TOSO interacts with SYK and enhances BCR pathway activation in chronic lymphocytic leukemia.

ABSTRACT: BACKGROUND:TOSO, also named Fas inhibitory molecule 3 (FAIM3), has recently been identified as an immunoglobulin M (IgM) Fc receptor (Fc?R). Previous studies have shown that TOSO is specifically over-expressed in chronic lymphocytic leukemia (CLL). However, the functions of TOSO in CLL remain unknown. The B-cell receptor (BCR) signaling pathway has been reported to be constitutively activated in CLL. Here, we aimed to investigate the functions of TOSO in the BCR signaling pathway and the pathogenesis of CLL. METHODS:We over-expressed TOSO in B-cell lymphoma cell lines (Granta-519 and Z138) by lentiviral transduction and knocked down TOSO by siRNA in primary CLL cells. The over-expression and knockdown of TOSO were confirmed at the RNA level by polymerase chain reaction and protein level by Western blotting. Co-immunoprecipitation with TOSO antibody followed by liquid chromatography coupled with tandem mass spectrometry (IP/LCMS) was used to identify TOSO interacting proteins. Western blotting was performed to detect the activation status of BCR signaling pathways as well as B-cell lymphoma 2 (BCL-2). Flow cytometry was used to examine the apoptosis of TOSO-over-expressing B lymphoma cell lines and TOSO-down-regulated CLL cells via the staining of Annexin V and 7-AAD. One-way analyses of variance were used for intergroup comparisons, while independent samples t tests were used for two-sample comparisons. RESULTS:From IP/LCMS, we identified spleen tyrosine kinase (SYK) as a crucial candidate of TOSO-interacting protein and confirmed it by co-immunoprecipitation. After stimulation with anti-IgM, TOSO over-expression increased the phosphorylation of SYK, and subsequently activated the BCR signaling pathway, which could be reversed by a SYK inhibitor. TOSO knockdown in primary CLL cells resulted in reduced SYK phosphorylation as well as attenuated BCR signaling pathway. The apoptosis rates of the Granta-519 and Z138 cells expressing TOSO were (8.46?±?2.90)% and (4.20?±?1.21)%, respectively, significantly lower than the rates of the control groups, which were (25.20?±?4.60)% and (19.72?±?1.10)%, respectively (P?

SUBMITTER: Zhang YR 

PROVIDER: S-EPMC7478576 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature