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. 2020 Jul 1;16(7):e1008883. doi: 10.1371/journal.pgen.1008883

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© 2020 Li et al

This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Fig 7 — (A) His pull-down assays of PRE2/3 and UPB1. His-SUMO-UPB1, GST-PRE2, GST-PRE3, and GST were purified from E. coli. Purified proteins were used for the pull-down assay. His-SUMO-UPB1 protein was detected with an anti-His antibody, and the GST-PRE2, GST-PRE3, and GST proteins were detected with anti-GST antibodies. (B) Co-IP assays of UPB1 and PRE2/3. Protein extracts from 4-week-old 35S::PRE2-GFP/35S::HA-Flag, 35S::PRE2-GFP/35S::UPB1-HA-Flag, 35S::PRE3-GFP/35S::HA-Flag, and 35S::PRE3-GFP/35S::UPB1-HA-Flag seedlings, An anti-HA antibody was used for immunoprecipitation, and immunoblot analysis was performed with anti-HA and anti-GFP antibodies. (C) Phenotypes of 5-day-old seedlings of Col-0, upb1-1, pre-amiR/pre3, pre-amiR/pre3/upb1-1, 35S::UPB1-HA-Flag, 35S::PRE2-GFP/35S::UPB1-HA-Flag, and 35S::PRE3-GFP/35S::UPB1-HA-Flag. Bar = 0.5 cm. (D) Root meristem of Col-0, upb1-1, pre-amiR/pre3, pre-amiR/pre3/upb1-1, 35S::UPB1-HA-Flag, 35S::PRE2-GFP/35S::UPB1-HA-Flag, and 35S::PRE3-GFP/35S::UPB1-HA-Flag in 5-day-old seedlings. White arrowheads (below) mark the position of the quiescent center (QC), and white arrowheads (above) mark the end of the meristem where cells start to elongate. Bar = 50 μm. The primary root length (E), meristem size (F), and meristem cell number (G) in the seedlings shown in (C). Date means ± SD (n≥20). Double asterisk represent highly significant differences (**, P<0.01; Student’s t test). (H) The UPB1 protein levels were detected in 35S::UPB1-HA-Flag, 35S::PRE2-GFP/35S::UPB1-HA-Flag, and 35S::PRE3-GFP/35S::UPB1-HA-Flag transgenic lines. UPB1 was detected with an anti-HA antibody. Actin was used as a control.