IRF4 Deficiency Leads to Altered BCR Signalling Revealed by Enhanced PI3K Pathway, Decreased SHIP Expression and Defected Cytoskeletal Responses

Abstract

The graded expression of transcription factor interferon regulatory factor 4 (IRF4) regulates B cell development and is critical for plasma cell differentiation. However, the mechanisms, by which IRF4 elicits its crucial tasks, are largely unknown. To characterize the molecular targets of IRF4 in B cells, we established an IRF4-deficient DT40 B cell line. We found that in the absence of IRF4, the expression of several molecules involved in BCR signalling was altered. For example, the expression of B cell adaptor for PI3K (BCAP) was upregulated, whereas the SHIP (SH2-containing Inositol 5´-Phosphatase) expression was downregulated. These molecular unbalances were accompanied by increased BCR-induced calcium signalling, attenuated B cell linker protein (BLNK) and ERK activity and enhanced activity of PI3K/protein kinase B (Akt) pathway. Further, the IRF4-deficient cells showed dramatically diminished cytoskeletal responses to anti-IgM cross-linking. Our results show that IRF4 has an important role in the regulation of BCR signalling and help to shed light on the molecular mechanisms of B cell development and germinal centre response.

BLIMP‐1 is insufficient to induce antibody secretion in the absence of IRF4 in DT40 cells

“…The resulting DKO cells were verified to lack both the IRF4 and BCL6 expression (Figure A,B). BCL6 and IRF4 single knockouts were described previously . The DKO cell line had the same growth characteristics as IRF4 single knockout and WT cell lines (Figure ).…”

“…BCL6 and IRF4 single knockouts were described previously. 4,17 The DKO cell line had the same growth characteristics as IRF4 single knockout and WT cell lines ( Figure S1). As BCL6KO cells secrete antibodies, 4 we analysed whether DKO cells lacking both IRF4 and BCL6 also secrete.…”

“…To obtain DKO cell line, the BCL6 ‐encoding alleles were removed with previously described targeting vectors from IRF4 ‐deficient cell line (IRF4KO) . The DT40 cell line carries three alleles of IRF4 due to trisomy of IRF4 harbouring chromosome 2 .…”

“…To obtain DKO cell line, the BCL6-encoding alleles were removed with previously described targeting vectors 4 from IRF4-deficient cell line (IRF4KO). 17 The DT40 cell line carries three alleles of IRF4 due to trisomy of IRF4 harbouring chromosome 2. 18 Prior to the targeting of BCL6, the expression vector Mer-Cre-Mer-Puro 19 was stably transfected into IRF4KO cells to allow p-lox-mediated removal of Bsr and Neo selection cassettes.…”

Interferon Regulatory Factor 4 Correlated With Immune Cells Infiltration Could Predict Prognosis for Patients With Lung Adenocarcinoma

“…In detail, we found that all three tumor infiltrating B lymphocytes (TIL-B) subsets, activated or effector memory CD8+ and activated or effector memory CD4+T cells were significantly correlated with IRF4 expression (|r| > 0.3 and P < 0.001) (Figure 4). In fact, several studies showed that IRF4 plays a key role in diverse pathways related immune cell (21)(22)(23)(24)(25). Above all, high IRF4 expression might be reflect a state having predominantly tumor-specific TILs which play an important role in tumor control and prevent tumor progression (26).…”