An annual gene expression profile of markers for programmed cell death and polyphenols biosynthesis across heartwood forming stems in Robinia pseudoacacia

Heartwood formation is a developmental defense strategy in trees that transforms the inner stem core through the deposition of natural antimicrobial preservatives. The defining cellular events in this process are the accumulation of specialized metabolites conjoined with the death of living cells in the transition zone. While previous studies have characterized heartwood compounds and deposition, the cell death aspects are often neglected. Thus, the correlation of these defining cellular events remained unclear, while the exact timing of occurrence and regional involvement within the stem became topics for debate. In this study, members of metacaspases and vacuolar processing enzymes were isolated as programmed cell death (PCD) markers to explore their correlation with those of polyphenol biosynthesis. We collected heartwood-forming stems from R. pseudoacacia monthly for a year and monitored their expression through real-time PCR. Our results revealed novel markers of PCD in xylem parenchyma cells and demonstrated their corresponding fluctuation in expression with markers of polyphenols across the stem regions. The annual expression profile demonstrated regional stem development along with weather data on a monthly scale. This work provides a framework to compare the xylem-specific PCD in stages of stem development and a preliminary overview of how they correspond to external conditions over an annual cycle.