Functional Characterization Of KATANIN Subunit OsKTN80a In Microtubule Dynamics In Rice

As a kind of cytoskeletons, microtubules are essential for cell morphogenesis, the intracellular transportation, cell division, composition of some novel cellular apparatus such as flagellum and cilia and the plant cell wall biosynthesis and so on. Microtubules are highly dynamic and one of the reasons is that it was regulated by microtubule associated proteins (MAPs). Katanin, a kind of MAP involved in microtubule severing, is composed of two subunits, one is P60which has ATPase and microtubule severing activity and the other is P80which has no microtubule severing activity but plays roles in targeting P60to the centrosome and regulating P60’s microtubule severing activity. P60itself are involved in many significant life activities while P80also functions essentially in C. elegans meiosis, human neuronal development, the male gamete genesis in mouse and in cytokinesis and flagella biogenesis in lower order species. KATANIN P60is necessary for the anisotropic cell expansion and cell division in root tip and also acts as the effector in cortical microtubule orientation in auxin signaling pathway in Arabidopsis cell. When it comes to the other KATANIN subunit P80in higher plants, its function remain largely unknown.In this study, we used rice KATANIN P60(KTN60) as the bait to screen the bacteria two hybrid cDNA library of rice immature panicle and obtained a KTN80b. Through BLAST the protein database of Oryza sativa using this protein, we found the other two homologous proteins (KTN80a and KTN80c) in rice. There are high sequence similarity between the3homologous proteins mentioned above and the KATANIN P80in human and mouse. Later the3homologs are demonstrated as katanin p80orthologs in rice after yeast two hybrid assay, the domain analysis and the phylogenic analysis. The overexpression of OsKTN80a in rice (KTN80a-OX) caused the delayed primary root growth of rice seedling and two reasons results in it:the first is that cell elongation dysfunction in root tip in KTN80a-OX which is thought as the disordered cortical microtubule arrangement in elongation zone after Ktn80a overexpression; the second is that cell division are stalled at cytokinesis which are mainly caused by the incomplete separation of genetic materials into the daughter cells and the abnormal phragmoplast morphology and function. The above two reasons could be explained that the KTN60microtubule severing activity are repressed by KTN80a and the event caused the broken microtubule dynamics. There are3evidence supporting the idea:1) KTN80a interacts with KTN60;2) The phenotype of the shorter cell in root tip and the disordered cortical microtubule array in elongation zone when KTN80a is overexpressed could perfectly mimic the case in KTN60mutant (fra2);3) Compared with the wild type, the root tip growth and cortical microtubules in elongation zone are more resistant to treatment of oryzalin, a kind of microtubule-special destabilizing drug, which implies that KTN80a stabilizes microtubules.We discuss the molecular mechanism of the role of KTN80a plays in cell expansion and in cell division. From the above mechanism of KTN80a function in, we can get the conclusion that controlling the KATANIN activity is essential for cell expansion and cell division in rice root tip.