Cell - 8 September 2016

Figure S5. CPR5 Function Requires Homo-oligomerization, Related toFigure 5
(A) qRT-PCR determined relative expression levels ofCPR5during ETI triggered byPsm/AvrRpt2at OD600nm= 0.1. Samples were collected until 8 hr post
inoculation (hpi) before ETI-induced PCD occurred. Data are presented as mean±SDM (n = 3 biological replicates). Similar results were obtained in two in-
dependent experiments.
(B) GFP-CPR5 protein levels in35S:GFP-CPR5transgenic plants during ETI triggered byPsm/AvrRpt2at OD600nm= 0.1 as measured by western blot using the
GFP antibody.
(C) Fluorescence intensity measurement of GFP-CPR5 in the NE in35S:GFP-CPR5transgenic plants during ETI triggered byPsm/AvrRpt2at OD600nm= 0.1. Data
are presented as mean±SDM (n = 15 cells for each time point and treatment).
(D) CPR5 homomeric interaction is mediated by its N-terminal domain. CPR5-N, rather than CPR5-C, mediated the homomeric interaction. In vitro pull-down
assays were performed using wheat germ-synthesized recombinant proteins. Immunoprecipitation was performed using HA antibody-conjugated agarose
beads. Immunoblotting was performed using GFP and HA antibodies. Stars indicate non-specific signals from immunoglobulins.
(E) The G120D mutation compromises heteromeric interactions of CPR5. BiFC was performed by transiently coexpressing nYFP-CPR5/cYFP-CPR5 or nYFP-
G120D/cYFP-G120D inN. benthamiana. The BiFC intensity in the NE was normalized using averaged expression levels of YFP-CPR5/G120D measured in
separate experiments and plotted as relative values (upper panel). Data are presented as mean±SDM (n = 15 cells for each BiFC combination). The number of
Z-whorl structures in each cell was measured when WT CPR5 or G120D mutant was transiently expressed inN. benthamiana(lower panel). Data are presented as
mean±SDM (n = 15 cells for each construct). Student’s t test and non-parametric test were used, **** p-value < 0.0001.