| CD4+CD25+T regulatory cells (Tregs) are a subpopulation of CD4+T lymphocytes and engaged in the maintenance of immunological self-tolerance and immune homeostasis by suppressing aberrant or excessive immune responses, such as autoimmune disease and allergy. Transcriptional factor Foxp3 is the master gene for differentiation and function of regulatory T cells and plays an important role in the negative immune regulation. Dysfunction of Foxp3 gene results in fatal, early onset autoimmune disease in both human and mouse. Foxp3 mutation in mice results in lethality in hemizygous males (Xsf/Y) 16-25 days after birth, and is characterized by overproliferation of CD4+CD8-T lymphocytes, extensive multiorgan infiltration and elevation of numerous cytokines. Similarly, mutations of FOXP3 gene in human are responsible for the disease called immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX). Therefore, Foxp3 as a transcription factors plays a critical role by controlling immune homeostasis mediated by CD4+CD25+ regulatory T cells.In human CD4+CD25+ regulatory T cells, Foxp3 is expressed as two isoforms: the full-length isoform (Foxp3FL) and a splice variant lacking the exon 2 (Foxp3△2). Studies demonstrate that Foxp3 isoforms possess similar capacities to converts conventional CD4+CD25-T cells into bona fide Tregs; however, the underlying molecular mechanism remains almost entirely undefined. Recent studies indicate that Foxp3 may form a dynamic superamolecular complex with a variety of molecular partners including transcription factors and enzymatic proteins to regulate transcription, such as NFAT (nuclear factor of activated T cells), NF-кB (nuclear factorкB), AML1 (acute myeloid leukemia 1)/Runx1 (runt-related transcription factor 1), AP-1 (Activator protein 1), RORγt (retinoicacid-related orphan receptorγt) and TIP60 (Tat-interactive protein, 60 kDa). During our research, however, several reports were published, describing the interaction of Foxp3 with other proteins (AP-1, AML1/Runx1, RORγt and TIP60). However, it is not sufficient to elucidate the mechanism of how Foxp3 orchestrates the cellular and molecular programs involved in Tregs function. Therefore, the further study of unknown Foxp3 interacting protein has become a hotspot of Foxp3 transcription regulation research.In this study, Foxp3△2 was used to screen a human leukocyte yeast two-hybrid library and 40 positive clones which encode 9 different protein within the same open reading frame of GAL4 activation domain were obtained by reporter genes screening. The interaction of Foxp3△2 and ubiquitously expressed transcript (UXT) was further confirmed in yeast and in mammalian cells. The confocal fluorescence microscopy result indicates that both Foxp3△2 and UXT were predominantly colocalized in the nucleus, which raises the possibility that these proteins interact to regulate gene expression. The proline-rich domain in the N terminal of Foxp3△2 was necessary for the interaction of Foxp3△2 and UXT.In addition, we attempted to do some research for the function of Foxp3△2 and UXT interaction. Recent studies showed that Foxp3 was expressed not only in T lymphocyte, but also in tumor cells which probably took part in the immune escape of tumor. Our teams also have confirmed that Foxp3 expressed in gastric carcinoma cell line SGC-7901. However, will the overexpression of UXT influence on the expression of Foxp3? Preliminary result showed that the forced overexpression of UXT in gastric carcinoma cell line SGC-7901 could elevate the Foxp3 protein level. However, the mechanism of action was not clear.In summary, we performed yeast two-hybrid screening using Foxp3△2 as a bait protein and obtained 9 interacting proteins. Physical interaction and interacting domain of Foxp3△2 were proved in vivo. Moreover, the finding of a novel Foxp3△2 interactant will lay a basis for the elucidation of Foxp3 action. |
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