Immune responses in neonates are often defined by dominant production of anti-inflammatory Th2 cytokines, a condition referred to as the neonatal Th2 bias. This pattern of cytokine secretion is thought to contribute to the susceptibility of young animals to infection and to the development of Th2-mediated diseases, such as asthma. Thus, vigorous Th2 function is a key defining feature of immunity in early life. Learning how this arises is of central importance for our understanding of the ontogeny of the adaptive immune system. The Th2 biased responses of neonates are due, in part, to rapid, high-level Th2 cytokine production. We have demonstrated that the Th2 cytokines IL-4 and IL-13 are both produced early and in copious amounts by newly activated murine neonatal CD4+ cells. In adult Th2 cells, coordinate expression of high levels of Th2 cytokines requires an intergenic regulatory region called conserved non-coding region 1 (CNS-1). Therefore, the CNS-1 genetic element may contribute importantly to the early, robust Th2 cytokine production in neonates. Indeed, this DNA element exists in a relatively permissive epigenetic state in early ontogeny. We have found that, unlike in naive adult cells, CNS-1 is hypomethylated at CpG residues in naive fetal and neonatal T lineage cells. This hypomethylated state is strongly linked to rapid, high level Th2 cytokine production by neonatal CD4+ cells. Methylation of CNS-1 occurs rapidly post birth, with adult levels of methylation being attained by the end of the first week o life. Increasing methylation of CNS-1 with increasing development is associated with the progressive silencing of rapid Th2 cytokine gene expression. Together, these observations indicate that CNS-1 may play a centrally important role in defining the neonatal Th2 bias. Our long-term goals are to understand the genetic and epigenetic contributions to the Th2 dominant state of early life. In this proposal, we will focus on the well-defined CNS-1 region in the Th2 cytokine locus. Our preliminary data lead to two clear hypotheses.
Specific Aim 1 will test the idea that the physical presence of the CNS-1 region is required to achieve Th2 cytokine overproduction in neonates. This will be achieved by comparing the epigenetic state of the Th2 locus and Th2 cytokine production in wild-type and CNS-1-deficient neonates.
Specific Aim 2 will examine the premise that hypomethylation of CNS-1 in fetal and neonatal life is essential for generating the neonatal Th2 bias. This will be tested by creating transgenic mice with forced methylation of CNS-1 throughout development. Constructs containing a CNS-1-targeted zinc finger protein coupled to the catalytic domain of the de novo DNA methyltransferase Dnmt3a will be created~ expression of the constructs will be confined to T lineage cells. The epigenetic state of the Th2 locus and Th2 function will be assessed in the transgenic mice. These studies will be the first to dissect genetic and epigenetic regulation at the Th2 cytokine locus in early ontogeny. Results from these studies will ultimately lead to interventional strategies aimed at improving pediatric health by mitigating the Th2 dominant state in early life.
This proposal will examine the contribution of selective components of the genome to the poor immunity in newborn animals. This information will provide an important foundation for developing novel approaches to meet the health challenges of pediatric life, such as (a) enhancing vaccine responsiveness, (b) mitigating pediatric-onset asthma, and (c) increasing resistance to pathogenic microorganisms.