Fig. 3
NCP participates in photobody biogenesis and PIF degradation in the nucleus. a Confocal images showing the subcellular localization pattern of transiently expressed NCP-CFP-FLAG in tobacco leaf cells. NCP-CFP-FLAG signals (green) were detected in chloroplasts (marked by yellow arrowheads) and the nucleus (indicated by a white arrow). The nucleus was labeled with DAPI. Chloroplasts were labeled with DAPI in the blue channel, as well as by chlorophyll autofluorescence in the red channel. Scale bars represent 10 μm. b Immunoblots showing NCP-HA-His from total, (T), nuclear (N), and plastidial (P) protein fractions of 2-day-old NCP-HA-His/ncp-10 seedlings grown in 10 μmol m−2 s−1 R light. In vitro translated (IVT) NCP-HA-His and NCPΔ48-HA-His (indicated by red arrowheads) were used as molecular size controls. The HA-tagged NCP proteins were detected via anti-HA antibodies. Ferredoxin:sulfite reductase (SiR) and RNA Pol II were used as controls for the plastidial and nuclear fractions, respectively. c Confocal images of PHYB-GFP photobodies in epidermal cells from the top one third of the hypocotyls of 4-day-old PBG and ncp-1/PBG seedlings grown in 10 μmol m−2 s−1 R light. Scale bars represent 5 μm. d Box-and-whisker plots showing the numbers of large (>0.73 μm3, left panel) and small (<0.73 μm3, right panel) photobodies per nucleus in the epidermal cells of the top one third of the hypocotyls of PBG and ncp-1/PBG seedlings. The boxes represent from the 25th to the 75th percentiles, the whiskers extend to the maximum and minimum data points, and the center lines in each box indicate the median value. Asterisks (***) indicate a statistically significant difference between the value in ncp-1/PBG and that in PBG (Student’s t-test, p ≤ 0.001). e Immunoblots showing the PIF1 and PIF3 levels in 4-day-old Col-0, ncp-10, hmr-22, PBG, and ncp-1/PBG seedlings grown in 10 μmol m−2 s−1 R light. RPN6 was used as a loading control. The source data for immunoblots in b, e and photobody analysis in d are provided in Source Data file
