Volume 7, Issue 12, Pages 1788-1792 (December 2014) A Novel Single-Nucleotide Mutation in a CLAVATA3 Gene Homolog Controls a Multilocular Silique Trait in Brassica rapa L.  Chuchuan Fan, Yudi Wu, Qingyong Yang, Yang Yang, Qingwei Meng, Keqiao Zhang, Jinguo Li, Jinfang Wang, Yongming Zhou  Molecular Plant  Volume 7, Issue 12, Pages 1788-1792 (December 2014) DOI: 10.1093/mp/ssu090 Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 1 A Pro-to-Leu Substitution in BrCLV3 Gene Controls a Multilocular Silique Trait in Brassica rapa L. (A) Siliques of the ml4 mutant consisting of two to four carpels with an extra gynoecium inside the silique at a high frequency (arrow). (B) SAMs of wild-type (wt) and ml4 mutant at indicated stages. From left to right: the embryos observed by differential interference contrast microscopy (bar = 10 μm); longitudinal section of 10-day-old SAMs and the primary inflorescence meristems (bar = 10 μm); scanning electron micrographs of the primary inflorescence meristems (bars = 100 μm). (C) Cross-sections of gynoecia of wt and ml4 mutant at indicated stages. Scale bar = 10 μm; lc, locule; O, ovule; M, medial region; L, lateral region. (D) Fine mapping of the ML4 gene on Scaffold000094 of B. rapa. The numbers between the molecular markers represent recombination events. The solid circles indicate the locations of the predicted genes in B. rapa. (E) The BrCLV3 gene structure and natural variations between the alleles from wt and ml4 mutant. The positions of the coding regions (black boxes), translation start codon, and translation stop codon are indicated. The ml4 mutant has three single-nucleotide substitutions and one ATAT deletion compared with wild-type in the Brclv3 gene region, which includes the entire coding sequence, 2.3-kb promoter region, and 2.5-kb 3'-flanking sequence. (F) Alignment of the putative protein sequences of CLV3 from B. rapa (ml4 mutant, wt, and Chiifu-401) and Arabidopsis. The putative signal sequence cleavage sites is indicated by arrow, the conserved domain (CLE motif) at its C-terminus is underlined, and the amino acid change in ml4 mutant is highlighted by the black box. (G) Carpel numbers of seven independent 35S::BrCLV3 transgenic lines. The data and error bars represent the mean ± SD (n ≥ 10 plants for each line). Uppercase letters indicate a significant difference at the 0.01 probability level. (H) Area of the SAM of Arabidopsis after different peptide treatments for 9 d (left) and B. rapa after different peptide treatments for 12 d (right). The data and error bars represent the mean ± SD (n ≥ 15 plants for each line). ** Significant at p < 0.01 compared with the corresponding SAM size grown on 1/2 MS media based on Student's t-test. (I) Expression of Arabidopsis homologous genes involved in the CLV3 signaling pathway in the inflorescence meristem of wt and ml4 mutant. The expression levels were determined by real-time PCR and normalized to 18S rRNA. The values are presented as the mean ± SD (n = 3 biological replicates). (J) A model for CLV signaling in B. rapa. The CLV/WUS feedback loop restricts the stem cell fate in the SAM and permits the formation of only two carpels in wild-type B. rapa (left); the Pro-to-Leu substitution in CLV3 leads to a non-functional, weak receptor-binding peptide (CLV3Leu9), preventing downstream signal transduction (right). Consequently, the expression of WUS without inhibition markedly promotes the expression of Brclv3 and its receptors, resulting in an enlarged SAM and extra locules in the ml4 mutant. Molecular Plant 2014 7, 1788-1792DOI: (10.1093/mp/ssu090) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions