Cold stress involves CBF dependent regulatory pathway to remodel wood formation in Eucalyptus gunnii hybrids.

While being the most planted tree worldwide, most Eucalyptus species are sensitive to frost. Long-term exposure to cold temperatures, among other abiotic cues, triggers modification of secondary xylem differentiation in Eucalyptus, but the molecular mechanisms behind it remain unknown. Overexpression of key players of cold signalling pathway, the CRT-repeat binding factors (CBF), not only causes the expected increase in freezing tolerance but also remodels wood in a similar trend to cold acclimation, making it a good candidate for wood functional adaptation to cold stress. To gain insight in CBF role in cold-induced secondary cell wall (SCW) remodelling, we used both targeted and untargeted methodologies to show that chilling and freezing temperatures induced the deposition of a thick SCW with alterations in lignin and polysaccharides composition as well as modifications of wood anatomy in a Eucalyptus cold-tolerant hybrid. Using co-expression network approaches, we identified CBF transcription factors (TFs) as a regulatory hub in xylem cold response. Direct targets of the CBF TFs were identified by DAPseq and unravelled promising candidates involved in SCW deposition and hormonal signalling pathways. Our results shed new light on the interplay between cold response and wood formation, bringing new evidence for the role of the cell wall in trees tolerance to abiotic stresses.