Extended Data Fig. 1: Characterization of wild-type and TCR-transgenic T_β-syn cells in vitro and in vivo.

a, Identification of the immunogenic β-synuclein peptides in Lewis rats. Screening of a β-synuclein peptide library covering the full-length protein for identification of immunogenic peptides. OVA, MBP, concanavalin A (ConA) and a mix of all peptides were used as control. The proliferation of αβTCR⁺CD4⁺ T cells isolated from β-synuclein-immunized wild-type rats was measured by flow cytometry. Data are mean + s.e.m. from two independent experiments. b, Antigen response of established effector T cell lines. Wild-type T_β-syn or T_MBP cell lines were cultured in the absence of antigen (No) or stimulated with ConA or the indicated antigens for 72 h. Proliferation was determined by ³H-thymidine incorporation. Data are mean + s.e.m. Representative data of at least four independent experiments. c, Phenotype of T_β-syn and T_MBP cell lines. Expression of αβTCR, CD4, CD8 and of surface activation markers CD134 and CD25 in cultured wild-type T_β-syn and T_MBP cells. Flow cytometry was performed two days (blast T cells) and seven days (resting T cells) after antigen stimulation. IgG, isotype control. d, Histological quantification of T_β-syn and T_MBP cells in cortical grey matter and corpus callosum (white matter) at the peak of T cell infiltration. Data are mean + s.e.m., n = 5 from 2 independent experiments. Unpaired two-tailed t-test. ***P < 0.001. e, Wild-type T_β-syn cells induce clinical disease. Representative disease course upon intravenous transfer of wild-type T_β-syn or T_MBP cells. Data are mean + s.d., n = 6–8 per group. Representative data of at least six independent experiments. f, Photographs illustrating frontlimb paralysis induced by T_β-syn cells and classical EAE hindlimb paralysis induced by T_MBP cells. g, Expression of Trbv genes in wild-type T_β-syn cells isolated ex vivo at the onset of T_β-syn cell brain infiltration. Relative frequency was calculated in T_β-syn cells from blood and CNS and expressed as ratio of reads mapping to a particular Trbv genomic segment to the number of reads mapping to the constant region of the TCR β chain encoded by Trbc1 and Trbc2 (RNA-seq analysis, n = 3 per group). Data are mean + s.d. h, Production of IFNγ by lymph node cells isolated from naive wild-type and TCR-transgenic (Tg) donors in response to β-syn_93–111 measured by ELISA. Data are mean ± s.d. i, Immunization of TCR-transgenic rats with β-syn_93–111 results in a monophasic clinical disease. Bars and lines indicate clinical score and relative weight change, respectively (mean ± s.d.). Representative data from two independent experiments (n = 10). j, Expression of surface markers on wild-type T_β-syn (blue) and transgenic T_β-syn (green) cell lines. k, TCR repertoire in transgenic T_β-syn cell lines is dominated by transgenic TCR Vβ8.3 chain expression. j, k, Flow cytometry data. l, Expression of Trbv genes in transgenic T_β-syn cell lines determined by RNA-seq analysis. Analysis and normalization of data as in g. n = 3 per group. m, n, Proliferative response measured by ³H-thymidine incorporation (m) and IFNγ secretion measured by ELISA (n) in wild-type T_β-syn (blue) and transgenic T_β-syn (green) cell lines stimulated with the indicated amount of cognate (β-syn) or non-cognate (MBP, OVA) antigens. Data are mean ± s.d. o, Intracellular staining for IFNγ and IL-17 in T_MBP and T_β-syn cell lines stimulated with anti-CD3 monoclonal antibodies. Representative data from at least three independently established T cell lines. p, Quantification of transgenic T_β-syn cells infiltrating the cortical grey matter or the corpus callosum (white matter) measured by flow cytometry at the peak of T cell infiltration (n = 5). q, Clinical disease induced by transgenic T_β-syn effector cells (n = 15). p, q, Data are mean + s.e.m.