Plokhovska S. The protective role of nitric oxide from the low temperatures influence on the organization of actin filaments of Аrabidopsis thaliana

The role of cytoskeleton as one of nitric oxide (NO) downstream targets is gaining the increasing recognition because of its involvement in plenty of NO-controlled processes in plants throughout the entire life cycle - starting from seed germination to pollination as well as (a)biotic stress tolerance. It has been revealed that low temperature has an inhibitory effect on A. thaliana primary root growth and causes an anisotropic increase of epidermal cells diameter in elongation zone (+0.5°C) and a formation of large number of deformed (ectopic) root hairs in differentiation zone (+4°C). Cold treatment of A. thaliana roots resulted in distinct changes in the initial organization of actin filaments in the cells of all studied zones of the primary root. A thinner and sparser network of actin filaments in epidermal cells of the meristematic zone after 1 h from the beginning of exposure at +4°C and its partial depolymerization after 2 h was observed. In the elongation zone of A. thaliana actin filaments reoriented after 1 h exposure, and after 2 h became thinner and were visible as short bundles of F-actin. It was found that the increased sensitivity to cold (+4°C) was typical for actin filaments in the cells of differentiation zone and root hairs. Thin actin network and a partial depolymerization of microfilaments were observed after 1 h and after 2 h only brightly colored dot structures or short bundles of F-actin could be seen. A change in orientation of the actin filaments in epidermal cells of the root apex after cold treatment with +0.5°C took place after 1 h exposition and after 2 h of cold treatment, disordered network of microfilaments became sparser and its partial depolymerization was observed. After 1 h cold treatment of root thinner and sparser network of actin filaments in meristematic cells was noted and in some cells their complete depolymerization after 2 h was observed. Microfilaments in epidermal cells of the elongation zone formed network with more pronounced transverse orientation after 1 h treatment with +0.5°C and after 2 h partial or complete depolymerization of microfilaments in same cells was observed. Actin filaments network in cells of root hairs is gradually becoming rare only bright shining rods or point of actin were observed and after 2 h of cold treatment at +0.5°C a bright stains of F-actin and a partial depolymerization of microfilaments were observed which can lead to disruption in initiation of root hairs formation and growth. On next step of our research the low temperature and NO donor or scavenger combined effects on the actin filaments organization in primary root cells of A. thaliana were studied. We found that the growth and differentiation of main roots of A. thaliana were sensitive to exogenous NO content. SNP (NO donor) stimulated differentiation processes such as the formation of numerous germs of root hairs with active growth. After cPTIO (NO scavenger) treatment the cell size increase (swelling) in transition and elongation zones of primary roots, induction of primordial formation of root hairs were observed. Furthermore, actin filaments organization of epidermal cells in different zones of primary roots is modulated by NO content. Thus, the exogenous NO donor (SNP) favours to microfilaments network reorganization, while both cold and NO scavenger (cPTIO) increase its randomization and fragmentation. We have found that not only the sparseness of actin network and microfilaments polymerization/depolymerization in cells of different zones of the root apex occurs, but actin filaments orientation changes also after cold treatment and combined treatment with low temperature and exogenous NO. The obtained results testify to the existence of a functional relationship between changes in the intracellular NO content and the organization of actin filaments when exposed to cold on the plant cell. This allows us to conclude that microfilaments are important intermediaries in the realization of cold effect on the plant cell and NO is involved in the cell response to the low temperatures by signaling through these cytoskeletal structures.