ABSTRACT
Hair-like trichomes cover the aerial organs of many plant species and act as a barrier between a plant and its environment. They function in defense against biotic and abiotic stresses, while also serving as sites for synthesis and storage of secondary metabolites. Previously, the transcription factor PtaMYB186 was identified as a positive regulator of trichome initiation during early stages of leaf development in Populus tremula x P. alba (IRNA 717-1B4). However, trichome regulation in poplar remains largely unexplored, as does the functional redundancy of duplicated poplar genes. Here, we employed CRISPR/Cas9 to target a consensus region of PtaMYB186 and its close paralogs for knockout. Regeneration of glabrous mutants suggested their essential roles in poplar trichome development. No apparent differences in growth and leaf transpiration rates between the mutants and the controls were observed, but trichomeless poplars showed increased insect pest susceptibility. RNA-seq analysis revealed widespread down-regulation of circadian- and light-responsive genes in the mutants. When exposed to a high light regime, trichomeless mutants accumulated significantly higher levels of photoprotective anthocyanins. Cuticular wax and whole leaf analyses showed a complete absence of triterpenes in the mutants, suggesting biosynthesis and storage of triterpenes in poplar occurs in the non-glandular trichomes. This work also demonstrates that a single gRNA with SNP-aware design is sufficient for multiplex targeting of paralogous genes in outcrossing and/or hybrid species with unexpected copy number variations.
ONE SENTENCE SUMMARY Non-glandular trichomes in poplar have roles both as a physical barrier and a chemical factory to mediate plant interactions with the environment.
Footnotes
The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors is Chung-Jui Tsai (cjtsai{at}uga.edu).
FUNDING The work was funded in part by The Center for Bioenergy Innovation, a U.S. Department of Energy Research Center supported by the Office of Biological and Environmental Research in the DOE Office of Science, the Division of Integrative Organismal Systems (grant nos. IOS-1546867) of the National Science Foundation, the Community Science Program of the Joint Genome Institute, a DOE Office of Science User Facility. The work conducted by the U.S. Department of Energy Joint Genome Institute is supported by the Office of Science of the U.S. Department of Energy under Contract No DE-AC02-05CH11231.