Figure 2: RSS1 encodes an uncharacterized protein containing a D-box.
From: RSS1 regulates the cell cycle and maintains meristematic activity under stress conditions in rice
(a) (top) The schematic structure of RSS1. Rectangles indicate the exons. The first ATG, the stop codon, and the insertion sites of the Tos17 retrotransposon (arrows) in the three mutant alleles are shown. (bottom) The RSS1 protein including its conserved motifs. The three regions containing amino acid (a.a.) sequences that are conserved among the RSS1 homologues, the DEN and double D-Box, the WAGE motif, and the C-terminal acidic amino-acid-rich region are shown. (b) Assay for the interaction of RSS1 with the subunits of APC/C by the yeast two-hybrid system. (top) The GAL4 DNA-binding domain fused to the truncated RSS1 (a.a. 1–207) and GAL4 activation domain fused to OsCdh1;1 (Os01g0972900), OsCdh1;2 (Os03g0123300) or OsCdc20;1 (Os04g0599800) were co-expressed in the yeast cells carrying the reporter genes, ADE2 and HIS3. The yeast cells were grown on the appropriate medium (which did not contain the indicated amino acids). (bottom) The two-hybrid assay using truncated RSS1 (a.a. 1–104) with or without the D-boxes as the bait. Empty indicates the vector control. (c, d) The cell cycle-dependent accumulation of GFP (c) and RSS1-GFP (d). Rice Oc cells carrying the respective transgenes were synchronized and used for ploidy analysis (top) and immunoblot analysis using anti-GFP (middle) at the indicated time points after release from G1 arrest by starvation. (top) The rates of 2C cells (blue) and 4C cells (magenta) are plotted. The 59-kDa band corresponds to the full-length RSS1-GFP protein. (bottom) Protein staining for the loading control of the immunoblot. (e) A simplified phylogeny of plants and the schematic structure of RSS1-related proteins representative of the respective lineages are illustrated. The percent amino-acid identity among sequences homologous to regions I and II is indicated. The C-terminal regions containing acidic amino acids are not necessarily homologous to region III. The dashed line indicates truncated sequence information. GenBank accession and code numbers are as follows: Arabidopsis AT3G14910 (NM_112353), Oryza sativa Os02g0606700 (RSS1) (NC_008395), Aristolochia fimbriata (FD758252), Nuphar advena (PUT-165a-Nuphar_advena-6361, Plant GDB (http://www.plantgdb.org/), Amborella trichopoda (CK757545), Picea sitchensis (ABK26784), Adiantum capillus-veneris (DK961685), and Syntrichia ruralis (CN201203).
