Supplementary Materials

Dynamic signaling by T follicular helper cells during germinal center B cell selection

Ziv Shulman, Alexander D. Gitlin, Jason S. Weinstein, Begoña Lainez, Enric Esplugues, Richard A. Flavell, Joseph E. Craft, Michel C. Nussenzweig

Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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  • Materials and Methods
  • Figs. S1 to S8
  • Full Reference List

Images, Video, and Other Other Media

Movie S1
Dynamics of T and B cell interactions in the GC. Collapsed 4D images depicting OT-II DsRed+ T cells (red), B1-8hi GFP+Ly75+/+ B cells (light blue) and T-B contacts (colocalized pixels, green) in the germinal center after αDEC-CS (control) injection. Scale bar, 20 µm.
Movie S2
Dynamics of T and B cell interactions in the GC during selection. Collapsed 4D images depicting OT-II DsRed+ T cells (red), B1-8hi GFP+Ly75+/+ B cells (light blue) and T-B contacts (colocalized pixels, green) in the germinal center after αDEC-OVA injection. Scale bar, 20 µm.
Movie S3
TFH cell interactions with B cells in GCs are transient. High magnification of a part of movie S2. Scale bar, 20 µm.
Movie S4
Polyclonal follicular B cells within the mantle zone are unaffected by αDEC-OVA injection. Collapsed 4D images depicting OT-II GFP+ T cells (green), B1-8hi CFP+Ly75+/+ B cells (blue), follicular dendritic cells (FDCs; red, non-mobile) and follicular DsRed+ B cells (red, mobile) in GCs after αDEC-CS or αDEC-OVA injection. The FDCs were labeled by injection of NP conjugated to tdTomato. Scale bar, 30 µm.
Movie S5
T cell interactions with selected and non-selected GC B cells. Collapsed 4D images depicting OT-II DsRed+ T cells (red), B1-8hi GFP+Ly75-/-B cells (green) and CFP+Ly75+/+ cells (blue) after injection of αDEC-OVA. T-B contacts (colocalized pixels) of T cells with B1-8hiLy75+/+ and Ly75-/- B cells are shown in yellow and gray, respectively. Scale bar, 20 µm.
Movie S6
In vivo imaging of GCaMP3 expressing T cells. Collapsed 4D images depicting OT-II GCaMP3+ T cells (green) in the T zone after injection of OVA323-339 peptide. Myeloid autofluorescent cells are in white. False color representation of one cell is shown at the bottom left. Scale bar, 20 µm.
Movie S7
TFH cell Ca2+ signaling during B cell selection. Collapsed 4D images depicting OT-II DsRed+ GCaMP3+ T cells (red, Ca2+ low and yellow, Ca2+ high) and B1-8hi CFP+ Ly75+/+ B cells (light blue) in the GC during control or after αDEC-OVA treatment. Myeloid autofluorescent cells are in white. Scale bar, 15 µm.
Movie S8
An example of TFH cell Ca2+ signaling in the GC. Collapsed 4D images depicting OT-II DsRed+ GCaMP3+ T cell (red, Ca2+ low and yellow, Ca2+ high) in the GC without injection of αDEC-OVA. The motile TFH cell slows down while displaying high levels of intracellular Ca2+. Scale bar, 15 µm.
Movie S9
An example of Ca2+ signaling in motile GC TFH cell. Collapsed 4D images depicting OT-II DsRed+ GCaMP3+ T cell (red, Ca2+ low and yellow, Ca2+ high) in the GC without injection of αDEC-OVA. Scale bar, 15 µm.
Movie S10
An example of a TFH cell transiently mobilizing Ca2+ and slowing down during contact with a GC B cell. Collapsed 4D images depicting an OT-II GCaMP3+ T cells (gray) and a B1-8hi tdTomato+Ly75+/+ B cells (red) in the GC after injection of αDEC-OVA. Scale bar, 10 µm.
Movie S11
An example of TFH cell Ca2+ signaling during GC B cell contact. Collapsed 4D images depicting an OT-II GCaMP3+ T cell (gray) interacting with a B1-8hi tdTomato+Ly75+/+ B cell (red) in the GC after injection of αDEC-OVA. Scale bar, 10 µm.
Movie S12
An example of multiple Ca2+ mobilization events in TFH cell during a contact with a GC B cell. Collapsed 4D images depicting OT-II GCaMP3+ T cell (gray) and B1-8hi tdTomato+Ly75+/+ B cell (red) in the GC after injection of αDEC-OVA. Scale bar, 10 µm.