Identification and Characterization of the Amphioxus Lck and Its Associated Tyrosine Phosphorylation-Dependent Inhibitory LRR Receptor

Copyright © 2021 Zhou, Xiao, Zhan, Qu, Mou, Deng, Zhang, Lan, Huang and Li..

Amphioxus (e.g., Branchiostoma belcheri, Bb) has recently emerged as a new model for studying the origin and evolution of vertebrate immunity. Mammalian lymphocyte-specific tyrosine kinase (Lck) plays crucial roles in T cell activation, differentiation and homeostasis, and is reported to phosphorylate both the ITIM and ITSM of PD-1 to induce the recruitment of phosphatases and thus the inhibitory function of PD-1. Here, we identified and cloned the amphioxus homolog of human Lck. By generating and using an antibody against BbLck, we found that BbLck is expressed in the amphioxus gut and gill. Through overexpression of BbLck in Jurkat T cells, we found that upon TCR stimulation, BbLck was subjected to tyrosine phosphorylation and could partially rescue Lck-dependent tyrosine phosphorylation in Lck-knockdown T cells. Mass spectrometric analysis of BbLck immunoprecipitates from immunostimulants-treated amphioxus, revealed a BbLck-associated membrane-bound receptor LRR (BbLcLRR). By overexpressing BbLcLRR in Jurkat T cells, we demonstrated that BbLcLRR was tyrosine phosphorylated upon TCR stimulation, which was inhibited by Lck knockdown and was rescued by overexpression of BbLck. By mutating single tyrosine to phenylalanine (Y-F), we identified three tyrosine residues (Y539, Y655, and Y690) (3Y) of BbLcLRR as the major Lck phosphorylation sites. Reporter gene assays showed that overexpression of BbLcLRR but not the BbLcLRR-3YF mutant inhibited TCR-induced NF-κB activation. In Lck-knockdown T cells, the decline of TCR-induced IL-2 production was reversed by overexpression of BbLck, and this reversion was inhibited by co-expression of BbLcLRR but not the BbLcLRR-3YF mutant. Sequence analysis showed that the three tyrosine-containing sequences were conserved with the tyrosine-based inhibition motifs (ITIMs) or ITIM-like motifs. And TCR stimulation induced the association of BbLcLRR with tyrosine phosphatases SHIP1 and to a lesser extent with SHP1/2. Moreover, overexpression of wild-type BbLcLRR but not its 3YF mutant inhibited TCR-induced tyrosine phosphorylation of multiple signaling proteins probably via recruiting SHIP1. Thus, we identified a novel immunoreceptor BbLcLRR, which is phosphorylated by Lck and then exerts a phosphorylation-dependent inhibitory role in TCR-mediated T-cell activation, implying a mechanism for the maintenance of self-tolerance and homeostasis of amphioxus immune system and the evolutionary conservatism of Lck-regulated inhibitory receptor pathway.

Medienart:

E-Artikel

Erscheinungsjahr:

2021

Erschienen:

2021

Enthalten in:

Zur Gesamtaufnahme - volume:12

Enthalten in:

Frontiers in immunology - 12(2021) vom: 09., Seite 656366

Sprache:

Englisch

Beteiligte Personen:

Zhou, Jiatao [VerfasserIn]
Xiao, Zhihui [VerfasserIn]
Zhan, Yanli [VerfasserIn]
Qu, Xuemei [VerfasserIn]
Mou, Sisi [VerfasserIn]
Deng, Chong [VerfasserIn]
Zhang, Tianxiang [VerfasserIn]
Lan, Xin [VerfasserIn]
Huang, Shengfeng [VerfasserIn]
Li, Yingqiu [VerfasserIn]

Links:

Volltext

Themen:

Amphioxus
Amphioxus Lck associated LRR (BbLcLRR)
Biomarkers
Costimulatory and Inhibitory T-Cell Receptors
EC 2.7.10.2
Immunoreceptor tyrosine-based inhibitory motif
Interleukin-2
Journal Article
Leucine rich repeat receptor (LRR)
Lymphocyte Specific Protein Tyrosine Kinase p56(lck)
Lymphocyte-specific tyrosine kinase (Lck)
Receptors, Antigen, T-Cell
Research Support, Non-U.S. Gov't
TCR-mediated T cell activation
Tyrosine phosphorylation
Tyrosine phosphorylation-dependent inhibitory immunoreceptor

Anmerkungen:

Date Completed 28.09.2021

Date Revised 28.09.2021

published: Electronic-eCollection

Citation Status MEDLINE

doi:

10.3389/fimmu.2021.656366

funding:

Förderinstitution / Projekttitel:

PPN (Katalog-ID):

NLM326971475