Physiological, Photosynthetic Characteristic and Transcriptome Analysis of PsnWRKY70 Transgenic Populus simonii × Populus nigra Under Salt Stress

The <i>Psn</i>WRKY70 transcription factor (TF) was reported to play an important role in the salt stress response mechanism of <i>Populus simonii × Populus nigra</i> in our previous research, and we also produced several <i>PsnWRKY70</i> overexpression (OEXs) and RNAi suppression (REXs) <i>P. simonii</i> × <i>P. nigra</i> lines. In order to further compare the photosynthetic and physiological characteristics of NT (non-transgenic line) and transgenic lines under salt stress, the dynamic phenotypic change, Na⁺ and K⁺ content in leaf and root tissues, superoxide dismutase (SOD) and peroxidase (POD) activity, malondialdehyde (MDA) content, chlorophyll content (Chl), photosynthesis parameters (net photosynthetic rate, P_n; stomatal conductance, Gs; intercellular CO₂ concentration, C_i; transpiration rate, T_r), chlorophyll fluorescence parameters (electron transport rate, ETR; maximum photochemical efficiency of photosystem II (PSII), F_v/F_m; actual efficiency of PSII, Φ_PSII; photochemical quenching coefficient, q_P; non-photochemical quenching, NPQ; the photosynthetic light-response curves of Φ_PSII and ETR) and RNA-seq of NT, OEX and REX lines were detected and analyzed. The phenotypic observation, MDA content and Chl detection results indicate that the stress damage of REXs was less severe than that of NT and OEX lines under salt stress. Photosynthesis parameter (P_n, Gs, T_r and C_i) and chlorophyll fluorescence parameter (ETR, F_v/F_m, Φ_PSII q_P and NPQ) detection results indicate that the REX lines exhibited much better photosynthetic adaptability than NT and OEX lines during salt stress. The photosynthetic light-response curves of Φ_PSII and ETR of NT, OEX and REX lines indicate that REXs exhibited better ability to activate the photosynthetic protection mechanism and adapt to a certain degree of strong light than NT and OEX lines under salt stress. RNA-seq analysis indicates that the DEGs between OEX1 vs. NT and REX1 vs. NT in different tissues (apical bud and fifth functional leaf) were all different in category and change trend. The expression of <i>PsnWRKY70</i> was significantly up-regulated in both the apical bud and fifth functional leaf of OEX1, and showed no significant change (namely maintained low expression level) in both the apical bud and fifth functional leaf of REX1, thus indicating the negative regulation role of <i>PsnWRKY70</i> in <i>P. simonii</i> × <i>P. nigra</i> under salt stress. Additionally, there were a lot of stress response-related TF genes (such as <i>bHLH</i>, <i>WRKY</i>, <i>MYB</i>, <i>NAM</i> and <i>AP2/EREBP</i>) and photosynthesis-related genes among all the DEGs. In REX1, the expression of three <i>Photosystem I P700 chlorophyll a apoprotein A1</i> genes (Potri.003G065200, Potri.013G141800 and Potri.019G028100) and a <i>Photosystem II protein D1</i> gene (Potri.013G138300) were significantly up-regulated after 6 days of salt stress. In OEX1, the <i>Heterodimeric geranylgeranyl pyrophosphate synthase small subunit</i> gene (Potri.015G043400) and <i>Phospho-2-dehydro-3-deoxyheptonate aldolase 1</i> gene (Potri.007G095700) were significantly down-regulated after 6 days of salt stress. These photosynthesis-related genes are probably regulated by <i>Psn</i>WRKY70 TF in response to salt stress. In conclusion, the REX lines suffered less severe salt damage and exhibited better photosynthetic adaptability than NT and OEXs under salt stress. The differences among the DEGs between OEX1 vs. NT and REX1 vs. NT in apical bud and fifth functional leaf, and the significantly differentially expressed photosynthesis-related genes are probably the key clues for discovering the photosynthesis adaptability mechanism of <i>PsnWRKY70</i> transgenic <i>P. simonii</i> × <i>P. nigra</i> under salt stress.