Cellulosimicrobium funkei strain AR6 alleviate Cr(VI) toxicity in Lycopersicon esculentum by regulating the expression of growth responsible, stress tolerant and metal transporter genes
Introduction
Heavy metal contaminations in agricultural land is an important environmental concern, which has augmented extremely since the beginning of the industrial revolution (Ali et al., 2019). Particularly, chromium (Cr) is one of the prominent and common environmental pollutant, that has been largely released into the environment from various industrial processes such as leather tanning, textile dyeing, chrome plating, etc., (Gill et al., 2015). It is well known that Cr has nine valence states (from –II to +VI), out of them Cr(III) and Cr(VI) are recognized as relatively stable oxidation form and found predominantly in the environment. However, the strong oxidizing nature of Cr(VI) can potentially induce various carcinogenic and mutagenic effects on all form of living organisms due to its toxicity and mobility. Moreover, Cr(VI) gets easily enter in to the cell through non-specific membrane transport channels and modulates the expression of various functional genes (Urbano et al., 2008). Further, intensive accumulation of Cr in soil adversely affects the microbial biomass, diversity, structure and enzymatical activity (Ashitha et al., 2020). Although, Cr is a non–essential element for plant growth and metabolism, a certain amount has been transported to the tissues along with the essential nutrients from the soil via metal transporters and symplast/apoplast pathway (Ghori et al., 2019). Such excess accumulation of Cr in plant tissues induces various biochemical changes including induction of lipid peroxidation, biosynthesis of ethylene, generation of reactive oxygen specis (ROS), alteration of antioxidant activities and loss in plasma membrane intregrity, which collectivley reduce the plant growth and yield (Gill et al., 2015; Gupta et al., 2018). In addition, Cr toxicity has a significant importance in altering the expression of specific genes associated with plant growth, abiotic stress tolerance and metal uptake (Oliveira, 2012).
In this context, free living soil bacteria harboring plant roots collectively known as plant growth promoting rhizobacteria (PGPR), acquired significant important in combating heavy metal stress and enhancing plant growth (Rajkumar et al., 2017). In general, PGPR have the ability to regulate metal accumulation and reduce the toxicity in plants through altering (precipitation, biotransformation, adsorption, biosorption, bioaccumulation, immobilization, complexation and alkalization) the metal availability in the rhizospheric soil (Manoj et al., 2020). Moreover, they can also promote the plant growth in metal contaminted soil through direct (production of phytohormones and other plant growth promoting substances) and indirect (soil nutrient mobilization/recycling, metal toxicity reduction and pathogen and nematode bicontrol) plant growth promoting mechanisms (Manoj et al., 2020; Rajkumar et al., 2012). In addition, such PGPR have also been shown to improve the plant tolerance towards metal stress by altering the osmolyte accumulation and antioxidant enzyme activities (Karthik et al., 2016).
Lycopersicon esculentum Mill. is a major economic crop and widely cultivated across the world due to its high nutritive value and processed industrial products. The plant is highly sensitive to heavy metal particularly Cr, that affect the growth and various physiological behavior of the plant (Singh et al., 2020; Khanna et al., 2019a). Although numerous PGPR have been exploited to study the effects on L. esculentum under heavy metal stress, most of these studies exclusively spotlight the physiological parameters and metal accumulation (Khanna et al., 2019b; Khan et al., 2017). The alteration in gene expression is also responsible for metal accumulation and synthesis of compatible osmolytes, which adapted by the plants as a protective mechanism to tolerate metal stress. However, the molecular mechanism behind the PGPR mediated plant growth promotion, stress tolerance and metal transports in L. esculentum to Cr stress is still unclear. Hence, the present study was aimed to identify the influence of Cellulosimicrobium funkei AR6 inoculation on growth (LeEXP1, LeEXP2 and LeEXP3), stress mitigation (P5CS, DHN, HSP21, HSP70, HSP90, MT1, MT2, MT3 and MT4) and metal transporter (NRAMP1, NRAMP2 and NRAMP3) genes expression in L. esculentum seedlings under Cr(VI) stress with an objective to explore the possible mechanisms involved in PGPR-mediated Cr tolerance.
Section snippets
Chemicals and Cr(VI) stock solution preparation
For experimental studies, analytical grade chemicals were purchased from HiMedia (Mumbai) and Merck (India). PCR master mix was purchased from Bioline (UK). RNAiso Plus (Total RNA extraction reagent), SYBR Premix Ex Taq™ (Tli RNaseH Plus) and ROX plus were purchased from TaKaRa (TAKARA BIO INC, Japan). cDNA synthesis kit was purchased from Thermo Fisher Scientific (USA). For Cr(VI) stock solution preparation (1000 mg/l), potassium dichromate (2.829 g) was dissolved in double distilled (1000 ml)
Effect of Cr(VI) and C. funkei AR6 inoculation on growth of L. Esculantum
In the present study, the positive correlation between C. funkei AR6 inoculation and L. esculantum under Cr(VI) stress was validated through various physiological, biochemical and molecular expression analysis. In growth parameter analysis, higher concentrations of Cr(VI) significantly reduced the growth parameters of L. esculantum. The maximum reduction in root length (55.95%), shoot length (52.34%) and biomass (57.30%) were found in uninoculated seedlings treated with 200 μg/ml exposure
Discussion
Rhizobacterial strain AR6 produced various plant growth promoting substances (Karthik et al., 2016), which motivated us to assess its effect on physiological, biochemical and molecular response of L. esculentum seedlings under Cr(VI) stress. When L. esculentum seedlings exposed to different concentrations of Cr(VI), growth parameters were decreased drastically, which could be due to the toxicity effect excreted by Cr(IV). Cr(VI) is a well-known toxic heavy metal, which affects the growth and
Conclusion
Cr(VI) exposure could induce a wide range of toxic effects on growth and other biochemical processes of the plants, which could be minimized by inoculating potential Cr(VI) reducing PGPR strains. The present finding revealed that the inoculation of C. funkei AR6 could improve the growth and ameliorate Cr(V) induced toxicity by regulating the expression of major metal transporter NRAMP genes, resulting decrease in the accumulation of Cr. The bacterial inoculation also influences the cell wall
Declaration of competing interest
The authors declare that they have no conflict of interest.
Acknowledgments
Dr. C.K thankful to the Science and Engineering Research Board (SERB, India) for providing National Post-doctoral Fellowship (File No: PDF/2016/002058). The authors would like to express our heartfelt thanks to DRDO – BU – Center for Life Sciences for providing laboratory facilities.
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