Elevated expression of chemokine receptors in tumors has been reported in many instances and is related to a number of survival advantages for tumor cells including abnormal activation of prosurvival intracellular pathways. In this work we demonstrated an inverse correlation between expression levels of p53 tumor suppressor and CXCR5 chemokine receptor in MCF-7 human breast cancer cell line. Lentiviral transduction of MCF-7 cells with p53 shRNA led to elevated CXCR5 at both mRNA and protein levels. Functional activity of CXCR5 in p53-knockdown MCF-7 cells was also increased as shown by activation of target gene expression and chemotaxis in response to B-lymphocyte chemoattractant CXCL13. Using deletion analysis and site-directed mutagenesis of the cxcr5 gene promoter and enhancer elements, we demonstrated that p53 appears to act upon cxcr5 promoter indirectly, by repressing the activity of NFκB transcription factors. Using chromatin immunoprecipitation and reporter gene analysis, we further demonstrated that p65/RelA was able to bind the cxcr5 promoter in p53-dependent manner and to directly transactivate it when overexpressed. Through the described mechanism, elevated CXCR5 expression may contribute to abnormal cell survival and migration in breast tumors that lack functional p53.
Interleukin-1 (IL-1) plays a crucial role in numerous inflammatory diseases via action on its only known signaling IL-1 receptor type 1 (IL-1R1). To investigate the role of IL-1 signaling in selected cell types, we generated a new mouse strain in which exon 5 of the Il1r1 gene is flanked by loxP sites. Crossing of these mice with CD4-Cre transgenic mice resulted in IL-1R1 loss of function specifically in T cells. These mice, termed IL-1R1ΔT, displayed normal development under steady state conditions. Importantly, isolated CD4 positive T cells retained their capacity to differentiate toward Th1 or Th17 cell lineages in vitro, and strongly proliferated in cultures supplemented with either anti-CD3/CD28 or Concanavalin A, but, as predicted, were completely unresponsive to IL-1β administration. Furthermore, IL-1R1ΔT mice were protected from gut inflammation in the anti-CD3 treatment model, due to dramatically reduced frequencies and absolute numbers of IL-17A and interferon (IFN)-γ producing cells. Taken together, our data shows the necessity of intact IL-1 signaling for survival and expansion of CD4 T cells that were developed in an otherwise IL-1 sufficient environment.
CD58 is expressed on the surface of antigen-presenting cells, including B-cells, and provides co-stimulation to regulatory T-cells (Treg) through CD2 receptor binding. Tregs appear to be essential suppressors of tissue-specific autoimmune responses. Thereby, CD58 plays protective role in multiple sclerosis (MS) and CD58 was identified among several loci associated with MS susceptibility. Minor (C) variant of the single-nucleotide polymorphism (SNP) rs1335532 is associated with lower MS risk according to genome-wide association studies (GWAS) and its presence correlates with higher CD58 mRNA levels in MS patients. We found that genomic region containing rs1335532 has enhancer properties and can significantly boost the CD58 promoter activity in lymphoblast cells. Using bioinformatics and pull-down assay we found that the protective (C) rs1335532 allele created functional binding site for ASCL2 transcription factor, a target of the Wnt signaling pathway. Both in B-lymphoblastoid cell lines and in primary B-cells, as well as in a monocytic cell line, activation of Wnt signaling resulted in an increased CD58 promoter activity in the presence of the protective but not the risk allele of rs1335532, whereas ASCL2 knockdown abrogated this effect. In summary, our results suggest that ASCL2 mediates the protective function of rs1335532 minor (C) allele in MS.
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