MLK4-mediated Phosphorylationof Histone H3T3 Promotes Flowering by Transcriptional silencing of FLC/MAFin Arabidopsis thaliana.

Casein kinase I (CK1), a ubiquitous Ser/Thr protein kinase in eukaryotes, plays a critical role in higher plant flowering. Arabidopsis CK1 family member MUT9-LIKE KINASEs, such as MLK1 and MLK3, have been shown to phosphorylate histone H3 at threonine 3 (H3T3), an evolutionarily conserved residue, and the modification is associated with transcriptional repression of euchromatic and heterochromatic loci. This study demonstratesthat mlk4-3, aT-DNA insertion mutant of MLK4, flowered late and overexpression of MLK4 caused early flowering. The nuclear protein MLK4 phosphorylated histone H3 at Thr3in vitro and in vivo, and thecatalytic activity required the conserved lysine residue K175. The mutation K175Rof MLK4failed to restore the H3T3ph level or complement the phenotypic defects of mlk4-3. The FLC/MAF-clade genes includingFLC, MAF4 and MAF5 were significantly up-regulated in mlk4-3. The double mutant mlk4-3 flc-3 flowered earlier than mlk4-3, suggesting functional FLC is crucial for flowering repression in mlk4-3. ChIP assay showed that MLK4 bound to FLC/MAF chromatin and the H3T3ph occupancy at the promoter of FLC/MAF was negatively associated with its transcriptional level. Consistently, H3T3ph accumulated at FLC/MAF in 35S::MLK4/mlk4-3, but diminished in 35S::MLK4(K175R)/mlk4-3plants. Moreover, the amount of RNA Pol II deposited at FLC/MAFwas clearly enrichedin mlk4-3 relative to wild type. Therefore, MLK4-dependent phosphorylation of H3T3 contributes to accelerating flowering by repressing the transcription of flowering negative regulatorFLC/MAF. This study sheds light on the delicate control of flowering by the plant-specific CK1, MLK4,via posttranslational modification of histone H3.