Fig. 1. Light impacts the transcriptional status of cotyledon nuclei.

A Analyses in this panel employed cotyledons dissected from dark- or light-grown seedlings. B RNA Pol II activity in individual cotyledon nuclei was determined by flow cytometry as the ratio of fluorescent signals after immunolabeling of its Serine 2 phosphorylated (S2P) and non-phosphorylated (NP) forms for each ploidy class (Supplementary Fig. S1A). The intensity of individual signals was measured consistently with the same settings and is plotted as arbitrary units. For the intensity ratio, the mean of the medians of the 2 C Dark samples was arbitrarily set to 1. Each dot represents the measure in a single nucleus. Three independent biological replicates were used, each comprising cotyledons from several seedlings. Values from at least 437 nuclei were aggregated (i.e., the number of measured nuclei in a single ploidy class of a single biological replicate). The p-value from a two-sided t test (see “Methods”) is given between dark and light for each ploidy class (Source data are provided as a Source Data file). C Live imaging of cotyledon mesophyll nuclei expressing RNA Pol II S2P mintbody-GFP and H2B-mRuby reporting local signals of the elongating RNA Pol II and chromatin. 3D projections of deconvolved images showing the levels of RNA Pol II S2P (green), H2B-mRuby (magenta), and their ratio (“Fire”)³¹. See Supplementary Fig. S1C for further details. Source data are provided as a Source Data file. D Quantification of RNA Pol II-S2P and H2B total nuclear signals calculated as the sum of pixel intensity in each analyzed nucleus, and RNA Pol II-S2P/H2B signal ratio calculated as the mean of the intensity ratio across the pixels of each nucleus (n nuclei = 14, n plants = 3 and n nuclei = 11, n plants = 5 for dark- and light-grown seedlings, respectively). The p-values are calculated from a two-sided Welch’s t test comparing the means of biological replicates (individual plants) between Dark and Light. E RT-qPCR analysis of AtRS31 pre-mRNA accumulation. The bars indicate the ratio of the signal obtained with primer pairs recognizing a proximal and a distal position on the AtRS31 pre-mRNA. Error bars represent the standard deviation between two independent biological replicates. Source data are provided as a Source Data file. F ChIP-seq meta-profiles of RNA Pol II S2P and H2Bub over the ensemble of genes marked in Dark or Light samples.