DEV035436 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Interactions between pan and the weak lfy-5 allele. (A) pan-3 mutant flower. (B) Flower of the weak lfy-5 allele, which usually bears four sepals, two to three petals, two to three stamens (that are sometimes sepalloid) and an apparently normal gynoecium. These flowers are only infrequently sterile. (C) pan-3 lfy-5 flower bearing four sepals, two petals, four stamens and two unfused carpels in the center (arrowheads). Secondary flowers are frequently visible in the axils of the first-whorl sepals (arrow). (D) The same pan lfy flower shown in C with some organs removed to expose ectopic fifth-whorl structures growing internal to the fourth-whorl carpels (arrowheads). These flowers are almost always sterile.

• Supplemental Figure S2 -

  Fig. S2. In situ localizations of WUS mRNA in pan flowers. (A) View of a pan-3 shoot apex hybridized with a WUS probe. Expression in the shoot apical meristem is indicated (arrowhead). (B-G) Serial sections of a close-up view of the indicated flower (dashed box in A). No WUS expression is visible at the base of the gynoecium. The section outlined in blue (B) is the same as Fig. 1K. Scale bars: 100 µm.

• Supplemental Figure S3 -

  Fig. S3. In situ localizations of WUS mRNA in lfy flowers. (A) View of a lfy-6 shoot apex hybridized with a WUS probe. Expression in a stage 3 flower is marked (arrowhead). (B-M) Serial sections of a close-up view of the indicated flower (dashed box in A). No WUS expression is visible at the base of the gynoecium. The section outlined in blue (E) is the same as Fig. 1L. Scale bars: 50 µm.

• Supplemental Figure S4 -

  Fig. S4. In situ localizations of WUS mRNA in lfy pan double mutants. (A) View of a lfy pan shoot apex. Arrowhead marks the approximate position of the shoot apical meristem, lost in this glancing section. (B-I) Serial sections of a close-up view of the indicated flower (dashed box in A). WUS expression is visible at the base of the gynoecium in panels D-F. The section outlined in blue (E) is the same as Fig. 1M. Scale bars: 50 µm.

• Supplemental Figure S5 -

  Fig. S5. pan mutants reveal determinacy defects under sensitized conditions. Views of the same silique (mature fruit) from either wild-type (L-er) (A-C) or pan-3 (D-F) plants grown at long days with mixed white and gro-lux light. (A,D) Top view of wild-type (A) and pan (D) siliques, showing two carpels in the wild type and three in pan. Individual carpel valves are outlined (dashed lines). (B,E) View of the base of the gynoecium with valves indicated (arrows). (C,F) Side view of the gynoecium with one carpel valve removed. The growth of ectopic floral structures (arrowhead) is observed in pan (F) but not in wild-type (C) siliques.

• Supplemental Figure S6 -

  Fig. S6. Interactions between pan mutants and mutants with fourth-whorl phenotypes. (A-F) Double mutant phenotypes of pan-3 with crabs claw-1 (crc-1). Flowers from wild type (L-er accession) (A); pan-3 mutants (B); crc-1 mutants (C), with some outer organs removed to reveal carpels that are incompletely fused at their distal tips (arrow); and pan-3 crc-1 double mutants (D), displaying an additive effect in whorls one to three (five sepals, five petals and five stamens) and a synergistic interaction in the fourth whorl that involves a complete loss of carpel fusion (arrowheads). (E) crc gynoecium from the flower shown in C with one carpel removed. No ectopic organs are observed within. (F) The same pan crc flower shown in D with all primary floral organs, including the two carpels, removed. All the visible internal organs are carpelloid in nature, with stigmatic papillae (arrows). (G-I) Double mutant phenotypes of pan-3 with superman-1 (sup-1). (G) A representative sup-1 flower bearing four sepals, four petals, ten stamens and a masculinized carpel in the center. (H) The sup-1 flower shown in (G), with outer organs removed to show the central structure with both staminoid characteristics such as pollen (arrowhead), as well as female characteristics such as ovules (arrow). Part of a stamen filament is also visible. (I) A representative pan-3 sup-1 flower displaying five sepals, five petals, eleven stamens and a fertile, morphologically normal carpel at the centre (red arrowhead).

• Supplemental Figure S7 -

  Fig. S7. The AP1(1.7-kb) promoter fragment drives expression throughout the flower. pAP1-driven ER-GFP signal in an inflorescence meristem (left). Longitudinal optical sections taken through flowers at stages 3 (right, top), 4 (right, middle) or 5 (right, bottom). Sepals (arrowheads) and the central dome of the floral meristem (arrow) are indicated in each panel.

• Supplemental Figure S8 -

  Fig. S8. An inducible pPAN::alcR−alcA::PAN-RD produces causes floral indeterminacy. (A,B) Gynoecia from pan-2 flowers carrying a pPAN::alc−alcA::PAN-RD construct, wherein PAN-RD is expressed under the PAN promoter in an ethanol-inducible manner. (A) Gynoecium from a control (uninduced) flower is indistinguishable from the pan mutant and from induced wild-type plants (not shown). (B) Gynoecium from a plant induced with ethanol vapours over 4 days. The two primary fourth-whorl carpels (arrows) are completely unfused, as in pAP1(1.7-kb)::PAN-RD flowers, and reiterations of carpel whorls are visible within (arrowhead).

• Supplemental Figure S9 -

  Fig. S9. Results of particle bombardment experiments. (A-D) Onion epidermal cells were bombarded with either DNA of a published pAGi-3′::GUS reporter construct alone (A,C) or with an equal quantity of p35S::PAN-VP16. Images are the duplicate experiments summarized in Fig. 3C. (A,C) Basal GUS activity is observed for the reporter construct. (B,D) GUS activity is observed in 4- to 5-fold more cells when the reporter is co-bombarded with a p35S::PAN-VP16 fusion.