CD45 Controls Interleukin-4-mediated IgE Class Switch Recombination in Human B Cells through Its Function as a Janus Kinase Phosphatase

Abstract

CD45 plays a critical regulatory role in receptor signaling through its protein tyrosine phosphatase and Janus kinase (JAK) phosphatase activities. To investigate whether CD45 also plays a regulatory role in Ig class switching in human B cells, we examined the effects of CD45 triggering on Ig class switching to IgE and its relationship with CD45 JAK phosphatase activity. Anti-CD45 triggering of CD45 significantly inhibited interleukin-4 ؉ anti-CD40-induced switch recombination in a switch recombination vector assay in stably transfected Ramos 2G6 human B cells, as well as Ig ⑀ germ-line transcription and S-S⑀ switch recombination in primary human B cells. These negative regulatory effects on Ig class switching were concomitant with the ability of CD45 to dephosphorylate the induced phosphorylation of JAK1, JAK3, and signal transducer and activator of transcription 6, but not on stress-activated/mitogenactivated protein kinases. We also showed that phosphorylated JAK1 and JAK3 were directly dephosphorylated by recombinant CD45 in vitro. These results indicate that CD45 is able to function as JAK phosphatase in human B cells and that this activity is directly associated with the negative regulation of the class switch recombination to IgE. CD45 may be an appropriate target drug for modulating IgE in allergic diseases.

Inhibition of Interleukin-4-induced Class Switch Recombination by a Human Immunoglobulin Fcγ-Fcϵ Chimeric Protein

“…DC-PCR for Human S-S⑀ Recombination-DC-PCR was performed as described (16) and was used to quantify the levels of S-S⑀ recombination in stimulated primary human B cells. Genomic DNA was digested with BglII followed by ligation under conditions favoring selfligation.…”

Human Activation-Induced Cytidine Deaminase Is Induced by IL-4 and Negatively Regulated by CD45: Implication of CD45 as a Janus Kinase Phosphatase in Antibody Diversification

“…We have previously shown that CD45, via its JAK phosphatase activity, is able to negatively regulate IgE CSR in human B cells through inhibition of IL-4 plus anti-CD40-induced Ig ⑀ germline transcription (35). However, whether CD45 plays a role in AID expression is not known.…”

“…We have previously shown that CD45 functions as a JAK phosphatase to negatively regulate IL-4-dependent IgE CSR via inhibition of Ig ⑀ germline transcription (35). To investigate whether IL-4-dependent AID induction is also regulated by CD45 signaling, we examined the effect of anti-CD45 on IL-4-dependent AID induction, as well as on Ig ⑀ germline transcripts, in BL-2 cells.…”

“…The sizes of the PCR products are expected to be 581 bp (AID1-AID4 primer pair) and 332 bp (AID3-AID4 primer pair) for the normally spliced AID. For detection of Ig ⑀ germline transcripts (⑀GTs), the primers GM3 (5Ј-AGCT GTCCAGGAACCCGACAGGGAG-3Ј) and C⑀2B (5Ј-GTTGATAGTC CCTGGGGTGTA-3Ј) were used to amplify a 518-bp PCR product (35). The primers for GAPDH have been described previously and amplify a 981-bp product (37).…”

Hematopoietic cytokine receptor signaling

“…Although Src family kinases were identified as primary molecular targets for CD45, targeted disruption of the CD45 gene leads to enhanced cytokine and IFN receptor-mediated activation of JAKs and STAT proteins. In vitro, CD45 directly dephosphorylates and binds to JAKs and negatively regulates IL-3-mediated cellular proliferation, Epo-dependent hematopoiesis, IFN-mediated antiviral responses (Irie-Sasaki et al, 2001) and IL-4/anti-CD40-induced switch recombination (Yamada et al, 2002).…”