Altered Root Growth, Auxin Metabolism and Distribution in Arabidopsis thaliana Exposed to Salt and Osmotic Stress

Salt and osmotic stress are the main abiotic stress factors affecting plant root growth and architecture. We investigated the effect of salt (100 mM NaCl) and osmotic (200 mM mannitol) stress on the auxin metabolome by UHPLC-MS/MS, auxin distribution by confocal microscopy, and transcript levels of selected genes by qRT-PCR in Arabidopsis thaliana ecotype Columbia-0 (Col-0) and DR5rev::GFP (DR5) line. During long-term stress (13 days), a stability of the auxin metabolome and a tendency to increase indole-3-acetic acid (IAA) were observed, especially during salt stress. Short-term stress (3 h) caused significant changes in the auxin metabolome, especially NaCl treatment resulted in a significant reduction of IAA. The data derived from auxin profiling were consistent with gene expressions showing the most striking changes in the transcripts of YUC, GH3, and UGT transcripts, suggesting disruption of auxin biosynthesis, but especially in the processes of amide and ester conjugation. These data were consistent with the auxin distribution observed in the DR5 line. Moreover, NaCl treatment caused a redistribution of auxin signals from the quiescent center and the inner layers of the root cap to the epidermal and cortical cells of the root elongation zone. The distribution of PIN proteins was also disrupted by salt stress; in particular, PIN2 was suppressed, even after 5 min of treatment. Based on our results, the DR5 line was more sensitive to the applied stresses than Col-0, although both lines showed similar trends in root morphology, as well as transcriptome and metabolome parameters under stress conditions.

Mutual Promotion of LAP2 and CAT2 Synergistically Regulates Plant Salt and Osmotic Stress Tolerance

Almost all abiotic stresses induce reactive oxygen species (ROS) overaccumulation, causing oxidative damages to plant cells. Catalase (CAT) plays a vital role in plant oxidative stress tolerance by scavenging stress-induced excess H2O2; thus, the identification of factors regulating catalase function will shed light on the underlying regulatory mechanisms. Here, we identified leucine aminopeptidase 2 (LAP2) as a novel CAT2-interacting protein and showed a mutual promotion effect of the two proteins in plant stress responses. LAP2 has a physical interaction with CAT2 in plant cells. The loss-of-function mutant of LAP2, lap2-3, is hypersensitive to salt or osmotic stress with increased ROS accumulation and malondialdehyde content and decreased catalase activity. The lap2-3 mutant has less CAT2 protein levels as CAT2 protein stability is impaired in the mutant. Scavenging excess ROS by glutathione or overexpressing CAT2 in the lap2-3 mutant recovers its hypersensitive phenotype to salt or osmotic stress. Further study showed that CAT2 promotes LAP2 hydrolysis activity with leucine-4-methylcoumaryl-7-amides as a substrate in vivo and in vitro, and thus, similar to the lap2-3 mutant, the cat2-1 mutant also has lower γ-aminobutyric acid content than the wild type. Together, our study reveals mutual promotion effects of CAT2 and LAP2 in conferring plant salt and osmotic stress tolerance.

Conserved Aquaporins in Gossypium and Silencing of Ghpip2;7 and Ghtip2;1 Decreased Salt and Osmotic Stress Tolerances in Upland Cotton

Aquaporins (aquaporin), membrane channel proteins, facilitate the transport of water and small molecules across intrinsic membranes and play a critical role in abiotic stresses. Here, 111, 54, and 56 candidate AQP genes were identified in Gossypium hirsutum (AD1), G. arboreum (A2), and G. raimondii (D5), and classified into five subfamilies including PIP, TIP, NIP, SIP, and XIP, respectively. Some GhPIPs and GhTIPs exhibited high expression levels in drought and salt stress verified by transcriptome analysis and Quantitative Real-time PCR (qRT-PCR). The chlorophyll content, SOD activity, and POD activity were decreased in GhPIP2;7 gene-silenced plants versus the control (using blank vector) under 400 mM NaCl treatment, which indicated a negative role of GhPIP2;7 in cotton salt tolerance. Comparing with mock plants, GhTIP2;1 silenced cotton plant was more sensitive to osmotic stress. Overexpressed GhTIP2;1 plants exhibited less accumulation of H2O2 and MDA with higher proline contents detected under osmotic stress. Taken together these results, provide in-depth knowledge into plant response to abiotic stress and gene resource.

Phenological evaluation of the established somaclonal line of camelina sativa resistant to salt and osmotic stress

The biological features of сamelina sativa make it possible to grow it in different soil and climatic conditions with high economic efficiency, to obtain environmentally friendly products and make full use of the natural potential of the region. Camelina oil is used for the production of varnishes, paints, soaps, plastics, biodiesel. It is also used for dietary and medical nutrition. The introduction of high-yielding adaptive varieties is the main condition for increasing the production of camelina sativa. Biotechnological methods are used to increase the efficiency of the selection process. Plant forms obtained by biotechnological methods must be evaluated according to a set of economically valuable traits. The length of the growing season is one of the main breeding characteristics of crop varieties. The aim of the work was to analyze the duration of the vegetation period and phenological phases of development of the created somaclonal lines of, resistant to salt and osmotic stress. Phenological evaluation of somaclonal lines of сamelina sativa obtained from explants of Stepovyi 1, Peremoha, Klondike and Yevro 12 varieties was performed during 2018–2020. Created by cell selection, stress-resistant plant material (sodium chloride, mannitol) plant material after microclonal propagation, rooting and adaptation was grown in the research areas of the Department of Genetics, Plant Breeding and Biotechnology, Uman National University of Horticulture. The analysis of the duration of the vegetation period and phenological phases of development of somaclonal lines of сamelina sativa resistant to salinity and osmotic stress was performed. It is established that the lack of moisture and elevated air temperatures contribute to the acceleration of the phases of ontogenesis and the reduction of the total growing season. The duration of «sowing-germination» period plants over the years of research, depending on the genotype, varied from 9 to 13 days, the formation of the rosette — from 8 to 13 days, the period of stalking and budding — from 18 to 20 days, the duration of flowering — from 8 to 27 days, the phase of the green pod — from 13 to 15 days, the period from the beginning of pod browning to the onset of full ripeness of seeds — from 26 to 31 days. The vegetation period of the created samples of сamelina sativa in 2018 averaged 80 days, in 2019 — 89 days and in 2020 — 88 days. According to the Methodology of examination of сamelina sativa for difference, homogeneity, stability the obtained data make it possible to rank the created somaclonal plant lines by the duration of the growing season on medium- ripening – C-234-8, C-326-9, C-402-6, С-419-6, С-586-7, П-46-2, П-46-5, П-202-6, П-202-7, П-248-8, П-485-4, П-618-6, П-646-3, П-658-8, Є-405-5, Є-405-8, K-478-2, K-480-2, K-480-4 and late-ripening – C-87-4, C-87-7, C-121-2, C-121-11, C-384-4.

MsCIPK, a CBL-interacting Protein Kinase in Medicago sativa L., Confers Salt and Osmotic Stress Tolerance in Transgenic Tobacco

Alfalfa is an important perennial forage but suffers from salt and osmotic stresses worldwide. Calcineurin B-like proteins (CBLs) and CBL-interacting protein kinases (CIPKs) are reported to play important roles in response to diverse plant stresses, but are largely unvalidated in alfalfa. In this study, we cloned a MsCIPK gene, which contained 1530 bp, coding 509 amino acids, with typical CIPK functional domains. The expression pattern of MsCIPK was measured using qRT-PCR under salt, drought, heat, cold and ABA stresses. Under NaCl, heat and ABA treatment, the expression pattern of MsCIPK was generally similar, with a first steady decrease and then a gradual increase pattern. The highest expression of MsCIPK was all observed at the start point of all treatments, except in cold treatment. Using transgenic tobaccos of MsCIPK, we further measured the content of malondialdehyde (MDA), superoxide dismutase (SOD), soluble protein (SOP), and proline (Pro) under 21 days’ salt and 24 hours’ cold treatment. Under both salt and cold conditions, the content of MDA, SOP and Pro had a similar overall increase pattern with the time of treatment. These results indicated that the MsCIPK played an important role in improving alfalfa’s salt and osmotic tolerance.

Dual role of MdSND1 in the biosynthesis of lignin and in signal transduction in response to salt and osmotic stress in apple

Clarifying the stress signal transduction pathway would be helpful for understanding the abiotic stress resistance mechanism in apple (Malus × domestica Borkh.) and could assist in the development of new varieties with high stress tolerance by genetic engineering. The key NAC transcription factor SND1, which is involved in the lignin biosynthesis process in apple, was functionally analyzed. The results of the stress treatments indicated that MdSND1 could be induced by salt, mannitol and ABA. Compared with wild-type GL-3 plants, MdSND1-overexpressing apple plants with greater antioxidant capacity and lignin were more resistant to salt and simulated osmotic stress, while RNAi plants were more vulnerable. Additionally, molecular experiments confirmed that MdSND1 could regulate the biosynthesis of lignin by activating the transcription of MdMYB46/83. Moreover, genes known to be involved in the stress signal transduction pathway (MdAREB1A, MdAREB1B, MdDREB2A, MdRD29A, and MdRD22) were screened for their close correlations with the expression of MdSND1 and the response to salt and osmotic stress. Multiple verification tests further demonstrated that MdSND1 could directly bind to these gene promoters and activate their transcription. The above results revealed that MdSND1 is directly involved in the regulation of lignin biosynthesis and the signal transduction pathway involved in the response to both salt and osmotic stress in apple.