Defense of the dissertation of Beisekova Moldir Kudiyarbekovna for the degree of Doctor of Philosophy (PhD) in the educational program «8D05107 - Biology»

L.N. Gumilyov Eurasian National University, a dissertation defense for the degree of Doctor of Philosophy (PhD) by Beisekova Moldir Kudiyarbekovna on the topic «Stress – inducible modulation of molybdohydroxylase activity in barley (Hordeum vulgare L.) under salinity» to the educational program «8D05107 – Biology».

The dissertation was carried out at the «General Biology and Genomics education department» of L.N. Gumilyov Eurasian National University.

The language of defense is kazakh

Official reviewers:

Rysbekova Аiman Bokenovna – Candidate of biological science, Professor at the Institute of Agriculture and Forestry of NСJSC «S.Seifullin Kazakh Agrotechnical Research University» (Astana, Kazakhstan).

Sapakhova Zagipa Beisenovna – Doctor of Philosophy (PhD), Associate Professor, leading researcher at the Laboratory of Breeding and Biotechnology, Institute of Plant Biology and Biotechnology (Almaty, Kazakhstan).

Temporary members of the Dissertation Committee:

Bisenbaev Amangeldy Kuanbaevich – Academician of the National Academy of Sciences of the Republic of Kazakhstan, Doctor of Biological Sciences, Professor of the Department of Molecular Biology and Genetics of Al-Farabi Kazakh National University (Almaty, Kazakhstan).

Kakimzhanova Almagul Apsalamovna – Doctor of biological science, Professor, Head of Plant biotechnology and selection laboratory, National Center of Biotechnology (Astana, Kazakhstan).

Вakhsh Allah – Doctor of Philosophy (PhD), Associate Professor University of the Punjab (Lahore, Pakistan).

Scientific advisors:

Akbassova Alua Zholdasbayevna – Doctor of Philosophy (PhD), Associate Professor at the Department of Biotechnology and Microbiology, L.N. Gumilyov Eurasian National University (Astana, Kazakhstan).

Edyta Zdunek-Zastocka – Doctor of Philosophy (PhD), Professor, Institute of Biochemistry and Microbiology, Warsaw University of Life Sciences (Warsaw, Poland).

The defense will take place on November 06, 2026, at 11:00 AM in the Dissertation Council for the training direction «8D051 – Biological and related sciences» in the educational program «8D05107 – Biology» of L.N. Gumilyov Eurasian National University. The defense meeting is planned to be held offline and online.

Link: https://us06web.zoom.us/j/83936592218?pwd=XNnvE4Px9o9sLj3ehz1haXP3qcp7Kb.1

Conference ID: 839 3659 2218

Access code: 984984

Address: Astana, Satpayev str., 2, building №1, audience №302.

Abstract (English): General characteristics of the study. The dissertation was devoted for investigating the role of molybdohydroxylases in plant adaptation to adverse environmental conditions. In addition, the study examines the role of molybdenum in regulating stress responses and enhancing the resistance of different cultivars of barley to salinity stress. Relevance of the research. The productivity of agricultural crops is reduced by adverse abiotic factors that induce oxidative stress. During oxidation stress accumulation of hydrogen peroxide (H2O2), and one of the molybdohydroxylases, aldehyde oxidase (AO), is involved in its production. Another enzyme, xanthine dehydrogenase (XDH), participates in the formation of uric acid, which has strong antioxidant properties and decreases the level of oxidative stress. The third enzyme studied in research, nitrate reductase (NR), catalyzes the reduction of nitrates and is involved in the formation of signaling agent NO, which serves as a signaling agent and is involved in the regulation of plant responses to abiotic stress. In addition, sulfite oxidase (SO), another molybdohydroxylase, also oxidizes sulfite to sulfate. Molybdenum is a vital micronutrient required for the functioning of molybdohydroxylases, plant growth, development, and productivity. Its deficiency leads to decreased molydohydroxylase activity and disruption of metabolic processes, especially under stress conditions. Therefore, optimizing molybdenum supply to plants can be important for increasing their resilience and productivity. Molybdate is currently used as a component of fertilizers in soils with insufficient levels of available molybdenum. However, excessive and improper use of molybdenum-containing fertilizers can have negative impacts on the environment and living organisms, highlighting research into optimal conditions for its use. According to current findings, molybdenum can increase plant tolerance to salinity. To elucidate the mechanisms of this effect, we hypothesized that molybdenum may enhance plant stress resistance by activating AO-dependent ABA biosynthesis, in which, in turn, promotes proline accumulation. Proline is an important osmoprotectant and antioxidant involved in protecting cells from stress. To test this hypothesis, tungsten (W) was used as a functional antagonist of molybdenum. Due to its ability to replace for molybdenum in the molybdenum cofactor (Moco), tungsten can reduce the activity of molybdenum-containing enzymes, particularly aldehyde oxidase (AO), thereby enabling an assessment of the contribution of the AO-dependent pathway to ABA regulation and proline accumulation. However, the existence of ABA-independent mechanisms for regulating its synthesis indicates the need for further study of the relationship between AO activity, ABA levels, and proline accumulation. Thus, exploring the role of molybdohydroxylases in resistance of plant to salinity is of both fundamental and practical importance. The knowledge gained from research may contribute to the development of more effective and environmentally sustainable approaches to increasing the resilience and productivity of agricultural crops in saline soils and insufficient molybdenum availability. Object of the study: «Baysheshek» and «Astana 2000» barley cultivars. Subject of the study: Modulation of the activity of molybdoenzymes (AO, XDH, NR, SO), in studied barley plants (Hordeum vulgare L.) under abiotic stress conditions as an indicator of the degree of oxidative stress development and as a marker of plant tolerance to salinity. Purpose and objectives of the study. The aim of this study was to explore the role of oxidative stress markers and stimulation of molybdohydroxylase enzymes in the regulation of plant stress resistance under salinity conditions. To achieve this aim, the following objectives were set: 1. To study the effect of molybdenum and tungsten on the accumulation of oxidative stress markers in plants under salinity conditions. 2. To study the activity of antioxidant enzymes (superoxide dismutase, catalase) in barley under the combined effects of metals and salinity. 3. To investigate the role of molybdenum and tungsten in modulating the activity of molybdohydroxylases (AO, XDH, NR, and SO) under salinity conditions. 4. To determine the effect of molybdenum and tungsten on the content of the phytohormone abscisic acid (ABA) in plants under salt stress. 5. To investigate the effect of molybdenum and tungsten on the accumulation of proline and proline metabolism under salinity conditions. Research methods. Native electrophoresis, spectrophotometric methods, real-time PCR, digital data processing using the Image J program, and analysis of variance (ANOVA) were used in the course of the study. Scientific novelty. For the first time, it was established at the molecular physiological level that the accumulation of proline in barley (Hordeum vulgare L.) under salt stress is predominantly regulated by an ABA-independent pathway. Molybdenum has been shown to activate aldehyde oxidase and increase ABA levels, but this is not accompanied by a significant increase in proline content. Expression analysis of key proline metabolism genes (P5CS and ProDH) revealed that proline accumulation is primarily associated with increased synthesis of proline and can occur independently of the ABA-dependent signaling pathway. A relationship was established between xanthine dehydrogenase (XDH) activity and catalase (CAT) activity under salt stress. Increased activity of the second catalase isoforms in roots was detected in response to molybdenum and tungsten treatment, indicating its possible role in the plant antioxidant defense system. It was also shown that superoxide dismutase (SOD) isoforms have similar activity toward molybdenum and tungsten, indicating that ABA is not an inducer of SOD isoforms. These results allow us to clarify the role of XDH in regulating the antioxidant defense system of the plant under salt stress. The functional interactions between AO, SOD, and CAT in response to abiotic stress factors remain poorly understood. The role of AO in regulating proline accumulation and shaping plant defense responses to salinity also remains unclear. Theoretical and practical significance. The theoretical significance of this study lies in its expanded understanding of the molecular and physiological mechanisms of cereal crop adaptation to abiotic stress. The obtained results demonstrate that proline accumulation under salt stress can be regulated not only by ABA-dependent pathways but also through ABA-independent mechanisms associated with changes in the cellular redox state. These findings expand our understanding of the role of AO, XDH, and their catalytic reaction products in maintaining redox homeostasis under salt stress. Analysis of the expression of key proline metabolism genes (P5CS and ProDH) linked changes in proline content to the regulation of proline synthesis and degradation. Taken together, these results complement existing understanding of the mechanisms of plant stress tolerance and provide a better understanding of the specific characteristics of barley varietal tolerance to salinity. The practical significance of the study lies in the potential use of molybdenum to enhance the tolerance and maintain productivity or cereal crops under salt stress. The findings obtained on the role of molybdenum, molybdoenzymes, and ABA can be used to develop markers for early assessment of salt tolerance and the selection of resistant barley cultivars. The results of the study can also be applied to the development of agricultural practices to enhance the tolerance of grain crops to salinity. Main results submitted for defense. - Salinity stress in barley plants leads to increased accumulation of ROS and MDA in tissues. In the absence of salinity, molybdenum does not alter their levels, whereas tungsten increases ROS and MDA levels. Under salinity stress, molybdenum increases the accumulation of anthocyanins and allontoin, whereas tungsten decreases their content. - Neither molybdenum nor tungsten do not affect SOD activity under non-saline conditions, while their combined affect with NaCl leads to a decrease in SOD activity in barley shoots and roots. In the absence of salinity, molybdenum treatment leads to increased catalase activity, while tungsten causes only slightly increase in catalase activity. - Salinity stress of barley plants leads to increased activity of molybdohydroxylases. In the absence of salinity, molybdenum also increases molybdohydroxylase activity, while tungsten reduces significantly their activity. The obtained results confirm the important role of molybdohydroxylases in barley plant adaptation to salt stress. - Increased molybdoenzyme activity improves seed germination and plant morphological health during the early stages of growth and development. - Proline accumulation in barley under salt stress is independent of ABA levels, but is associated with the degree of oxidative stress, which is regulated by molybdenum and tungsten. Connection of the Dissertation with Research Programs The dissertation research was carried out within the framework of grant-funded projects under the program of the Ministry of Science and Higher Education of the Republic of Kazakhstan project AP05135013 «The involvement of ROS producing Mo-enzymes in root development and stress tolerance of plants» and the «Zhas Galym» Young Researchers Competition project AP22685648 «Development of innovative technology for express purification of soil from heavy metals». The research was conducted at the R.T. Omarov Laboratory of Plant Biotechnology, Research Institute of Cell Biology and Biotechnology, L.N. Gumilyov Eurasian National University (Astana, Kazakhstan), and at the Institute of Biochemistry and Microbiology, Warsaw University of Life Sciences (SGGW) (Warsaw, Poland). Publications related to the research topic. The main results of the study are presented in 7 published papers, including: 2 articles in international journals indexed in the Scopus and Web of Science databases: Horticulturae, (Q1, impact factor 3); and International Journal of Molecular Sciences (Q1, impact factor 4.9); 4 articles in the journals of the Republic of Kazakhstan recommended by the Committee for Quality Assurance in Science and Higher Education of the Ministry of Science and Higher Education of the Republic of Kazakhstan: ENU Bulletin (3) and Eurasian Journal of Applied Biotechnology (1); and 1 publication in the proceedings of international scientific and practical conferences. Structure and scope of the dissertation. The dissertation consists of an introduction, four chapters, a conclusion, and 242 references. The main text comprises 104 pages. The dissertation includes 35 figures and 4 table.

Dissertation

List of Scientific Works

Consultant's Review

Foreign Consultant's Review

Conclusion of the Research Ethics Committee