Genome-Wide Identification of Novel Genes Involved in Early Th1 and Th2 Cell Differentiation

Abstract

Th cell subtypes, Th1 and Th2, are involved in the pathogenesis or progression of many immune-mediated diseases, such as type 1 diabetes and asthma, respectively. Defining the molecular networks and factors that direct Th1 and Th2 cell differentiation will help to understand the pathogenic mechanisms causing these diseases. Some of the key factors regulating this differentiation have been identified, however, they alone do not explain the process in detail. To identify novel factors directing the early differentiation, we have studied the transcriptomes of human Th1 and Th2 cells after 2, 6, and 48 h of polarization at the genome scale. Based on our current and previous studies, 288 genes or expressed sequence tags, representing ∼1–1.5% of the human genome, are regulated in the process during the first 2 days. These transcriptional profiles revealed genes coding for components of certain pathways, such as RAS oncogene family and G protein-coupled receptor signaling, to be differentially regulated during the early Th1 and Th2 cell differentiation. Importantly, numerous novel genes with unknown functions were identified. By using short-hairpin RNA knockdown, we show that a subset of these genes is regulated by IL-4 through STAT6 signaling. Furthermore, we demonstrate that one of the IL-4 regulated genes, NDFIP2, promotes IFN-γ production by the polarized human Th1 lymphocytes. Among the novel genes identified, there may be many factors that play a crucial role in the regulation of the differentiation process together with the previously known factors and are potential targets for developing therapeutics to modulate Th1 and Th2 responses.

A Network Modeling Approach to Analysis of the Th2 Memory Responses Underlying Human Atopic Disease

“…4 As illustrated in Fig. 1B, most of these genes were elevated in the atopic responses, including several previously recognized members of the Th2 cascade (IL-4R, IL-5, IL-13, GFI-1, ITK), as well as a cohort of novel Th2-associated genes (IL-17RB, CAMK2D, CISH, DACT1, DPP4, MAL, NDFIP2, NSMCE1, PLXDC1) reported recently by us (16) and confirmed in an independent study (28).…”

“…Of note, application of network analysis techniques to the microarray data not only confirmed the importance of the bulk of Th2-associated genes identified by more conventional analyses in previous studies (16,28), but it also added a second tier of previously unrecognized Th2-associated genes identified purely on the basis of their patterns of interconnectivity with other members of the atopy module (Table I). These novel genes encode proteins mainly involved in generic functions such as transcriptional regulation and signal transduction; however, their precise function in allergen-driven Th cell responses remains to be investigated.…”

“…This finding justifies further pursuit of this overall approach, in particular studies related to the network regulatory functions of other "novel" hub genes. The most direct approach would be to employ small interfering RNA-mediated knockdown of hub(s), as has been performed in Th cell lines (28); however, effective methodology is not available to achieve this in primary human Th memory responses.…”

“…However, it is not clear why the atopy module contains gene expression signatures associated with the early differentiation of Th2 cells (28), Th2 memory responses (16), and Treg function (30). One possibility is that because the Th memory compartment is phenotypically and functionally heterogeneous (36), the above analyses on total CD4 T cells are likely to have provided an oversimplified picture.…”

Genome-Wide Analysis Reveals Unique Regulation of Transcription of Th2-Specific Genes by GATA3

“…Transcriptional profiling by DNA microarray analysis also helped clarify the genes whose expression is controlled by STAT6, and the results argue that STAT6 is clearly important for regulating Th2 status (28,29). However, these studies do not address whether these genes are directly regulated by STAT6 or not.…”