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S4 Table. Clones for transformation.

A-F) Each point represents the length (A, C, E) or width (B, D, F) of a single epidermal cell against distance of that cell from the base of the hypocotyl (x-axis). Seedlings were grown in short days for 2 (A, B), 3 (C, D) or 4 (E, F) days. Cells were measured from confocal z-stack projections using ML1:RFP PM-localized fluorescence (examples in S3G, S3H and S3I Fig). Data are from 3 cell files per seedling, for two seedlings per time point, except that wild type at 4 days and ost2-2 at 3 days data are from just one seedling each. In a replicate experiment with 2-day-old seedlings only, P35S:SAUR63:YFP:HA hypocotyls also had a greater proportion of longer cells than did wild-type hypocotyls.

A) Western blot showing presence of fusion proteins in extracts used in lipid blot experiments in Fig 5A. Arrows indicate locations of full-length SAUR63:YFP:HA and truncated SAUR6326-142:YFP:HA fusion proteins (upper arrow) and SAUR631-25:YFP:HA N-terminal domain fusion protein (lower arrow). B) Longer exposures of two lipid blots from Fig 5A. C) Mock experiment in which extracts were incubated for 70 minutes in protein extraction buffer at the indicated temperatures, and then run on a gel for western blots. For both SAUR63:YFP:HA and SAUR6326-142:YFP:HA, similar amounts of protein are present after incubation at -20 C or after incubation at 22 C, as during the lipid blot binding experiment.

A) Genomic map showing positions of genes and locations of mutations from CRISPR/Cas9 mutagenesis in the 9x-saur mutant based on the TAIR10 Arabidopsis genome annotation. The first sgRNA directed cuts in both SAUR61 and SAUR64 (green arrows), creating a deletion between them (green bar) and leaving behind a hybrid gene with a frameshift at the junction site (symbolized by a green X). The second sgRNA directed cuts in the remaining genes (blue arrows, with lighter blue indicating slight mismatches between the sgRNA and the genome), leading to deletions (blue bars) and/or frameshift mutations (blue X’s). SAUR gene names are abbreviated as S61 etc. SAUR61-SAUR68 are on chromosome 1 and SAUR75 is on chromosome 5. B,C) 5-day-old seedlings grown on 1x MS/1% Suc medium in long days. Scale bar, 1 mm. D) Hypocotyl lengths of seedlings grown for 4d in short days on 0.5x MS medium. n, 27 (wild type), 22 (9x-saur). E) Cotyledon area of seedlings grown on vertically oriented plates for 6d on MS/1% Suc medium. n, 16 (wild type), 22 (9x-saur). Graphs show means ± s.d. No statistical differences were detected between wild-type and 9x-saur mutant measurements by t-test. F) Sequences of guide RNAs used for CRISPR/Cas9-mediated mutagenesis, wild-type genes, and mutant alleles present in the 9x-saur mutant. Underlines indicate PAM motif adjacent to guide RNA target site, and any mismatches to the guide RNA sequence. Uppercase bold letters indicate insertion mutations. All alleles create frameshift mutations except for saur75-1, which has an in-frame deletion of 13 amino acids in the SAUR domain.

S3 Table. Primers used.

S2 Table. NAAIRS mutants of SAUR63 and root tortuosity index.

A) Appearance of genotypes used for crosses presented in A and in Fig 2C and 2D, grown for 6d in long days in the presence of sucrose. Scale bar, 3 mm. B) Hypocotyl lengths of seedlings of indicated genotypes grown for 4d in short days in the absence of sucrose. Plants measured were F1 progeny of crosses of transgenic lines with each other or with wild-type Columbia, and were hemizygous for the indicated transgene(s). Fig 2C shows a subset of this data. A replicate experiment gave similar results. C) Appearance of PEST:SAUR63:CerFP:HA and wild-type seedlings grown with estradiol and in the absence or presence of 15 mM LiCl. Seedlings were grown for 3d under control conditions, and then transferred to plates with estradiol and with or without 15 mM LiCl, and grown for an additional 3d before imaging. Dots mark positions of root tips at the time of transfer to estradiol plates. Scale bar, 5 mm. D) Root growth of indicated genotypes in the absence (open bars) or presence (closed bars) of 15 mM LiCl. Seedlings were grown without LiCl for 5d, transferred to plates containing 0 or 15 mM LiCl, and root growth over the next three days was measured. E) HPTS fluorescence ratios around root cells of indicated genotypes. Data are pooled from measurements taken on three different days, each normalized to the average of wild-type values on those days. Graphs show means ± s.d. Letters in graphs indicate statistical classes based on Tukey’s Honestly Significant Difference test. n, from left to right: panel B: 28, 24, 24, 30, 26, 27, 26, 25, 26, 25, 24, 20, 24, 26, 25; panel D: 21, 27, 16, 23, 19, 25, 20, 30, 18, 28, 21, 30, 23, 31, 18, 29; panel E: 26, 13, 14, 16.

A) Cotyledon area of seedlings of indicated genotypes grown on vertically oriented plates for 7d in long days in the absence (open bars) or presence (filled bars) of 1% sucrose. B) Root length of seedlings of indicated genotypes grown on vertically oriented plates for 4d in long days in the absence (open bars) or presence (filled bars) of 1% sucrose. C,F) Hypocotyl lengths (E) and hypocotyl tortuosity index [F, 1 –(distance between ends)/(contour length)] of seedlings grown on vertically oriented plates for 3d in darkness without sucrose. D) Cotyledon areas of P35S:SAUR63 and P35S:CBL11-12:SAUR6326-142 seedlings grown for 6d on vertically oriented MS 1% Suc plates. E) Hypocotyl lengths of P35S:SAUR63 and P35S:CBL11-12:SAUR6326-142 seedlings grown for 4d on vertically oriented 0.5x MS plates. Panels D and E show data for the three homozygous single-locus P35S:SAUR63 lines that differed most from wild type among seven lines analyzed. Graphs show means ± s.d. Letters in graphs indicate statistical classes based on Tukey’s Honestly Significant Difference test. n, from left to right: panel A: 26, 20, 25, 15, 22, 23, 24, 20, 22, 23, 19, 19, 21, 19, 21, 20; panel B: 19, 16, 18, 14, 20, 20, 21, 16, 21, 17, 19, 16, 18, 15, 16, 17; panel D: 77, 18, 17, 17, 19, 20; panel E: 119, 25, 24, 27, 21, 24; panels C and F: 35, 36, 45, 43, 47, 38, 41, 47.

A,B) Hypocotyl length (A) and hypocotyl tortuosity index [B, 1 –(distance between ends)/(contour length)]) of seedlings of indicated genotypes grown for 3d in darkness on plates with 0.5x MS medium. C,D) Root length (C) and root tortuosity index (D) of seedlings of indicated genotypes grown for 4d in long days on plates with 1x MS 1% Sucrose medium. E,F) Root length (E) and root tortuosity index (F) of seedlings of indicated genotypes grown for 4d in short days on plates with 0.5x MS medium. Graphs show means ± s.d. Letters in graphs indicate statistical classes based on Tukey’s Honestly Significant Difference test. n from left to right: Panels A,B: 51, 24, 26, 28, 23, 25, 25; Panels C,D: 34, 17, 23, 14, 19, 15, 16; Panels E,F: 43, 21, 18, 17, 16, 15, 17. The same genotypes were measured in panels A-F, with genotype designations shown only in panels E and F. G,H,I) Hypocotyl epidermal cells of seedlings of indicated genotypes grown in short days for 2 days, visualized with the ML1:RFP shoot epidermis plasma membrane marker. Shown are z-stack confocal image projections of the near side of the hypocotyl. Scale bar, 0.1 mm. S2 Fig shows measurements of cell sizes in this experiment.

S5 Table. Antibodies.

For self-pollinating plants to reproduce, male and female organ development must be coordinated as flowers mature. The Arabidopsis transcription factors AUXIN RESPONSE FACTOR 6 (ARF6) and ARF8 regulate this complex process by promoting petal expansion, stamen filament elongation, anther dehiscence, and gynoecium maturation, thereby ensuring that pollen released from the anthers is deposited on the stigma of a receptive gynoecium. ARF6 and ARF8 induce jasmonate production, which in turn triggers expression of MYB21 and MYB24, encoding R2R3 MYB transcription factors that promote petal and stamen growth. To understand the dynamics of this flower maturation regulatory network, we have characterized morphological, chemical, and global gene expression phenotypes of arf, myb, and jasmonate pathway mutant flowers. We found that MYB21 and MYB24 promoted not only petal and stamen development but also gynoecium growth. As well as regulating reproductive competence, both the ARF and MYB factors promoted nectary development or function and volatile sesquiterpene production, which may attract insect pollinators and/or repel pathogens. Mutants lacking jasmonate synthesis or response had decreased MYB21 expression and stamen and petal growth at the stage when flowers normally open, but had increased MYB21 expression in petals of older flowers, resulting in renewed and persistent petal expansion at later stages. Both auxin response and jasmonate synthesis promoted positive feedbacks that may ensure rapid petal and stamen growth as flowers open. MYB21 also fed back negatively on expression of jasmonate biosynthesis pathway genes to decrease flower jasmonate level, which correlated with termination of growth after flowers have opened. These dynamic feedbacks may promote timely, coordinated, and transient growth of flower organs.