Sedum alfredii Hance, a cadmium (Cd)/zinc (Zn)/lead (Pb) co-hyperaccumulating species, is a promising phytoremediation candidate because it accumulates substantial amounts of heavy metal ions without showing any obvious signs of poisoning. The heat shock transcription factor (Hsf) family plays crucial roles in plant growth, development, and stress responses. Although the roles of some Hsfs in abiotic stress have been well studied in model plants, the Hsf family has not been systematically investigated in heavy metal hyperaccumulators. Here, we comprehensively analyzed the Hsf gene family in S. alfredii based on a transcriptome under Cd stress. There were 22 Hsf members that were identified and phylogenetically clustered into three classes, namely, SaHsfA, SaHsfB, and SaHsfC. All of the three classes shared similar motifs. The expression profiles of the 22 Hsf members showed significant differences: 18 SaHsfs were responsive to Cd stress, as were multiple SaHsp genes, including SaHsp18.1, SaHsp22, SaHsp26.5, SaHsp70, SaHsp90, and SaHsp101. Two class A4 members, SaHsfA4a and SaHsfA4c, exhibited transcriptional activation activities. Overexpression of SaHsfA4a and SaHsfA4c in transgenic yeast indicated an improved tolerance to Cd stress and Cd accumulation. Our results suggest SaHsfs play important regulatory roles in heavy metal stress responses, and provide a reference for further studies on the mechanism of heavy metal stress regulation by SaHsfs.

Salix viminalis L., a dioecious species, is widely distributed in riparian zones, and flooding is one of the most common abiotic stresses that this species suffers. In this study, we investigated the morphological, anatomical, and physiological responses of male vs. female plants of S. viminalis to flooding. The results showed that the plant height and root collar diameter were stimulated by flooding treatment, which corresponded with higher dry weight of the stem and leaf. However, the dry weight of the underground part decreased, which might be due to the primary root having stopped growing. The little-influenced net photosynthesis rate (P_n) under flooding treatment could guarantee rapid growth of the aboveground part, while the unaffected leaf anatomical structure and photosynthetic pigment contents could ensure the normal operation of photosynthetic apparatus. Under a flooding environment, the production ratio of superoxide free radical (O₂^∙-) and malondialdehyde (MDA) contents increased, indicating that the cell membrane was damaged and oxidative stress was induced. At the same time, the antioxidant enzyme system, including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX), and osmotic adjustment substances, involving proline (Pro) and solute protein (SP), began to play a positive role in resisting flooding stress. Different from our expectation, the male and female plants of S. viminalis performed similarly under flooding, and no significant differences were discovered. The results indicate that both male and female plants of S. viminalis are tolerant to flooding. Thus, both male and female plants of S. viminalis could be planted in frequent flooding zones.