Intrinsic defect in B-lymphoblastoid cell lines from patients with X-linked lymphoproliferative disease type 1. II. receptor-mediated Akt/PKB and ERK1/2 activation and transcription factors expression profile.

BACKGROUND X-linked lymphoproliferative disease type 1 (XLP1) belongs to genetically determined primary immunodeficiency syndromes with mutations in SH2D1A/DSHP/SAP gene. The dramatic increase of the risk of B-cell lymphoma development in XLP1 patients is linked not only to SAP deficiency of NK, NKT and T cells, but probably to the impairment of B cell differentiation. AIM To analyze the receptor-mediated Akt/PKB and ERK1/2 phosphorylation and expression of transcription factors that are involved in B cell maturation in EBV-transformed B-lymphoblastoid cell lines (B-LCLs) from XLP1 patients (XLP B-LCLs) in comparison with conventional B-LCLs. METHODS Studies were performed on EBV-transformed XLP B-LCLs IARC 739, SC-XLP and RP-XLP in comparison with SAP-negative B-LCL T5-1 and SAP-positive B-LCL MP-1. Western blot analysis was used for evaluation of Akt (Ser473) and ERK1/2 (Thr202/Tyr204) phosphorylation in response to ligation of CD150, CD40, and IgM cell surface receptors. The expression levels of transcription factors IRF4, IRF8, BCL6, BLIMP1, SPIB, PU.1 and MITF were assessed using quantitative RT-PCR. RESULTS It was shown that SAP deficiency in XLP B-LCL did not abrogate CD150-mediated Akt and ERK1/2 phosphorylation. At the same time, ligation of CD150 or IgM affects kinetics and amplitude of ERK1/2 activation. In XLP B-LCL the CD150 signaling with IgM coligation play the dominant role in both Akt and ERK1/2 phosphorylation. We found that significantly reduced IRF4, IRF8 and PU.1 expression levels are the key features of XLP B-LCLs. CONCLUSION XLP B-LCLs and conventional B-LCLs have differences in kinetics and amplitude of Akt and ERK1/2 phosphorylation. Analysis of transcription factors profile revealed the distinguishing features of XLP B-LCLs with SAP deficiency that may impair B cell differentiation.Key Words: B-lymphoblastoid cell lines, X-linked lymphoproliferative disease type 1, CD150, CD40, BCR, Akt/PKB, ERK1/2, transcription factors.