بررسی عناصر تنظیم کننده ژن‌های NAD(P)H oxidase در گندم

نوع مقاله : مقاله پژوهشی

نویسندگان

1 بخش بیوتکنولوژی گیاهی، دانشکده کشاورزی، دانشگاه شیراز، شیراز، ایران

2 بخش تحقیقات زراعی و باغی، مرکز تحقیقات و آموزش کشاورزی و منابع طبیعی استان کرمانشاه، سازمان تحقیقات، آموزش و ترویج کشاورزی، کرمانشاه،

چکیده

مقدمه: تغییرات آب و هوایی و افزایش شدت و مدت تنش‌های محیطی بر عملکرد و تولید جهانی اکثر گیاهان زراعی از جمله گندم تاثیر گذاشته است. بر این اساس بررسی اثر تنش‌های محیطی بر تغییرات سلولی در سطح ملکولی جهت تشخیص مکانیسم‌های تنظیمی و استفاده از آنها برای افزایش تحمل و مقاومت در گیاهان اهمیت ویژه‌ای پیدا کرده است. گیاهان برای ﻣﻘﺎﺑﻠﻪ ﺑﺎ ﺗﻨﺶﻫﺎی محیطی راهﮐﺎرﻫﺎی ﻣﺨﺘﻠﻔﯽ از ﺟﻤﻠﻪ ﺗﻮﻟﯿﺪ ﺳﺮﯾﻊ ﮔﻮﻧﻪﻫﺎی ﻓﻌﺎل اﮐﺴﯿﮋن (ROS) ﮐﻪ ﺑﻪ ﻋﻨﻮان اﻧﻔﺠﺎر اﮐﺴﯿﺪی ﻣﻌﺮوف اﺳﺖ، بهره می­گیرند. گونه­های ﻓﻌﺎل در ﻣﻘﺪار اﻧﺪک ﺑﻪ ﻋﻨﻮان ﭘﯿﺎم رﺳﺎن­های ثانویه در ﺟﺮﯾﺎن ﻓﺮاﯾﻨﺪﻫﺎی دﻓﺎﻋﯽ ﮔﯿﺎه و ﺳﺎزﮔﺎری در ﻣﻘﺎﺑﻞ اﺳﺘﺮسﻫﺎی ﻣﺨﺘﻠﻒ ﻧﻘﺶ اﺳﺎﺳﯽ را ﺑﺮ ﻋﻬﺪه دارﻧﺪ اما ﺗﺪاوم ﺗﻨﺶ و ﺗﺠﻤﻊ ﺑﯿﺶ از ﺣﺪ ﮔﻮﻧﻪﻫﺎی ﻓﻌﺎل، ﺗﻌﺎدل اﮐﺴﺎﯾﺶ-ﮐﺎﻫﺶ در ﮔﯿﺎه را ﺑﺮ ﻫﻢ زده و ﻣﻮﺟﺐ تنش اکسیدی ﻣﯽﺷﻮد. ﮔﻮﻧﻪﻫﺎی ﻓﻌﺎل اﮐﺴﯿﮋن در ﮐﻠﺮوﭘﻼﺳﺖ، ﻣﯿﺘﻮﮐﻨﺪری، ﭘﺮاﮐﺴﯽزوم در ﺳﯿﺘﻮﭘﻼﺳﻢ، ﭘﺮاﮐﺴﯿﺪاز و آﻣﯿﻦ­اﮐﺴﯿﺪاز ﻣﻮﺟﻮد در دﯾﻮاره ﺳﻠﻮﻟﯽ و آﻧﺰﯾﻢﻫﺎی خانواده NADP(H) oxidase (NOX) ﻏﺸﺎی ﺳﻠﻮﻟﯽ ﺗﻮﻟﯿﺪ ﻣﯽﺷﻮﻧﺪ. در این میان ژن­های NAD(P)H oxidase با تحریک ژن‌های مرتبط با ایجاد گونه­های فعال اکسیژن نقش بسیار مهمی در پاسخ گیاه به تغییرات محیطی ایفا می­کنند. این خانواده ژنی همچنین ﺑﺎ اﻧﺘﻘﺎل اﻟﮑﺘﺮون از NAD(P)H ﺑﻪ اﮐﺴﯿﮋن ﻣﻮﻟﮑﻮﻟﯽ ﯾﻮن ﺳﻮﭘﺮاﮐﺴﯿﺪ را ﺗﻮﻟﯿﺪ کرده و در ادامه قادر به تولید انواع گونه­های فعال اکسیژن هستند. این پژوهش با هدف بررسی ناحیه راه­انداز ژن رمزکننده NAD(P)H oxidase در ژنوم گندم جهت شناسایی عناصر تنظیمی تاثیرگذار بر بیان آن در گندم انجام گردید.
مواد و روش‌ها: راه انداز ژن­ها و عناصر تنظیمی مهمی که در آن قرار دارد نقش مهمی در تنظیم بیان ژن در شرایط مختلف و در در نتیجه در ایجاد سازگاری گیاهان با تغییرات محیطی را بر عهده دارد. جهت بررسی چگونگی تنظیم بیان ژن NAD(P)H oxidase ناحیه راه­انداز ژن با استفاده از پایگاه داده‌های بیوانفورماتیک و نرم افزارهای تحلیلی، مورد بررسی قرار گرفت. ابتدا با استفاده از سایت NCBI توالی مربوط به ژن رمزکننده این آنزیم در گندم شناسایی شد. جهت شناسایی عناصر تنظیمی در بالادست این ژن در ناحیه راه‌انداز ژنی 1500 جفت باز از ناحیه آغاز رونویسی جدا گردید و سپس این توالی‌ها با استفاده از سایت PlantEnsemble مورد بررسی قرار گرفت. برای شناسایی عناصر تنظیمی مهم در بیان ژن که هر کدام نقش مشخصی در تنش­های مختلف ایفا می کنند، توالی راه­انداز توسط سایت PlantCare بررسی شد.
یافته‌ها: نتایج نشان داد که عناصر تنظیمی مختلفی که در مسیرهای بیوشیمیایی مختلف دخالت دارند می‌توانند بر میزان بیان ژن های NAD(P)H oxidase در گندم تاثیر گذار باشند. از جمله عناصر تنظیمی که به وسیله آنالیز راه­انداز شناسایی شدند می‌توان ABRE, TCA motif, TGACG motif, ARE, CCAAT-Box, skn-1-motif, circadian را نام برد که هر کدام به وسیله هورمون­ها و فاکتورهای رونویسی مختلف شناسایی می شوند.
نتیجه‌گیری: تحلیل­ها نشان داد تغییرات شرایط محیطی توانایی ایجاد تغییر بیان ژن آنزیم NAD(P)H oxidase را دارند. هورمون­ها و فاکتورهای رونویسی هر کدام عناصر تنظیمی خاصی را در راه­انداز ژن مورد هدف قرار می­دهند و از این طریق موجب تغییر میزان بیان آنزیم می­شوند. آنزیم در ابتدا با ایجاد یون سوپراکسید و در ادامه تولید انواع گونه­های فعال اکسیژن که نقش پیام­رسان ثانویه در گیاه را دارند موجب سازگاری گیاه با شرایط محیطی می­شود. از جمله عناصر تنظیمی که به وسیله آنالیز راه­انداز شناسایی شدند می‌توان ABRE, TCA motif, TGACG motif, ARE, CCAAT-Box, skn-1-motif, circadian را نام برد. این عناصر به ترتیب در مسیرهای پاسخ به هورمون­های آبسیزیک اسید، ﺳﺎﻟﯿﺴﯿﻠﯿﮏ اﺳﯿد و جاسمونیک اسید، مسیرهای واکنش­های غیر هوازی، پاسخ به خشکی از طریق فاکتورهای رونویسی MYB، مسیر آندوسپرم گیاه و واکنش­های نوری نقش دارند که همگی موجب پاسخ بهتر گیاه در مراحل مختلف رشدی و سازگاری با شرایط متغیر محیطی می­شوند.

کلیدواژه‌ها

موضوعات

عنوان مقاله [English]

Investigation of NAD(P)H oxidase genes regulatory elements in wheat

نویسندگان [English]

  • Fatemeh Sohrabi 1
  • Armin Saed-Moucheshi 2
1 Depratment of Plant Biotechnology, College of Agriculture, Shiraz University, Shiraz, Iran
2 Department of Crop and Horticulture Reseach, Kermanshah Agricultural and Natural Resources Research and Education Center, AREEO, Kermanshah, Iran
چکیده [English]

Introduction: Climate changes and increase in severity and duration of abiotics stresses have considerably affected the crops’, e.g. wheat’s, productivity. Accordingly, evaluating the influences of environmental stresses at cellular and molecular level to distinguish regulatory and tolerance mechanisms in plants has become a priority in the world. In response to stress conditions plants employ altered mechanism including a rapid induction in production of reactive oxygen species (ROS) known as oxidative burst. ROS are helpful in a small amount and they are mostly used in cell signaling and as secondary messengers, but excessive accumulation of ROS under environmental stress disturbs the oxidation-reduction balance in the plant and causes oxidative stress. ROSs are normally produced in chloroplasts, mitochondria, peroxisomes in the cytoplasm, peroxidase and amine-oxidase in the plants’ cell walls along with NADP(H) oxidase (NOX) family enzymes available in plants’ cell membrane. NADP(H) oxidase genes play a very important role in plant response to environmental changes by regulating the transportation of electrons from NAD(P)H molecules to molecular oxygen and producing superoxide ion which further leads to create various types of ROS. This research was conducted aiming to investigate the promoter region of the gene encoding NADP(H) oxidase enzyme in wheat genome in order to find the effect of important regulatory elements on expression.
Materials and methods: The promoter region of genes and regulatory elements that are located within this region play an important role in the regulation of gene expression in plants under different conditions. In order to recognize the regulation of gene expression for NAD(P)H oxidase gene, the promoter region of the gene was investigated by using online tools. First, the sequence of the gene encoding this enzyme in wheat was downloaded using the NCBI site. Then, 1500 base pairs upstream from the transcription start point were assessed by using online tools in PlantEnsemble site. The promoter sequence was finally analyzed by PlantCare website to identify important regulatory elements with specific roles in gene expression.
Results: The results showed that a diverse set of regulatory elements involved in different biochemical pathways can affect the expression level of NAD(P)H oxidase gene in wheat. the most effective elements that were identified by promoter analysis were ABRE, TCA motif, TGACG motif, ARE, CCAAT-Box, skn-1-motif, circadian which their activities are regulated by some specific hormones and transcription factors.
Conclusion: The analysis showed that changes in environmental conditions have the ability to change NAD(P)H oxidase gene expression. Hormones and transcription factors each target specific regulatory elements in the gene promoter leading tochange the level of enzyme expression. NADPH oxidase enzyme manages the production of superoxide ion leading to production ofother types of ROSs in the plant. ABRE, TCA motif, TGACG motif, ARE, CCAAT-Box, skn-1-motif, circadian can be mentioned among the regulatory elements that were identified by promoter analysis and they are involvedin the response pathways to abscisic acid, salicylic acid, and jasmonic acid hormones, anaerobic reaction pathways, response to drought through MYB transcription factors, plant endosperm pathway, and light reactions, respectively. According to scientific contexts, the expression of these elements can improve the response of the plant to harsh conditions and environmental stresses in different growth stages and are playing an important role in plants adaptation to changed environments.

کلیدواژه‌ها [English]

  • NAD(P)H oxidase
  • reactive oxygen species
  • initiator
  • transcription factors

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