Journal: bioRxiv

Article Title: T cells compete via reverse MHC class I signaling at the synapse with dendritic cells to secure Golgi recruitment for activation

doi: 10.64898/2026.05.13.724773

Figure Lengend Snippet: (A) Scheme of synapse formation between murine Flt3L-differentiated bone marrow-derived DCs (BMDC) and OT-I T cells. LPS: lipopolysaccharide; CpG-B: oligodeoxynucleotide; OVA: ovalbumin; IC: immune complex; SIINFEKL: ovalbumin (257-264) peptide epitope; PFA: paraformaldehyde. (B) Representative confocal microscopy images of the BMDC-T cell synapses immunostained for cis-Golgi marker GM-130 (magenta), γ-tubulin (yellow), DAPI (blue) and α-tubulin (green). White arrowheads: MTOC polarization. Scale bar, 10 µm. (C) Scheme of quantification of MTOC polarization: the distance between the MTOC and the DC-T cell interface (a) was divided by the diameter of the DC (b). (D) Quantification of MTOC polarization with unstimulated BMDCs or TLR ligand-stimulated BMDCs unloaded or loaded with OVA, OVA-Immunocomplexes or SIINFEKL. For each of the 3 mouse cultures, 30-35 cells were analysed. Mean ± standard error of the mean, Kruskal-Wallis test, Dunn’s multiple comparison **P<0.01, ****P<0.0001 (E) Same as panel D, but now displayed as percentage DCs with polarized MTOC (defined as polarization index < 0.3) for individual mice. (F) Electron micrographs of two different immunological synapses between T cell and BMDC, showing MTOC, Golgi apparatus (GA), and immunological synapse (IS); Black boxes indicate magnified areas. Scale bars, 1 µm.

Article Snippet: From the femurs and tibias, bone marrow derived stem cells were obtained and cultured in RPMI-1640 medium with 10% fetal bovine serum (FBS, 758093, Greiner bio-one), 1% antibiotic-antimycotic (15240-062, Gibco), 1% ultraglutamine (BE17-605E/U1, Lonza Bioscience), 50 μM β-mercaptoethanol (60-24-2, Sigma-Aldrich) and 200 ng/ml human Flt3L (130-096-479, Miltenyi Biotec) for eight days at 37oC in an incubator with 10% CO 2 .

Techniques: Derivative Assay, Confocal Microscopy, Marker, Comparison

Journal: bioRxiv

Article Title: T cells compete via reverse MHC class I signaling at the synapse with dendritic cells to secure Golgi recruitment for activation

doi: 10.64898/2026.05.13.724773

Figure Lengend Snippet: (A) Scheme of MTOC polarization by antibody- or protein domain-coated bead stimulation of murine Flt3L-differentiated bone marrow-derived DCs (BMDCs). LPS: lipopolysaccharide; CpG-B: oligodeoxynucleotide; PFA: paraformaldehyde. (B) Representative immunofluorescence images of the BMDC-bead conjugates. Cells were immunostained for cis-Golgi marker GM-130 (magenta), γ-tubulin (yellow), DAPI (blue) and α-tubulin (green). Beads were coated with isotype or antibody against MHC class I. Straight outline: T cell; dotted outline BMDC; white arrowhead: MTOC polarization. Scale bar, 10 µm. (C) Quantification of MTOC polarization in BMDCs incubated with beads coated with antibodies against MHC class I, MHC class II, or ICAM-1, or with LFA-I or CD28 protein domains. One-way ANOVA Kruskal-Wallis test with Dunn’s Multiple Comparisons. (D) Quantification of MTOC polarization in BMDCs incubated with beads coated with antibodies against MHC class I treated with nocodazole inhibitor targeting the microtubules (Mt), the Src-I1 and PP1 inhibitors targeting tyrosine kinase Src, and the phospholipase C inhibitor (U73122; U22) and its negative control compound (U73343; U43) and the PKC inhibitor Gö6983. One-way ANOVA with Dunnett’s multiple comparison, Mean ± standard error of the mean (**P<0.01; ****P<0.0001).

Article Snippet: From the femurs and tibias, bone marrow derived stem cells were obtained and cultured in RPMI-1640 medium with 10% fetal bovine serum (FBS, 758093, Greiner bio-one), 1% antibiotic-antimycotic (15240-062, Gibco), 1% ultraglutamine (BE17-605E/U1, Lonza Bioscience), 50 μM β-mercaptoethanol (60-24-2, Sigma-Aldrich) and 200 ng/ml human Flt3L (130-096-479, Miltenyi Biotec) for eight days at 37oC in an incubator with 10% CO 2 .

Techniques: Derivative Assay, Immunofluorescence, Marker, Incubation, Negative Control, Comparison