Drought is one of the main environmental factors limiting plant growth and development. The AP2/ERF transcription factor (TF) ERF194 play key roles in poplar growth and drought-stress tolerance. However, the physiological mechanism remains to be explored. In this study, the ERF194-overexpression (OX), suppressed-expression (RNA interference, RNAi), and non-transgenic (WT) poplar clone 717 were used to study the physiology role of ERF194 transcription factor in poplar growth and drought tolerance. Morphological and physiological methods were used to systematically analyze the growth status, antioxidant enzyme activity, malondialdehyde (MDA), soluble sugars, starch, and non-structural carbohydrate (NSC) contents of poplar. Results showed that, compared with WT, OX plants had decrease in plant height, internode length, and leaf area and increased number of fine roots under drought stress. In addition, OX had higher water potential, activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), contents of chlorophyll, soluble sugar, starch, and NSC, implying that ERF194 positively regulates drought tolerance in poplar. The growth status of RNAi was similar to those of WT, but the relative water content and CAT activity of RNAi were lower than those of WT under drought treatment. Based on the transcriptome data, functional annotation and expression pattern analysis of differentially expressed genes were performed and further confirmed by RT-qPCR analysis. Gene ontology (GO) enrichment and gene expression pattern analysis indicated that overexpression of ERF194 upregulated the expression of oxidoreductases and metabolism-related genes such as POD and SOD. Detection of cis-acting elements in the promoters suggested that ERF194 may bind to these genes through MeJA-responsive elements, ABA-responsive elements, or elements involved in defense and stress responses. The above results show that ERF194 improved tolerance to drought stress in poplar by regulating its growth and physiological factors. This study provides a new idea for the role of ERF194 transcription factor in plant growth and drought-stress response.

Research Highlights: Sequence phylogeny, genome organisation, gene structure, conserved motifs, promoter cis-element and expression profiling of poplar NACs related to salt stress were detected. In addition, expression of two salt-induced NACs was analysed. Background and Objectives: NAC transcription factor (TF) proteins are involved in a wide range of functions during plant development and stress-related endurance processes. To understand the function of Populus NAC TFs in salt stress tolerance, we characterised the structure and expression profile of a total of 289 NAC members. Materials and Methods: Sequence phylogeny, genome organisation, gene structure, motif composition and promoter cis-element were detected using bioinformatics. The expression pattern of Populus NAC TFs under salt stress was also detected using RNA-Seq and RT-qPCR. Results: Synteny analysis showed that 46 and 37 Populus NAC genes were involved in whole-genome duplication and tandem duplication events, respectively. The expression pattern of Populus NAC TFs under salt stress showed the expression of the 289 PtNACs of 84K poplar was induced. Similar expression trends of NACs were found in Populus simonii × P. nigra T. S. Hwang et Liang and Arabidopsis thaliana (L.) Heynh. Conclusions: The correlation analysis showed that the expression of two differentially expressed NAC genes PtNAC024 and PtNAC182 was significantly associated with most of the 63 differentially expressed genes tested. The expression of PtNAC024 and PtNAC182 in different tissues was also analysed in silico and different expression patterns were found. Together, this study provides a solid basis to explore stress-related NAC TF functions in Populus salt tolerance and development.

The water deficits limit the growth and development of agricultural and forest organisms. The AP2/ethylene response factor (ERF) family has been identified as one of the largest plant-specific transcription factors (TFs) essential for plant development and stress response. The function of PtaERF194 in growth and drought tolerance was detected in the overexpression (OX) and RNA interference (RNAi) transgenic poplar 717 hybrids (Populus tremula × Populus alba). Plant growth, stem vessels, water-use efficiency (WUE), chlorophyll content and PtaERF194 co-expressed genes were analyzed using morphological, physiological and molecular methods. Overexpression seedlings showed a shorter and smaller phenotype along with smaller and more vessels compared with the wild-type (WT). Physiological indices indicated that OX with low transpiration and stomatal conductance improved the tolerance to drought by enhancing WUE, limiting water loss and maintaining high water potential. A total of 12 differentially expressed genes co-expressed with PtaERF194 were identified, and they worked together to regulate drought tolerance through the abscisic acid signaling and reactive oxygen species scavenging processes. However, RNAi plants showed similar morphology and physiology to WT, suggesting that the function of PtaERF194 was redundant with other ERF TFs. The findings of the current study may shed new light on the positive function of ERF TFs in plant drought stress tolerance.

The aggravation of soil salinization limits the growth and development of plants. The AP2/ERF transcription factors (TFs) have been identified and play essential roles in plant development and stress response processes. In this study, the function of PagERF16 was detected using the overexpressing (OX) and RNAi transgenic poplar 84K hybrids. Plant growth, stomatal conductance, antioxidant enzymes activity, and PagERF16 co-expressed TFs were analyzed using morphological, physiological, and molecular methods. OX showed a more robust lateral root system with a bigger diameter and volume compared to the wild-type plants (WT). Physiological parameters indicated the bigger stomatal aperture and lower stomatal density of OX along with the lower Catalase (CAT) activity and higher malondialdehyde (MDA) content contributed to the salt sensitivity. The plant height and rooting rate of OX and RNAi were significantly worse compared to WT. Other than that, the morphology and physiology of RNAi plants were similar to WTs, suggesting that the function of PagERF16 may be redundant with other TFs. Our results indicate that when PagERF16 expression is either too high or too low, poplar growth and rooting is negatively affected. In addition, a downstream target TF, NAC45, involved in Auxin biosynthesis, was identified and PagERF16 could directly bind to its promoter to negatively regulate its expression. These results shed new light on the function of ERF TFs in plant root growth and salt stress tolerance.

MYB transcription factors (TF) are recognized as significant regulators in facilitating plant adaptation to various biotic and abiotic stresses. In order to elucidate the involvement of MYB TFs in the growth, development, and salt tolerance of poplar, PagMYB73A was cloned from 84K poplar (Populus alba × P. glandulosa) and utilized as the target gene for generating PagMYB73A overexpressing transgenic lines (OE-1 , OE-8 , OE-12). Compared to the wild type (WT) plants under normal conditions, the OE lines exhibited enhanced plant height and increased leaf number. While under NaCl treatment, the OE lines displayed elongated adventitious roots but reduced plant height compared to WT. At the transcriptional level, the expression of four genes associated with adventitious root growth and development was examined, revealing higher expression levels in WT compared to OE under normal conditions, but the opposite pattern was observed under salt stress. Physiological index analysis demonstrated a significant decrease in stomatal density of WT under salt stress, whereas the reduction was not significant in OE lines. Furthermore, OE lines exhibited decreased content of malondialdehyde (MDA), a marker of cellular membrane damage, compared to WT under salt stress. In summary, this study preliminarily revealed the regulatory function of PagMYB73A in poplar growth and salt stress resistance, highlighting its active role in the adaptation of poplar to salt stress.