DEV001891 Supplementary Material

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• Supplemental Table S1 -

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• Supplemental Figure S1 -

  Fig. S1. Alignment of deduced amino acids of AtSWC6, Yeast SWC6/VSP71 and mammalian ZnF/HIT1. (A) Schematic of AtSWC6 protein with a HIT-type zinc-finger domain. (B) Multiple alignment of putative atswc6 homologs. SWC6 and ZnF/HIT1 (ZNHIT1) are known components of the yeast SWR1 and mammalian SRCAP (SWI2/SNF2-related CBP activator protein) complexes, respectively. Darkly shaded and lightly shaded residues represent identical and similar amino acids, respectively; dots indicate the conserved seven cysteines and one histidine typical of the HIT-type zinc-finger domain.

• Supplemental Figure S2 -

  Fig. S2. Western blot analyses for detection of myc:AtSWC6 and AtSWC6:GFP proteins. Protein was extracted from 10-day-old plants of Col, four independent 35S-myc:AtSWC6 atswc6 transgenic lines (A), and two independent 35S-AtSWC6:GFP atswc6 lines (B). Protein (50 μg per sample) was used for immunoblotting with anti-myc antibody or anti-GFP polyclonal antibody. Ponceau staining of the blots provides a loading control.

• Supplemental Figure S3 -

  Fig. S3. AtSWC6 expression in other flowering time mutants. RNA was extracted from 10-day-old plants grown in long-day conditions and analyzed by RT-PCR. (A) Expression of AtSWC6 in FRI, Col and the mutants involved in positive regulation of FLC (suf4, vip4, pie1, suf3, abh1) and the autonomous pathway (ld, fld, fca, fy, fpa). (B) Expression of AtSWC6 in flc and mutants in the photoperiod pathway (gi, co) and flowering pathway integrators (ft, soc1). TUB was used as an internal control.

• Supplemental Figure S4 -

  Fig. S4. Nuclear localization of AtSWC6:GFP fusion protein in the root of 35S-AtSWC6:GFP atswc6 transgenic plant. (A,B,D,E) Section images and (C,F) projection images of GFP florescence in roots of 7-day-old seedlings grown in long-day conditions detected by confocal microscopy. Scale bars: 10 μm

• Supplemental Figure S5 -

  Fig. S5. Comparison of transcript level and spatial expression pattern of HTA8, HTA9 and HTA11. (A) RT-PCR analysis for comparison of transcript level of HTA8, HTA9 and HTA11. HTA9 is more highly expressed than HTA11 and HTA8. (B) HTA8, HTA9 and HTA11 expression in different tissues. For RT-PCR analysis, RNA was extracted from 20-day-old Col grown in long-day conditions for whole plants (Wh), root (Rt), shoot apex (SA) and rosette leaves (RL). RNA for old leaves (OL), stem (St) and inflorescence with flowers (Inf) was extracted from 40-day-old plants.

• Supplemental Figure S6 -

  Fig. S6. Protein blots showing interaction between AtSWC6 and SUF3 in tobacco. The same interaction analysis as Fig. 6F but with antibodies for IP and western blot reversed. IP indicates immunoprecipitation of protein extracts from tobacco leaves transiently co-expressing epitope-tagged proteins, as described above the protein blot, with anti-myc (α-myc) or anti-Flag (α-Flag) antibodies. The immunoprecipitates and protein extracts were separated by 9% SDS-PAGE, transferred to polyvinylidene difluoride membranes and probed with anti-GFP (left four lanes) or anti-Flag (right two lanes) antibodies. About 10% of the total sample used in each IP was loaded as input control.