional ger-mline alleles that increase the risk of acquiring other somatic alterations that contribute to the pathogenesis of ALL.
It is likely that subsequent GWAS efforts in adult and pediatric ALL and in acute myeloid leukemia (AML) will identify additional loci
associated with increased risk of these diseases. In some cases, these loci might fall within genes encoding known members of hematopoietic pathways, while in other cases, these studies might implicate new genes and provide insight into new pathways relevant to leukemogen-esis. The current studies illustrate that only by incorporating clinical, pathological and molecular annotation into future GWAS will investigators elucidate the full complement of germline alleles that contribute to the patho-genesis of human malignancies.
1. Mullighan, C.G. et al. Nature 446, 758–764 (2007).2. Papaemmanuil, E. et al. Nat. Genet. 41, 1006–1010
(2009).3. Treviño, L.R. et al. Nat. Genet. 41, 1001–1005
(2009).4. Hasle, H., Clemmensen, I.H. & Mikkelsen, M. Lancet
355, 165–169 (2000).5. Morrell, D., Cromartie, E. & Swift, M. J. Natl. Cancer
Inst. 77, 89–92 (1986).6. Ford, A.M. et al. Nature 363, 358–360 (1993).7. Lichtenstein, P. et al. N. Engl. J. Med. 343, 78–85
(2000).8. Hemminki, K. & Jiang, Y. Leukemia 16, 297–298
(2002).9. Mullighan, C.G. et al. Nature 453, 110–114 (2008).10. Mullighan, C.G. et al. N. Engl. J. Med. 360, 470–480
(2009).11. Laken, S.J. et al. Nat. Genet. 17, 79–83 (1997).12. Campbell, P.J. Nat. Genet. 41, 385–386 (2009).
958 volume 41 | number 9 | september 2009 | nature genetics
Closing the gap between genotype and phenotypePeter K Gregersen
Making causative connections between genotypic and phenotypic variation is a major challenge for geneticists engaged in the study of human disease. a study drawing this connection for a type 1 diabetes risk locus now demonstrates the importance of focusing on specific quantitative traits and studying them in normal subjects.
Peter K. Gregersen is at the Robert S. Boas Center for Genomics and Human Genetics, The Feinstein Institute for Medical Research, North Shore Long Island Jewish Health System, Manhasset, New York, USA. e-mail: [email protected]
Over the last few years, genome-wide asso-ciation studies (GWAS) have reported an extraordinary harvest of new genetic associa-tions. This productivity has been particularly rich in the area of human autoimmune dis-orders, where in the aggregate well over 100 confirmed associations have been reported in diseases including type 1 diabetes1, inflamma-tory bowel disease2 and rheumatoid arthritis, among many others3. This has presented the scientific community with a tremendous chal-lenge because, with few exceptions, neither the causative variants nor the exact mechanisms underlying these associations are known.
On page 1011 of this issue, Linda Wicker and colleagues4 report a gene-to-phenotype association for IL2RA (or CD25) a locus pre-viously associated with type 1 diabetes5. In doing so, they demonstrate an approach to GWAS follow-up studies for exploring the functional basis for a disease association. This study depended on the availability of a population of normal subjects willing to donate blood samples repeatedly for immu-nological analysis, as part of the Cambridge BioResource, who were selected for this study on the basis of IL2RA genotype. Dendrou et al.4 used flow cytometry to survey the dif-ferences in cell surface expression of the IL2RA protein on several immune cell types and then examined the correlation between those data and IL2RA haplotypes. The results suggest that IL2RA haplotype–specific CD25 expression patterns are directly involved in the risk for autoimmunity.
Immunological phenotypesThe genetic architecture of most diseases is extremely complex, and therefore, following reports of associated genetic variants, we are usually reduced to handwaving about plau-sible pathways of pathogenesis. One way to get around this dilemma is to focus on less complex phenotypic variation. In the field of immunology, fine-grained analyses of phe-notypic variation have traditionally been carried out in murine systems; however, recent commentaries6—including one from a prominent mouse immunologist7—have emphasized the importance of examining the full range of functional and phenotypic diversity in the human immune system.
The study by Dendrou et al.4 illustrates the utility of selecting discrete immunologically related phenotypes for study in humans in order to begin to understand the functional significance of disease-associated alleles.
©20
09 N
atu
re A
mer
ica,
Inc.
All
rig
hts
res
erve
d.
news and v iews
IL2RA has been associated with a variety of autoimmune diseases in genome-wide asso-ciation and fine-mapping studies for type 1 diabetes5, Graves’ disease8 and multiple scle-rosis9. IL2RA is an essential component of the trimeric high-affinity receptor for IL-2, a major cytokine for lymphocyte activation and survival10. Given the importance of IL-2 for effector T cell function, it was somewhat surprising that knockout experiments in the 1990s revealed that autoimmunity, rather than immunodeficiency, results from defi-ciency of either IL-2 or its receptors10. This apparent paradox has been at least partially explained in recent years through the real-ization that regulatory T cells (Treg) are critically dependent on exogenous sources of IL-2, as they are unable to produce IL-2 themselves11. At the same time, a charac-teristic feature of Tregs is the constitutive expression of CD25. Thus, it is reasonable to ask whether the IL2RA haplotypes regu-late the level of expression of CD25 on Treg cells or other immunologically relevant cells. One might predict, for example, that high expression of CD25 would be associated with more robust Treg function and hence with protection from autoimmunity.
Toward disease mechanismsAs it turns out, Dendrou et al. show that high CD25 expression is indeed associated with IL2RA haplotypes that confer protec-tion from type 1 diabetes. However, this high CD25 phenotype is observed only on memory CD4+ T cells and not on the Treg population itself. Nevertheless, the associa-tion between protective CD25 haplotypes and high expression of CD25 on CD4+ memory cells is remarkably stable and con-vincing, with extremely high reproducibility on repeated measures in the same individual (with an r2 of >0.99). A reasonable hypoth-esis is that generally high CD4+ memory cell activity will produce a high–IL-2 state that will enhance Treg function, and in fact, activated memory CD4+ T cells do produce more IL-2 in vitro in individuals with the protective CD25 haplotype (Fig. 1). However, the findings of Dendrou et al. suggest that the connection may be more complex. For example, CD25 expression on naive T cells and monocytes is actually reduced by several of the disease-protective CD25 haplotypes. The sizes of these effects are more modest, but significant, and it remains unclear how these particular phenotypes are functionally relevant to disease protection. Interestingly, IL2RA haplotypes that are protective for multiple sclerosis do not include the high-CD25 phenotype on memory CD4+ T cells.
This may indicate that Treg function is less important for protection from multiple scle-rosis, and this is supported by the efficacy of CD25 inhibition in multiple sclerosis, despite the reduction in Tregs that results from this therapy12.
Collective bioresourceThese studies could not have been done without the Cambridge BioResource, a reg-istry currently consisting of ~5,000 normal subjects living near Cambridge, UK, who can be recalled for research studies on the basis of genotype. Note that this bioresource is quite different from the many large cohorts and biobanks that have been developed over the last decade13 because these collec-tions are not designed for selective recall of subjects. The ability to repeatedly study immunological phenotypes in subjects who do not have autoimmune disorders is criti-cal to correlating genotype to phenotype without the confounding variables of active inflammation and drug treatment. In many cases, the genotypes of interest may be quite uncommon, and thus a large pool of subjects is essential. Indeed, even larger sample sizes may be needed, as rare variants are emerging as important contributors to autoimmune disease14,15. In the United States, collabora-tive efforts are underway to develop a similar registry of genotyped normal controls that can be accessed by the scientific community. A large, geographically distributed registry of participants is desirable, as some phenotypes can only be reliably measured using fresh cells. At the same time, there is no reason that such a registry has to be limited to the investigation of immunological phenotypes.
Indeed, neuropsychological testing may be amenable to measurement at a distance using web-based tools. The emerging network of Clinical and Translational Science Centers (CTSC) should facilitate the sharing of such a population resource.
The potential for detailed phenotypic studies of genetically characterized normal populations is enormous. Phenotyping methods in all areas of biomedical research, from immunology to neuroimaging, are advancing rapidly. The emerging data from the 1,000 genomes project suggests that we are still far from a complete catalog of geno-typic variation in the human population. Closing the gap between these two sources of biological variation in the normal popula-tion will be a top priority for human genetics in the coming years.
1. Barrett, J.C. et al. Nat. Genet. 41, 703–707 (2009).
2. Barrett, J.C. et al. Nat. Genet. 40, 955–962 (2008).3. Gregersen, P.K. & Olsson, L.M. Annu. Rev. Immunol.
27, 363–391 (2009).4. Dendrou, C.A. Nat. Genet. 41, 1011–1015 (2009). 5. Lowe, C.E. et al. Nat. Genet. 39, 1074–1082
(2007).6. Hayday, A.C. & Peakman, M. Nat. Immunol. 9, 575–
580 (2008).7. Davis, M.M. Immunity 29, 835–838 (2008).8. Brand, O.J. et al. Clin. Endocrinol. 66, 508–512
(2007).9. Hafler, D.A. et al. N. Engl. J. Med. 357, 851–862
(2007).10. Malek, T.R. Annu. Rev. Immunol. 26, 453–479
(2008).11. Setoguchi, R., Hori, S., Takahashi, T. & Sakaguchi,
S. J. Exp. Med. 201, 723–735 (2005).12. Oh, U. et al. Arch. Neurol. 66, 471–479 (2009).13. Collins, F.S. & Manolio, T.A. Nature 445, 259
(2007).14. Nejentsev, S., Walker, N., Riches, D., Egholm, M. &
Todd, J.A. Science 324, 387–389 (2009). 15. Lee-Kirsch, M.A. et al. Nat. Genet. 39, 1065–1067
(2007).
nature genetics | volume 41 | number 9 | september 2009 959
IL-2 receptor surface expression
Genotype Quantitative trait Disease phenotype
IL2RAhaplotypes
CD4+
memoryT cell
IL-2
Risk Protective
Type 1 diabetes
Regulation ofTreg function andother mechanisms
IL-2receptor
Figure 1 IL2RA (CD25) haplotypes regulate levels of expression of IL-2 receptor on memory CD4+ T cells. High expression is associated with higher production of the cytokine IL-2 by these cells, which in turn may support the activity of regulatory T cells (Treg). Treg function is critical for the prevention of autoimmunity, and this may explain the protection conferred by high IL-2 receptor expression for type 1 diabetes and other IL2RA-associated autoimmune diseases.
©20
09 N
atu
re A
mer
ica,
Inc.
All
rig
hts
res
erve
d.
