N-terminal phosphorylation inhibits Arabidopsis katanin and affects vegetative and reproductive development in opposite ways

Katanin is an evolutionarily conserved microtubule-severing enzyme that is essential for cytoskeletal remodeling throughout the plant life cycle. However, the molecular mechanisms that tune katanin activity to meet distinct cellular requirements remain unclear. Here, we demonstrate that N-terminal phosphorylation of the Arabidopsis thaliana p60 katanin subunit (KTN1) serves as a key regulatory switch controlling microtubule severing during vegetative and reproductive development. Using in vitro biochemical assays, we show that combined phosphorylation of three conserved serine residues (S92, S147, S199) inhibits KTN1s microtubule-severing activity by reducing both microtubule-binding affinity and ATPase activity. Strikingly, phosphomimetic (DDD) and phosphonull (AAA) versions of KTN1 exhibit opposite developmental phenotypes. The constitutively active AAA mutant rescues defects in cortical microtubule organization and vegetative growth but leads to abnormal meiotic spindles, reduced pollen viability, and defective pollen tube growth, resulting in low male fertility. Conversely, the catalytically impaired DDD mutant fails to restore vegetative growth but supports normal male fertility. These findings reveal that phosphorylation differentially modulates KTN1 activity to balance the opposing requirements for high microtubule severing during interphase cell expansion versus limited severing during meiotic cell divisions, providing a sophisticated mechanism to coordinate cytoskeletal dynamics with plant developmental programs.