Phosphoproteomic analyses reveal that galectin-1 augments the dynamics of B-cell receptor signaling.
B-cell activation is important for mounting humoral immune responses and antibody production. Galectin-1 has multiple regulatory functions in immune cells. However, the effects of galectin-1 modulation and the mechanisms underlying the coordination of B-cell activation are unclear. To address this issue, we applied label-free quantitative phosphoproteomic analysis to investigate the dynamics of galectin-1-induced signaling in comparison with that following anti-IgM treatment. A total of 3247 phosphorylation sites on 1245 proteins were quantified, and 70-80% of the 856 responsive phosphoproteins were commonly activated during various biological functions. The similarity between galectin-1- and anti-IgM-elicited B-cell receptor (BCR) signaling pathways was also revealed. Additionally, the mapping of the 149 BCR-responsive phosphorylation sites provided complementary knowledge of BCR signaling. Compared to anti-IgM induction, the phosphoproteomic profiling of BCR signaling, along with validation by western blot analysis and pharmacological inhibitors, revealed that the activation of Syk, Btk, and PI3K may be dominant in galectin-1-mediated activation. We further demonstrated that the proliferation of antigen-primed B cells was diminished in the absence of galectin-1 in an animal model. Together, these findings provided evidence for a new role and insight into the mechanism of how galectin-1 augments the strength of the immunological synapse by modulating BCR signaling.


CONCLUSIONS
The current study revealed the first systematic phosphorylation-mediated signaling network and its dynamics in B cell activation. The comparative phosphoproteomic analysis on the dynamics of galectin-1 induced activation profiles not only showed that exogenously added galectin-1 augmented B-cell activation but also revealed its relatively enhanced activation in PI3K pathway. Together with proliferation assay, we further delineated that galectin-1 is important for B-cell proliferation in response to antigen challenge. Our phosphoproteomic study reveals a new role for galectin-1 in augmenting the strength of immunological synapse by modulating BCR signaling.
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Chih-Ming
CM
Tsai
Institute of Chemistry, Academia Sinica, Taipei 115, Taiwan.
Hsin-Yi
HY
Wu
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Tseng-Hsiung
TH
Su
Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan.
Chu-Wei
CW
Kuo
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Han-Wen
HW
Huang
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; Baylor Institute for Immunology Research, Dallas, TX 75204, United States.
Cheng-Han
CH
Chung
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Chien-Sin
CS
Chen
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan.
Kay-Hooi
KH
Khoo
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; Institute of Chemistry, Academia Sinica, Taipei 115, Taiwan. Electronic address: yujuchen@gate.sinica.edu.tw.
Yu-Ju
YJ
Chen
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan. Electronic address: kuoilin@gate.sinica.edu.tw.
Kuo-I
KI
Lin
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000494:pharmacology; D001402:B-Lymphocytes / Q000502:physiology*; D037483:Galectin 1 / Q000172:deficiency / Q000502:physiology*; D008213:Lymphocyte Activation / Q000187:drug effects*; D008810:Mice, Inbred C57BL; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000302:isolation & purification; D040901:Proteomics / Q000379:methods; D011947:Receptors, Antigen, B-Cell / Q000502:physiology*; D015398:Signal Transduction
D000888:Antibodies, Anti-Idiotypic; D037483:Galectin 1; D010750:Phosphoproteins; D011947:Receptors, Antigen, B-Cell; C044203:anti-IgM; D019869:Phosphatidylinositol 3-Kinases
B-cell activation; B-cell receptor; Galectin-1; Phosphoproteome