Loss of fumarylacetoacetate hydrolase causes light‐dependent increases in protochlorophyllide and cell death in Arabidopsis

Abstract

Fumarylacetoacetate hydrolase (FAH) catalyses the final step of the tyrosine degradation pathway, which is essential to animals but was of unknown importance in plants until we found that mutation of Short-day Sensitive Cell Death1 (SSCD1), encoding Arabidopsis FAH, results in cell death under short-day conditions. The sscd1 mutant accumulates succinylacetone (SUAC), an abnormal metabolite caused by loss of FAH. Succinylacetone is an inhibitor of d-aminolevulinic acid (ALA) dehydratase (ALAD), which is involved in chlorophyll (Chl) biosynthesis. In this study, we investigated whether sscd1 cell death is mediated by Chl biosynthesis and found that ALAD activity is repressed in sscd1 and that protochlorophyllide (Pchlide), an intermediate of Chl biosynthesis, accumulates at lower levels in etiolated sscd1 seedlings. However, it was interesting that Pchlide in sscd1 might increase after transfer from light to dark and that HEMA1 and CHLH are upregulated in the light-dark transition before Pchlide levels increased. Upon re-illumination after Pchlide levels had increased, reactive oxygen species marker genes, including singlet oxygen-induced genes, are upregulated, and the sscd1 cell death phenotype appears. In addition, Arabidopsis WT seedlings treated with SUAC mimic sscd1 in decline of ALAD activity and accumulation of Pchlide as well as cell death. These results demonstrate that increase in Pchlide causes cell death in sscd1 upon re-illumination and suggest that a decline in the Pchlide pool due to inhibition of ALAD activity by SUAC impairs the repression of ALA synthesis from the light-dark transition by feedback control, resulting in activation of the Chl biosynthesis pathway and accumulation of Pchlide in the dark.

Cell death resulted from loss of fumarylacetoacetate hydrolase in Arabidopsis is related to phytohormone jasmonate but not salicylic acid

“…Previously, we found that ROS marker genes such as APX2 , OXI1 , BAP1 and ZP were up-regulated before an occurrence of cell death in the sscd1 mutant 48 , so, we next investigated whether the repression of cell death in sscd1 by mutation of COI1 is correlated with the expression of these genes. Since the cell death phenotype of sscd1 seedlings that were grown under SD appeared on the 6th day 41 , 49 , therefore, we tested the expression of APX2 , OXI1 , BAP1 and ZP in seedlings grown under SD for 5 days.…”

“…Singlet oxygen is very unstable and difficult to detect within a cell 55 , however, some genes were specifically induced by singlet oxygen 40 . Recently, we found that the genes induced specifically by singlet oxygen 40 were up-regulated in sscd1 48 , suggesting that an effect of SUAC on Chl biosynthesis results in the generation of singlet oxygen in the sscd1 mutant. Furthermore, we found that treatment of Arabidopsis wild-type seedlings with SUAC activated the expression of JA-inducible genes (Fig.…”

“…In plants, we have found that treatment of Arabidopsis wild-type seedlings with SUAC mimicked the sscd1 cell death phenotype 41 and demonstrated that the cell death of sscd1 seedlings correlates with the accumulation of SUAC 54 . Recently, we found that SUAC affects Chl biosynthesis, resulting in the generation of ROS and then inducing cell death 48 . Some researcher's work has shown that JA could be synthesized in response to singlet oxygen that is one form of ROS 25,55 .…”

“…Afterwards, we found that sugar suppresses cell death caused by disruption of FAH in Arabidopsis , indicating that Tyr degradation is regulated by sugar in plants 49 . Recently, we found that cell death resulted from loss of FAH in sscd1 is related to chlorophyll (Chl) biosynthesis, suggesting a crosstalk between Tyr degradation and Chl biosynthetic pathways in mediating the sscd1 cell death 48 . Phytohormones such as SA and JA are involved in PCD 13 – 16 , 24 , 25 , 50 .…”

“…RT-qPCR analysis were performed as described 48 . Total RNA was isolated using TRIZOL reagent (LIFE TECHNOLOGIES, https://www.thermofisher.com/us/en/home/brands/life-technologies.html ).…”