Heat shock transcription factor (HSF) is an important TF that performs a dominant role in plant growth, development, and stress response network. In this study, we identified a total of 30 HSF members from poplar, which are unevenly distributed on 17 chromosomes. The poplar HSF family can be divided into three subfamilies, and the members of the same subfamily share relatively conserved domains and motifs. HSF family members are acidic and hydrophilic proteins that are located in the nucleus and mainly carry out gene expansion through segmental replication. In addition, they have rich collinearity across plant species. Based on RNA-Seq analysis, we explored the expression pattern ofPtHSFsunder salt stress. Subsequently, we cloned the significantly upregulatedPtHSF21gene and transformed it intoPopulus simonii×P. nigra. Under salt stress, the transgenic poplar overexpressingPtHSF21had a better growth state and higher reactive oxygen scavenging ability. A yeast one-hybrid experiment indicated PtHSF21 could improve salt tolerance by specifically binding to the anti-stress cis-acting element HSE. This study comprehensively profiled the fundamental information of poplar HSF family members and their responses to salt stress and specifically verified the biological function ofPtHSF21, which provides clues for understanding the molecular mechanism of poplar HSF members in response to salt stress.