Single-cell atlases reveal leaf cell-type-specific regulation of metal transporters in the hyperaccumulator Sedum alfredii under cadmium stress.

Hyperaccumulation in plants is a complex and dynamic biological process. Sedum alfredii, the most studied Cd hyperaccumulator, can accumulate up to 9000 mg kg Cd in its leaves without suffering toxicity. Although several studies have reported the molecular mechanisms of Cd hyperaccumulation, our understanding of the cell-type-specific transcriptional regulation induced by Cd remains limited. In this study, the first full-length transcriptome of S. alfredii was generated using the PacBio Iso-Seq technology. A total of 18,718,513 subreads (39.90 Gb) were obtained, with an average length of 2133 bp. The single-cell RNA sequencing was employed on leaves of S. alfredii grown under Cd stress. A total of 12,616 high-quality single cells were derived from the control and Cd-treatment samples of S. alfredii leaves. Based on cell heterogeneity and the expression profiles of previously reported marker genes, seven cell types with 12 transcriptionally distinct cell clusters were identified, thereby constructing the first single-cell atlas for S. alfredii leaves. Metal transporters such as CAX5, COPT5, ZIP5, YSL7, and MTP1 were up-regulated in different cell types of S. alfredii leaves under Cd stress. The distinctive gene expression patterns of metal transporters indicate special gene regulatory networks underlying Cd tolerance and hyperaccumulation in S. alfredii. Collectively, our findings are the first observation of the cellular and molecular responses of S. alfredii leaves under Cd stress and lay the cornerstone for future hyperaccumulator scRNA-seq investigations.