RNAi screening identifies the armadillo repeat-containing kinesins responsible for microtubule-dependent nuclear positioning in Physcomitrella patens

Abstract

Proper positioning of the nucleus is critical for the functioning of various cells. Actin and myosin have been shown to be crucial for the localisation of the nucleus in plant cells, whereas microtubule (MT)-based mechanisms are commonly utilised in animal and fungal cells. In this study, we combined live cell microscopy with RNA interference (RNAi) screening or drug treatment and showed that MTs and a plant-specific motor protein, armadillo repeat-containing kinesin (kinesin-ARK), are required for nuclear positioning in the moss Physcomitrella patens. In tip-growing protonemal apical cells, the nucleus was translocated to the centre of the cell after cell division in an MT-dependent manner. When kinesin-ARKs were knocked down using RNAi, the initial movement of the nucleus towards the centre took place normally; however, before reaching the centre, the nucleus was moved back to the basal edge of the cell. In intact (control) cells, MT bundles that are associated with kinesin-ARKs were frequently observed around the moving nucleus. In contrast, such MT bundles were not identified after kinesin-ARK downregulation. An in vitro MT-gliding assay showed that kinesin-ARK is a plus-end-directed motor protein. These results indicate that MTs and the MT-based motor drive nuclear migration in the moss cells, thus showing a conservation of the mechanism underlying nuclear localisation among plant, animal, and fungal cells.