**1. Introduction**

Iron (Fe) is a trace mineral element necessary for the normal life activities of almost all living organisms including plants. It is the fourth most abundant element in the earth's crust. Despite the high total iron content in soils, the soluble iron (Fe2+) fraction is easily fixed to the insoluble form (Fe3+) in an alkaline environment, which seriously affects the normal uptake of iron by plants. Iron deficiency impairs photosynthetic efficiency, plant growth and biomass yield [1,2]. As a redox-active metal, Fe is engaged in photosynthesis, mitochondrial respiration, nitrogen anabolism, hormone (ethylene, gibberellic acid, jasmonic acid) synthesis and pathogen defense [3]. Iron also acts as the cofactor of many antioxidant enzymes, and thus iron deficiency has a regulatory effect on the antioxidant mechanisms, including the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) in plants [4], which are in charge of protecting the biological system against the harmful effects of reactive oxygen species (ROS) [5]. ROS are produced in all forms of aerobic life under stress or normal conditions. The excessive production of ROS causes oxidative damage that has a negative impact on the function of important

**Citation:** Shi, Y.; Zhao, Y.; Yao, Q.; Liu, F.; Li, X.; Jin, X.; Zhang, Y.; Ahammed, G.J. Comparative Physiological and Transcriptomic Analyses Reveal Mechanisms of Exogenous Spermidine-Induced Tolerance to Low-Iron Stress in *Solanum lycopersicum* L. *Antioxidants* **2022**, *11*, 1260. https://doi.org/ 10.3390/antiox11071260

Academic Editor: Nafees A. Khan

Received: 7 June 2022 Accepted: 23 June 2022 Published: 27 June 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

macromolecules [6]. Thus, a better understanding of the mechanisms of plant response to iron stress can be useful to improve crop stress resilience and enhance crop yield and quality [7].

When plants are exposed to a low-Fe environment, insufficient iron uptake causes retarded growth, interveinal chlorosis and reduced plant fitness. In severe iron deficiency, chloroplasts are dissociated or vesiculated, thus affecting chlorophyll formation [8]. To ensure the normal growth of plants, phytohormone auxin accumulates in large amounts in the roots, promoting the development of lateral roots and positively regulating the transcriptional expression of the *FIT1* and *AHA2* genes. Thus, growth factors are involved in the plant Fe-deficiency response network through different pathways [9].

Under low-iron stress, plants enhance Fe uptake by the root system through two strategies: One strategy based on the reduction that occurs in all dicotyledons and nongrass monocotyledons, called strategy I, and another strategy relying on chelation, which is limited to monocotyledons, called strategy II [10]. Strategy-II plants produce plant iron carriers capable of chelating Fe3+, which are then absorbed by specific epidermal root cell plasma membrane transporters [11]. In tomatoes, on the one hand, as Strategy-I species, acidification of the root mesenchyme by plasma membrane H+-ATPase activity occurs to dissolve Fe3+, and reduction to Fe2+ by Fe3+-chelating reductase (FCR) activity increases iron solubility. Afterward, translocation of the resulting Fe2+ to the root cell via a specific Fe transporter (*IRT1*) takes place to accomplish iron acquisition in plants [12]. On the other hand, nitric oxide (NO) accumulates in the roots and promotes the expression of *FER/FIT*, as well as *IRT* and *FRO* genes, thus participating in the response to iron-deficiency stress in plants [13].

Polyamines are highly bioactive, low-molecular-weight aliphatic amines that occur as ubiquitous secondary metabolites in plants. Polyamines can bind to phospholipids and other biomolecules with negatively charged groups of nucleic acids and proteins through their ionic and hydrogen bonds, which widely affect the biological activity of plants [14]. In previous research reports, polyamines have been shown to perform an extremely important role in alleviating plant stress. Among the three widely distributed major polyamines, spermidine (Spd) plays a crucial role in abiotic stress tolerance. Due to its multivalent cation property, its physiological function is stronger and more associated with stress tolerance in plants [15]. Spd is a common polyamine in plants and is involved in adaptations to salinity [16], drought [17], cold [18] and heavy metals [19]. Some studies have shown that Spd modulates antioxidant enzyme activity and the expression of related genes in tomato seedlings exposed to high temperatures [20]. Exogenous Spd has been found to play an important role in remediating the effects of environmental stress on plants [21]. However, to date, few studies have reported on the Spd-mediated tolerance to iron stress, particularly in tomato plants.

In the present research, using 'Micro-tom' tomato as the object of study, we explored the effect of exogenously sprayed Spd on the growth, physiology and metabolism of tomato seedlings under low-Fe stress. The physiological analysis, combined with transcriptomic analyses, shed new light on the mechanism of Spd-mediated low-Fe tolerance in tomato seedlings from both physiological and molecular perspectives, which provides a theoretical basis for improving the uptake and utilization of Fe in protected cultivation.
