Mutational Analyses of a Fork Head Associated Domain Protein, DAWDLE, in iArabidopsis thaliana/i

摘要

DAWDLE (DDL) gene encodes a protein that contains an N-terminal arginine-rich domain and a C-terminal Fork Head Associated (FHA) domain in Arabidopsis thaliana. DDL protein is believed to function in microRNA biogenesis by mediating the recruitment of pri-microRNA to DICER-LIKE 1 and also stabilizing the microRNA. The aim of this study was to conduct a structure-function analysis to identify the regions in DDL that are of functional significance. Targeted Induced Local Lesions in Genome screen was performed in the Columbia erecta-105 background of Arabidopsis resulting in the identification of eight point mutations spanning DDL. The mutants were characterized by phenotypic and molecular analyses based on the prior knowledge on ddl knockout mutants. Height of the plant, hypocotyl and root length, and fertility were measured for phenotypic characterization, and microRNA172 levels were measured to assess the mutation effect at the molecular level. Phenotypic and molecular analyses of the mutants revealed effects resulting in ddl phenotypes of varying degrees in different organs and each mutant displayed at least one phenotype studied. Reduction in fertility and increase in stem length were two phenotypes that most of the mutants consistently displayed. Identification and characterization of several key residues in the arginine rich region and FHA domain will serve as an important tool for elucidation of DDL signaling pathway.