The BpFLA1 and BpFLA2 genes are involved in regulating secondary xylem development in Betula platyphylla and respond to artificial bending

Backgrounds and Aims Fasciclin-like arabinogalactan proteins (FLAs) function in diverse plant physiological processes, particularly cell wall biosynthesis and tension wood (TW) formation. This study aimed to identify Betula platyphylla FLA genes associated with secondary xylem and TW development, thereby elucidating molecular mechanisms underlying xylogenesis to facilitate molecular improvement strategies. Methods Tissue-specific expression of two FLA genes (BpFLA1, BPFLA2) was analyzed via quantitative real-time PCR. BpFLA1/2-overexpressing transgenic lines and bpfla1 knockout mutants were generated using the pROKⅡ vector and CRISPR-Cas9 technology. Anatomical phenotypes, cell wall composition, and TW development were subsequently investigated. Key Results Two FLA genes (BpFLA1, BpFLA2) were identified in the birch TW transcriptome. Phylogenetically, BpFLA1 and BpFLA2 exhibit conserved evolutionary relationships and structural domains. Both genes were highly expressed in mature stems but suppressed by mechanical stimuli. BpFLA1/2-overexpressing (OE) transgenics displayed slightly suppressed growth, reduced cell wall thickness, enlarged xylem vessel area, and notably inhibited TW formation following artificial bending. Conversely, bpfla1 knockout mutants exhibited significant thickening of xylem fiber cell walls—a phenotype contrasting sharply with OE lines. Conclusion BpFLA1/2 functions as a critical regulator of cell wall development and a negative regulator of xylem fiber and TW formation in birch. Therefore, this study identifies BpFLA1/2 as a novel target for genetic improvement. BpFLA1/2 overexpression potentially enhances wood straightness and quality, whereas its suppression or knockout may benefit cellulose production for pulpwood applications.