Donata Vercelli
Assistant Director, Arizona Respiratory Center (Functional Genomics)
Professor, Department of Cell Biology & Anatomy
Member, Graduate Interdisciplinary Program in Genetics
M.D., University of Florence, Italy
E-mail: donata@resp-sci.arizona.edu
FUNCTIONAL GENOMICS LABORATORY
Research in the Functional Genomics Laboratory focuses on the molecular mechanisms underlying the pathogenesis of complex diseases such as asthma and allergy. The aim of this work is to understand how the development of these diseases relates to the dysregulation of critical relevant genes. The ultimate goal is to establish a new paradigm that correlates genetic analysis, functional studies and patient phenotypes in order to understand the cause of disease and predict responsiveness to specific treatments.
Functional Genomics of IL-13
This project focuses on the functional analysis of the impact that complex patterns of genetic variation have on the expression of the IL-13 gene. The long-term objective is to determine the mechanisms that regulate human IL-13 expression, the influences of single nucleotide polymorphisms (SNP) on IL-13 expression, and the mechanisms by which the polymorphisms contribute to susceptibility to allergy and/or asthma.
IL-13 appears to be centrally involved in the pathogenesis of asthma and allergy. We recently identified seven polymorphisms in the human IL-13 gene. Two SNPs are in the 5’ regulatory region of IL-13, one is in the third intron, one is in the coding region and leads to a predicted amino acid change, and three are in the 3’ untranslated region (UTR). All SNPs, except the ones in the 5’ region, are in almost complete linkage disequilibrium, and the frequency of the rare genotype is 0.23. Notably, homozygosity for the rare variants was found to be strongly associated to increased total serum IgE and wheezing at age 6, suggesting that genetic variation results in dysregulation of IL-13 expression and/or activity. Because both regulatory and coding regions may contain SNPs, dysregulation may be quantitative and/or qualitative. We are currently studying the regulation of IL-13 gene expression and the mechanisms whereby the latter may be quantitatively altered by SNPs. In particular, we are assessing whether the polymorphisms affect chromatin accessibility at the IL-13 locus, and/or transcriptional or post-transcriptional regulatory mechanisms. The effects of the coding region SNP on the structure and function of IL-13 protein are also under investigation.
Recent Publications:
Vercelli D. The functional genomics of CD14 and IgE-mediated disease: An integrated view. J. Allergy Clin. Immunol. 109: 14-21 (2002)
Strempel JM, RB Webster and D Vercelli. CNS-1 is a regulator of human IL-13 transcription. FASEB 2002, New Orleans, April 2002
Webster RB and D Vercelli. IL-13 chromatin accessibility in human T cells. FASEB 2002, New Orleans, April 2002
Cameron L, M Kabesch and D Vercelli. Mechanisms underlying the attenuation of IL-13 transcription by a single nucleotide polymorphism in the promoter, 1112CÆT. FASEB 2002, New Orleans, April 2002
Functional genomics of innate immunity
Our interest in innate immunity began a number of years ago, with the realization that IL-4, the key cytokine in IgE isotype switching, downregulates the expression of CD14, the monocyte molecule that is a critical component of the receptor for bacterial pathogens (LPS/endotoxin). Later on, we found a similar activity for IL-13. Over the years, we have characterized several CD14-dependent functions, including its ability to trigger homotypic adhesion and regulate T cell activation. We also defined the structural requirements for CD14 activation by mycobacterial products. However, our most important finding was that engagement of CD14 modulates the synthesis of immunoglobulins, including IgE. This work provided a potential connection between innate CD14-dependent responses and IgE-mediated allergic disease. This link is pursued vigorously in the Functional Genomics Laboratory. Indeed, the researchers at the Arizona Respiratory Center had identified a SNP in the CD14 promoter that is associated with increased levels of soluble CD14, and decreased levels of IgE. We are investigating the molecular mechanisms responsible for the increased transcription of the polymorphic promoter. Using a combination of reporter assays and EMSA analysis, we have demonstrated that the SNP decreases the affinity of DNA/protein interactions at a GC box that binds member of the Sp family of transcription factors. Increased transcription results from decreased sensitivity to the repressive effect of Sp3. The cellular pathways that link CD14-dependent events with IgE regulation are also under investigation. The Arizona Respiratory Center recently obtained a four-year grant Program for Genomic Application on Innate Immunity in Diseases of Heart, Lung and Blood. This Program will allow us to deepen our understanding of the role that innate immunity genes and their variation play in the pathogenesis of allergy and asthma.
Recent Publications:
Vercelli D, M Baldini and F. Martinez. The monocyte/IgE connection: May polymorphisms in the CD14 gene teach us about IgE regulation? Int. Arch. Allergy Immunol. 124: 20-24 (2001)
Vercelli D, M Baldini, D Stern, IC Lohman, M. Halonen and F Martinez. CD14: A bridge between innate immunity and adaptive IgE responses. J. Endotoxin Res. 7: 45-48 (2001)
LeVan TD, JW Bloom, TJ Bailey, CL Karp, M Halonen, FD Martinez, and D Vercelli. A common single nucleotide polymorphism in the CD14 promoter decreases the affinity of Sp protein binding and enhances transcriptional activity. J. Immunol. 167: 5838-5844 (2001)
IL-4-dependent Isotype Switching: Differential Regulation of IgE and IgG4 Expression
Allergy and asthma are generally defined as IgE-dependent diseases due to a dysregulation of Th cell differentiation that results in dominant IL-4-induced Th2 responses. Increases in allergen-specific IgG4, such as those typically observed in successful allergen-specific immunotherapy, are therefore mostly ascribed to reprogramming of T cell functions towards Th1-dependent responses. However, a crucial issue not often appreciated is that IgG4 responses are themselves IL-4-dependent, so that their emergence and/or enhancement should rather be considered as a modified Th2-dependent effect. The important conceptual implication of this line of reasoning is that Th2-induced antibody responses against allergens may not necessarily be pathogenic, and when they are skewed towards IgG4 expression they may actually be protective.
The main theme of our work is the analysis and comparison of the molecular mechanisms that control the synthesis of IgE and IgG4 in human B cells. These two Ig isotypes are expressed as a result of IL-4-induced sequential switching and therefore share antigen specificity. Our data from epidemiological studies show that IL-4-dependent antibody responses can be uncoupled in vivo, and the ratio between allergen-specific IgE and IgG4 is significantly higher in sensitized asthmatics than in sensitive patients without asthma. Furthermore, IL-10 promoter polymorphisms associated with lower IL-10 production appear to be associated with increased IgE levels in early life. Interestingly, IL-10 is known to suppress IgE while enhancing IgG4 responses. Thus our overall hypothesis is that the ratio between allergen-specific IgE and IgG4, more than the absolute levels of either Ig class, is critical for the clinical outcome of allergic responses. IL-10 may play a critical role in regulating the IgE/IgG4 ratio. Therefore we are elucidating the molecular mechanisms underlying the differential regulation of IgE and IgG4, and the role played by IL-10 in this process. Because the isotype targeted by switch recombination is determined through the process of germline transcription, our analysis will focus on the mechanisms that control the activation of the human e and g4 GL promoters. In view of the role of IgE in the pathogenesis of allergy and asthma, and the demonstrated ability of allergen-specific IgG4 to block IgE-dependent responses to the same allergen in vivo, our long-term objective is to determine the molecular mechanisms that would lead to preferential production of antigen-specific IgG4.
Recent Publications:
Vercelli D. One cytokine, one isotype - A Trojan horse, Pandora's box and an evolving paradigm. Am. J. Respir. Crit. Care Med. 162:S86-S90 (2000)
Vercelli D. Immunoglobulin E and its regulators. Curr. Opin. Allergy Clin. Immunol. 1: 61-65 (2001)
Monticelli S and D. Vercelli. Molecular regulation of class switch recombination to IgE through e germline transcription. Allergy 56: 270-278 (2001)
Vercelli D. Novel insights into class switch recombination. Curr. Opin. Allergy Clin. Immunol. 2: 147-51 (2002)
Agresti A and D Vercelli. c-Rel is a selective activator of a novel IL-4/CD40 responsive element in the human Ig g4 germline promoter. Mol. Immunol. 38: 849-859 (2002)
Monticelli S, R Ghittoni, M Kabesch, and D Vercelli. Myb proteins repress human e germline transcription by inhibiting STAT6-dependent promoter activation. Mol. Immunol. (in press)
