This README_Arabidopsis_SAUR63_seedling_phenotypes_and_fluorescent_fusion_protein_localization.txt file was generated on 2022-07-13 by Jason W. Reed GENERAL INFORMATION 1. Title of Dataset: Arabidopsis SAUR63 seedling phenotypes and fluorescent fusion protein localization 2. Author Information A. Principal Investigator Contact Information Name: Jason W. Reed Institution: University of North Carolina at Chapel Hill Address: Department of Biology, CB #3280 Coker Hall, Chapel Hill, NC 27599-3280 Email: jreed@email.unc.edu B. Associate or Co-investigator Contact Information Name: Punita Nagpal Institution: University of North Carolina at Chapel Hill Address: Department of Biology, CB #3280 Coker Hall, Chapel Hill, NC 27599-3280 Email: punita@email.unc.edu 3. Date of data collection (single date, range, approximate date): between 2017 and 2022. 4. Geographic locations of data collection: Chapel Hill, NC, USA and Lyon, France 5. Information about funding sources that supported the collection of the data: This work was supported by U.S. National Science Foundation grants MCB-1615557 to Jason W. Reed and MCB-1613809 to William M. Gray, European Research Council grant 3363360-APPL under FP/2007-2013 to Yvon Jaillais, and by a visiting faculty fellowship from the Ecole Normale Superieure to Jason W. Reed. Access to the Leica sp8 confocal microscope was possible thanks to a grant from the European Research Council (ERC-2013-CoG-615739 ‘‘MechanoDevo’’) to Olivier Hamant. SHARING/ACCESS INFORMATION 1. Licenses/restrictions placed on the data: Data is licensed by the authors for use and distribution subject to citation of the original source in accordance with the Creative Commons Attribution (CC BY) license. 2. Links to publications that cite or use the data: P. Nagpal, P. H. Reeves, J. H. Wong, L. Armengot, K. Chae, N. B. Rieveschl, B. Trinidad, V. Davidsdottir, P. Jain, W. M. Gray, Y. Jaillais, J. W. Reed (2022) SAUR63 stimulates cell growth at the plasma membrane. PLoS Genetics, expected acceptance pending revision of manuscript PGENETICS-D-22-00174R1. A link to the publication will be provided upon final acceptance. 3. Recommended citation for this dataset: Reed, Jason; Nagpal, Punita; Trinidad, Brendan (2022), Arabidopsis SAUR63 seedling phenotypes and fluorescent fusion protein localization, Dryad, Dataset, https://doi.org/10.5061/dryad.8kprr4xr9 DATA & FILE OVERVIEW 1. File List: Arabidopsis SAUR63 seedling phenotypes and fluorescent fusion protein localization.xlsx This excel file has multiple tabs that have data corresponding to figures in the manuscript listed above. Each tab indicates the corresponding figure panels from the manuscript. A full list is provided below. 2. Data included in the dataset that are not in the corresponding manuscript: For Figures 2C, 4M, S4D,E, and S5B, some data are included in experiments in this dataset but were left out of the corresponding manuscript Figure, because they were not needed to support a conclusion. These are noted in the corresponding tabs. 3. Note on data: In most cases, different numbers of measurements (n) were made for different genotypes or samples. In these cases, there are blank cells below the last data entry for that sample, such that different columns have different numbers of entries. METHODOLOGICAL INFORMATION 1. Description of methods used for collection/generation of data: A link to the publication that incorporates this data will be provided upon acceptance. That publication describes methods for each experiment. 2. Methods for processing the data: The submitted data for most tabs are raw measurements and are not processed. Data for Figures S2 (calculated cell dimensions), 2I (calculated HPTS fluorescence ratios), and S5E (calculated HPTS fluorescence ratios) have already been processed, as indicated in the corresponding tabs. 3. Instrument- or software-specific information needed to interpret the data: A plugin for ImageJ was used to analyze HPTS fluorescence images in Figures 2I and S5E, as described in: Barbez E, Dunser K, Gaidora A, Lendl T, Busch W. Auxin steers root cell expansion via apoplastic pH regulation in Arabidopsis thaliana. Proceedings of the National Academy of Sciences of the United States of America. 2017;114(24):E4884-E4893. 4. Standards and calibration information, if appropriate: For length and area measurement data that are in raw format from ImageJ measurements of scanned images, the appropriate conversion factor to convert to standard units is listed with the data. 5. Environmental/experimental conditions: Conditions for each experiment are listed within the corresponding data tab. See also the list below and the Methods section of the associated publication. 6. People involved with sample collection, processing, analysis and/or submission: Data in this dataset were collected and analyzed by Punita Nagpal, Brendan Trinidad, and Jason W. Reed. DATA-SPECIFIC INFORMATION FOR: Nagpal_2022_SAUR63_dataset 1. Each table has multiple Arabidopsis thaliana genotypes as column or row headings, summarized in the following list. In some cases combinations among different transgenes or crosses among genotypes are indicated. Letters or numbers following the genotype names in the tables identify particular transgenic lines. Columbia = wild type. ost2-2 = open stomata2 mutant. 9x-saur = engineered nonuple mutant with mutations in SAUR61-SAUR68 and SAUR75. SAUR63:GUS = plant line with SAUR63 protein fused to E. coli beta-glucuronidase (GUS) reporter, expressed behind its native pSAUR63 promoter. SAUR63:GFP = plant line with SAUR63 protein fused to Green fluorescent Protein (GFP) reporter, expressed behind its native pSAUR63 promoter. p35S:SAUR63:YFP:HA = plant line with the SAUR63 protein fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63:GUS = plant line with the SAUR63 protein fused to E. coli beta-glucuronidase (GUS), expressed behind the p35S viral promoter. pp2c.d5 PP2C.D5:GFP = pp2c.d5 phosphatase gene mutant carrying a transgene of wild-type PP2C.D5 protein fused to Green Fluorescent Protein (GFP), expressed behind the native pPP2C.D5 promoter. pp2c.d5(M325K):GFP = plant line with a transgene of mutated PP2C.D5(M325K) protein (carrying a single amino acid change M325K) fused to Green Fluorescent Protein (GFP), expressed behind the native pPP2C.D5 promoter. p35S:SAUR63(1-25):YFP:HA = plant line with the N-terminal amino acids 1-25 of SAUR63 protein fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(26-142):YFP:HA = plant line with the amino acids 26-142 of SAUR63 protein fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:CBL1(1-12):SAUR63(26-142):YFP:HA = plant line with a fusion protein of i) amino acids 1-12 of Calcineurin Binding Like protein 1 (CBL1), ii) amino acids 26-142 of SAUR63 protein, iii) Yellow Fluorescent Protein (YFP) and iv) the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. cYFP:P4M = modified Yellow Fluorescent Protein fused to the P4M PI(4)P lipid-binding domain. Active MAP:mCherry:Sac1 = Myristoylation and Palmitoylation sequence (MAP) fused to mCherry fluorescent protein and Saccharomyces cerevisiae SAC1 phosphatase protein. Inactive MAP:mCherry:Sac1 = Myristoylation and Palmitoylation sequence (MAP) fused to mCherry fluorescent protein and inactive mutant Saccharomyces cerevisiae SAC1 phosphatase protein. p35S:SAUR63(m2):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m2, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(m8):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m8, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(m9):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m9, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(m11):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m11, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(m13):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m13, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:SAUR63(m15):YFP:HA = plant line with the SAUR63 protein carrying the six-amino-acid mutation m15, fused to Yellow Fluorescent Protein (YFP) and the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. ML1:RFP = marker line expressing a Red Fluorescent Protein behind the pML1 epidermal promoter p35S:SAUR63:HA = plant expressing the SAUR63 protein fused to the 3xHA (hemagglutinin) epitope tag behind the p35S viral promoter. p35S:CBL1(1-12):SAUR63(26-142):HA = plant line with a fusion protein of i) amino acids 1-12 of Calcineurin Binding Like protein 1 (CBL1), ii) amino acids 26-142 of SAUR63 protein, and iii) the HA (hemagglutinin) epitope tag, expressed behind the p35S viral promoter. p35S:CBL1(1-12):SAUR63(26-142) = plant line with a fusion protein of amino acids 1-12 of Calcineurin Binding Like protein 1 (CBL1) and amino acids 26-142 of SAUR63 protein, expressed behind the p35S viral promoter. p35S:SAUR63 = plant expressing the SAUR63 gene behind the p35S viral promoter. WAVE_1Y = marker line expressing a Yellow Fluorescent Protein fusion localized in the cytoplasm. WAVE_9Y = marker line expressing a Yellow Fluorescent Protein fusion localized to the tonoplast (vacuolar membrane). WAVE_138Y = marker line expressing a Yellow Fluorescent Protein fusion localized to the plasma membrane. 2. Listed in order are the tab names in the excel file with indication of the data in each: Figure 1E hyp lengths. Variable: Genotype. Measurement: Hypocotyl length. Experiment/condition: 4d in short days on 0.5x MS medium. n: 21-51 per genotype. Figure 1F cotyledon areas. Variable: Genotype, plate orientation. Measurement: Cotyledon area. Experiment/condition: 6d in long days on 1x MS 1% sucrose medium, on horizontal or vertical plates. n: 11-26 per genotype and orientation. Fig 1K cot sizes. Variable: Genotype. Measurement: Cotyledon area. Experiment/condition: 6d in long days on 1x MS 1% sucrose medium. n: 15-32 per genotype. Fig 2C, Fig S5B hyp epistasis. Variable: Genotype. Measurement: Hypocotyl length Experiment/condition: 4d in short days on 0.5x MS medium. n: 20-30 per genotype. Fig 2D cot areas epistasis. Variable: Genotype. Measurement: Cotyledon area. Experiment/condition: 6d in long days on 1x MS 1% sucrose medium. n: 15-27 per genotype. Fig 2I HPTS Variable: Genotype. Measurement: 458/405 fluorescence ratio. Experiment/condition: Confocal images of roots grown on 0.5X MS 1% Sucrose plates for 4-5 days. n: 17, 14, or 16 per genotype. Figure 3F,J,N,R Variable: Genotype. Measurement: Fluorescence intensity signal at each pixel along the vertical yellow lines in panels E,I,M,Q of Figure 3. Experiment/condition: Confocal images of roots grown on 0.5X MS 1% Sucrose plates for 4-5 days. n, 126 pixels per line. Fig 4I,J hyp lengths Variable: Genotype. Measurement: Hypocotyl length. Experiment/condition: 4d in short days on 0.5x MS medium. n: 19-32 per genotype. Fig 4M LiCl sensitivity Variables: Genotype, +/- LiCl. Measurement: Root growth over 3 days. Experiment/condition: 5-day-old seedlings on MS/1% Sucrose. n: 17-29 per genotype and [LiCl] combination. Figure 5H Sac1 phosphatase effect Variable: Different fluorescent fusion proteins; Active vs. inactive yeast MAP-Sac1. Measurement: Presence/absence of intracellular speckles. Experiment/condition: Nicotiana benthamiana leaf cells expressing indicated fusion proteins. n: 3 replicate experiments, 28-70 cells each per genotype and MAP-SAc1 version. Figure 7C,E naairs mutant line hyps Variable: Genotype. Measurement: Hypocotyl length Experiment/condition: 4d in short days on 0.5x MS medium. n: 17-55 per genotype. Figure 8, S12 NAAIRS line pixels Variable: Genotype. Measurement: Fluorescence intensity signal at each pixel along the vertical yellow lines in panels C,G,K,O of Figure 8 and C,G,K,O,S of Figure S12. Experiment/condition: Confocal images of roots grown on 0.5X MS 1% Sucrose plates for 4-5 days. n, 121 pixels per line. Fig S1D,E 9x saur mutant Variable: Genotype. Measurement: Hypocotyl length (D), Cotyledon area (E) Experiment/condition: D) 4d in short days on 0.5x MS medium; E) 6d on MS/1% Suc medium. n, panel D: 27 (wild type), 22 (9x-saur).n, panel E: 16 (wild type), 22 (9x-saur). Fig S2 cell measurements Variables: Genotype, Seedling age. Measurements: Length and width of single hypocotyl epidermal cells; distance of that cell from the base of the hypocotyl. Experiment/condition: 0.5x MS, short days for 2-4 days. n, 3 cell files per seedling, for 1-2 seedlings per time point. Fig S3A,B hypocotyls in darkness Variable: Genotype. Measurements: Hypocotyl contour length, distance between ends of hypocotyl. Experiment/condition: 3d in darkness on plates with 0.5x MS medium. n: 23-51 per genotype. Fig S3C,D root growth with Suc Variable: Genotype. Measurement: Root contour length, distance between ends of root. Experiment/condition: 4d in long days on plates with 1x MS 1% Sucrose medium. n: 14-34 per genotype. Fig S3E,F root growth no Suc Variable: Genotype. Measurement: Root contour length, distance between ends of root. Experiment/condition: 4d in short days on plates with 0.5x MS medium. n: 15-43 per genotype. Fig S4A cot areas Variables: Genotype, growth medium. Measurement: Cotyledon area. Experiment/condition: 7d in long days on 0.5x MS +/- 1% sucrose. n: 15-26 per genotype and growth medium. Fig S4B root lengths Variables: Genotype, growth medium. Measurement: Root length. Experiment/condition: 4d in long days on 0.5x MS +/- 1% sucrose. n: 14-21 per genotype and growth medium. Fig S4C,F dark hypocotyls Variable: Genotype. Measurements: Hypocotyl contour length, distance between ends of hypocotyl. Experiment/condition: 3d in darkness on plates with 0.5x MS medium. n: 35-47 per genotype. Fig S4D,E seedling phenotypes Variables: Genotype. Measurements: Cotyledon areas (D), hypocotyl length (E) Experiment/condition: 6d on vertically oriented MS 1% Suc plates (D), 4d on vertically oriented 0.5x MS plates (E). n: panel D: 17-77 per genotype; panel E: 19-119 per genotype. Fig S5D LiCl sensitivity Variables: Genotype, +/- LiCl. Measurement: Root growth over 3 days. Experiment/condition: 5-day-old seedlings on MS/1% Sucrose. n: 16-31 per genotype and [LiCl] concentration. Fig S5E HPTS Variable: Genotype. Measurement: 458/405 fluorescence ratio. Experiment/condition: Confocal images of roots grown on 0.5X MS 1% Sucrose plates for 4-5 days. n: 13-26 per genotype. Figure S6 WAVE line pixels Variable: Genotype. Measurement: Fluorescence intensity signal at each pixel along the vertical yellow lines in panels C,G,K,O,S. Experiment/condition: Confocal images of roots grown on 0.5X MS 1% Sucrose plates for 4-5 days. n, 121 pixels per line.