Fig. 1: Precursor granules form in close proximity to chloroplasts and reduce photosynthetic performance. | Nature Communications

Fig. 1: Precursor granules form in close proximity to chloroplasts and reduce photosynthetic performance.

From: Chloroplast precursor protein preClpD overaccumulation triggers multilevel reprogramming of gene expression and a heat shock-like response

Fig. 1

a Immunoblots showing preClpD-GFP accumulation in the t-i-ClpD-GFP line detected by anti-GFP antibodies. The numbers below the immunoblot represent the accumulation of preClpD-GFP relative to mature ClpD-GFP (in %). Seven-d-old seedlings were induced for the days indicated and whole seedlings were harvested for analyses. Actin served as loading control. pre: preClpD-GFP, mat: mature ClpD-GFP. b Representative images showing that preClpD-GFP granules closely associated with chloroplasts (see also Supplementary Fig. 1g). Seven-d-old t-i-ClpD-GFP seedlings were induced for 1 or 2 days, the newly emerged true leaves were analyzed. The boxed areas in the middle panel are magnified and shown below. Scale bars, 20 μm for the upper panel, and 5 μm for the middle and bottom panels. c Fluorescence intensity plots along the magenta lines in (b) measured by using Fiji software. d Colocalization of GFP and chlorophyll fluorescence of the boxed area in (b) analyzed by Coloc 2 in Fiji software. PCC, Pearson’s colocalization coefficient; MCC, Manders’ colocalization coefficient (tM1 and tM2 are the proportion of colocalized signal to GFP and chlorophyll fluorescence, respectively). e Proportion of cells containing preClpD-GFP granules in different transgenic lines at 2 days of induction. Data are presented as means ± s.d. (n = 10 images from 2 independent experiments). Two-tailed Student’s t-tests were performed to determine the significance of differences. f PreClpD accumulation after 2 days of heat stress at 37 °C. Newly emerged true leaves were harvested for analyses. The numbers below the immunoblot represent the accumulation of preClpD relative to the mature ClpD (in %). Pre, preClpD; mat, mature ClpD; asterisks, nonspecific cross-reactions. Actin served as loading control. g The maximum photochemical efficiency of PS II (Fv/Fm) of t-i-ClpD-GFP and untransformed toc75III seedlings after 4 days of induction. Scale bars, 1 cm. h Light-response curves of chlorophyll a fluorescence parameters after 2 days of induction. Y(II), quantum yield of PSII; ETR (II), electron transfer rate of PSII. The data are means ± s.d. (n = 30 biological replicates). In (f–h), 6-d-old seedlings were used for induction. Source data are provided as a Source Data file.

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