The heavy metal ATPase (HMA) family plays an important role in transition metal transport in plants. However, this gene family has not been extensively studied in Populus trichocarpa. We identified 17 HMA genes in P. trichocarpa (PtHMAs), of which PtHMA1–PtHMA4 belonged to the zinc (Zn)/cobalt (Co)/cadmium (Cd)/lead (Pb) subgroup, and PtHMA5–PtHMA8 were members of the copper (Cu)/silver (Ag) subgroup. Most of the genes were localized to chromosomes I and III. Gene structure, gene chromosomal location, and synteny analyses of PtHMAs indicated that tandem and segmental duplications likely contributed to the expansion and evolution of the PtHMAs. Most of the HMA genes contained abiotic stress-related cis-elements. Tissue-specific expression of PtHMA genes showed that PtHMA1 and PtHMA4 had relatively high expression levels in the leaves, whereas Cu/Ag subgroup (PtHMA5.1- PtHMA8) genes were upregulated in the roots. High concentrations of Cu, Ag, Zn, Cd, Co, Pb, and Mn differentially regulated the expression of PtHMAs in various tissues. The preliminary results of the present study generated basic information on the HMA family of Populus that may serve as foundation for future functional studies.

Trihelix genes play important roles in plant growth and development and responses to biotic and abiotic stresses. Here, we identified 56 full-length trihelix genes in Populus trichocarpa and classified them into five groups. Most genes within a given group had similar gene structures and conserved motifs. The trihelix genes were unequally distributed across 19 different linkage groups. Fifteen paralogous pairs were identified, 14 of which have undergone segmental duplication events. Promoter cis-element analysis indicated that most trihelix genes contain stress- or phytohormone-related cis-elements. The expression profiles of the trihelix genes suggest that they are primarily expressed in leaves and roots. Quantitative real-time reverse transcription polymerase chain reaction analysis indicated that members of the trihelix gene family are significantly induced in response to osmotic, abscisic acid, salicylic acid, methyl jasmonate and pathogen infection. PtrGT10 was identified as a target gene of miR172d, which is involved in the osmotic response. Repression of PtrGT10 could increase reactive oxygen species scavenging ability and decrease cell death. This study provides novel insights into the phylogenetic relationships and functions of the P. trichocarpa trihelix genes, which will aid future functional studies investigating the divergent roles of trihelix genes belonging to other species.

The AT-hook motif nuclear-localized (AHL) family is a plant transcription factor family, which plays an important role in growth and development and stress responses. We identified and analyzed 37 AHL genes in poplar (Populus trichocarpa). Phylogenetic analysis classified the PtrAHL members into three subfamilies based on their conserved domain. All PtrAHL paralogous pairs evolved under purifying selection. The promoter analysis revealed the presence of stress-related and phytohormone-related cis-elements of the PtrAHL genes. Our analysis of the tissue-specific expression pattern of PtrAHL genes indicated their significance in tissue and organ development. Network-based prediction suggested that PtrAHL genes may interact with histone deacetylases (HDAC) and participate in the development of organs, such as roots. Drought negatively impacts plant growth and development. ABA is produced under osmotic stress condition, and it takes an important part in the stress response and tolerance of plants. Real-time quantitative PCR (qRT-PCR) showed that PtrAHL genes were induced by drought stress and ABA treatment. These insights into the expression of PtrAHL genes under stress provide a basis for PtrAHL gene functional analysis. Our study will help develop new breeding strategies to improve drought tolerance in poplar.

Summary Adventitious rooting is an essential biological process in the vegetative propagation of economically important horticultural and forest tree species. It enables utilization of the elite genotypes in breeding programmes and production. Promotion of adventitious root (AR) formation has been associated with starvation of inorganic phosphate and some factors involved in low phosphorus (LP) signalling. However, the regulatory mechanism underlying LP‐mediated AR formation remains largely elusive. We established an efficient experimental system that guaranteed AR formation through short‐term LP treatment in Populus ussuriensis. We then generated a time‐course RNA‐seq data set to recognize key regulatory genes and regulatory cascades positively regulating AR formation through data analysis and gene network construction, which were followed by experimental validation and characterization. We constructed a multilayered hierarchical gene regulatory network, from which PuMYB40, a typical R2R3‐type MYB transcription factor (TF), and its interactive partner, PuWRKY75, as well as their direct targets, PuLRP1 and PuERF003, were identified to function upstream of the known adventitious rooting genes. These regulatory genes were functionally characterized and proved their roles in promoting AR formation in P. ussuriensis. In conclusion, our study unveiled a new hierarchical regulatory network that promoted AR formation in P. ussuriensis, which was activated by short‐term LP stimulus and primarily governed by PuMYB40 and PuWRKY75.