The Ethylene Responsive Factor (ERF) transcription factor family is important for regulating plant growth and stress responses. Although the expression patterns of ERF family members have been reported in many plant species, their role in Populus alba × Populus glandulosa, an important model plant for forest research, remains unclear. In this study, we identified 209 PagERF transcription factors by analyzing the P. alba × P. glandulosa genome. We analyzed their amino acid sequences, molecular weight, theoretical pI (Isoelectric point), instability index, aliphatic index, grand average of hydropathicity, and subcellular localization. Most PagERFs were predicted to localize in the nucleus, with only a few PagERFs localized in the cytoplasm and nucleus. Phylogenetic analysis divided the PagERF proteins into ten groups, Class I to X, with those belonging to the same group containing similar motifs. Cis-acting elements associated with plant hormones, abiotic stress responses, and MYB binding sites were analyzed in the promoters of PagERF genes. We used transcriptome data to analyze the expression patterns of PagERF genes in different tissues of P. alba × P. glandulosa, including axillary buds, young leaves, functional leaves, cambium, xylem, and roots, and the results indicated that PagERF genes are expressed in all tissues of P. alba × P. glandulosa, especially in roots. Quantitative verification results were consistent with transcriptome data. When P. alba × P. glandulosa seedlings were treated with 6% polyethylene glycol 6000 (PEG6000), the results of RT-qRCR showed that nine PagERF genes responded to drought stress in various tissues. This study provides a new perspective on the roles of PagERF family members in regulating plant growth and development, and responses to stress in P. alba × P. glandulosa. Our study provides a theoretical basis for ERF family research in the future.

D-type cyclins (CYCDs) are a special class of cyclins and play extremely important roles in plant growth and development. In the plant kingdom, most of the existing studies on CYCDs have been done on herbaceous plants, with few on perennial woody plants. Here, we identified a Populus d-type cyclin gene, PsnCYCD1;1, which is mainly transcribed in leaf buds and stems. The promoter of PsnCYCD1;1 activated GUS gene expression and transgenic Arabidopsis lines were strongly GUS stained in whole seedlings and mature anthers. Moreover, subcellular localization analysis showed the fluorescence signal of PsnCYCD1;1-GFP fusion protein is present in the nucleus. Furthermore, overexpression of the PsnCYCD1;1 gene in Arabidopsis can promote cell division and lead to small cell generation and cytokinin response, resulting in curved leaves and twisted inflorescence stems. Moreover, the transcriptional levels of endogenous genes, such as ASs, KNATs, EXP10, and PHB, were upregulated by PsnCYCD1;1. Together, our results indicated that PsnCYCD1;1 participates in cell division by cytokinin response, providing new information on controlling plant architecture in woody plants.

Wood is an abundant and essential renewable resource whose production is threatened in some parts of the world by drought. A better understanding of the molecular mechanisms underlying wood formation during drought is critical to maintaining wood production under increasingly adverse environmental conditions. In this study, we investigated wood formation in black cottonwood (Populus trichocarpa) during drought stress. The morphological changes during drought stress in P. trichocarpa included the wilting and drooping of leaves, stem water loss, and a reduction in whole plant biomass. The water embolism rate indicated that the water transport in stems was blocked under drought conditions. An anatomical analysis of the xylem and cambium revealed that drought stress changed the structure of vessel cells, increased lignin accumulation, and decreased the cambium cell layers. We subsequently identified 12,438 and 9156 differentially expressed genes from stem xylem and cambium tissues under well-watered and drought conditions, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that these genes were mainly involved in hormone signal transduction and amino sugar and nucleotide sugar metabolism. To further explore the molecular mechanism of wood formation in response to drought, we analyzed the expression patterns of the genes involved in lignin, cellulose, and hemicellulose biosynthesis in xylem and the genes involved in cambial activity in the cambium. To better understand the regulatory networks governing xylem development and cambium activity in response to drought, we analyzed the MYB (138), AP2 (130), bHLH (89), and NAC (81) transcription factor families to shed light on the interactions between the TFs in these families and the genes they regulate. Identifying the key genes that regulate wood formation in P. trichocarpa during drought provides a genetic foundation for further research on the molecular regulatory networks and physiology underpinning wood formation during drought stress.