NOL‐mediated functional stay‐green traits in perennial ryegrass (Lolium perenne L.) involving multifaceted molecular factors and metabolic pathways regulating leaf senescence

Abstract

SUMMARY Loss of chlorophyll (Chl) is a hallmark of leaf senescence, which may be regulated by Chl catabolic genes, including NON‐YELLOW COLORING 1 (NYC1)‐like (NOL). The objective of this study was to determine molecular factors and metabolic pathways underlying NOL regulation of leaf senescence in perennial grass species. LpNOL was cloned from perennial ryegrass (Lolium perenne L.) and found to be highly expressed in senescent leaves. Transient overexpression of LpNOL accelerated leaf senescence and Chl b degradation in Nicotiana benthamiana. LpNOL RNA interference (NOLi) in perennial ryegrass not only significantly blocked Chl degradation in senescent leaves, but also delayed initiation and progression of leaf senescence. This study found that NOL, in addition to functioning as a Chl b reductase, could enact the functional stay‐green phenotype in perennial grass species, as manifested by increased photosynthetic activities in NOLi plants. Comparative transcriptomic analysis revealed that NOL‐mediated functional stay‐green in perennial ryegrass was mainly achieved through the modulation of Chl catabolism, light harvesting for photosynthesis, photorespiration, cytochrome respiration, carbohydrate catabolism, oxidative detoxification, and abscisic acid biosynthesis and signaling pathways.

Functional Characterization of the Pheophytinase Gene, ZjPPH, From Zoysia japonica in Regulating Chlorophyll Degradation and Photosynthesis

“…Specifically, the identification of important stay-green mutants unmasked the key catalyzing steps in the chlorophyll breakdown pathway. In brief, the breakdown of chlorophyll was started with the conversion of chlorophyll b (Chl b ) to Chl a catalyzed by two Chl b reductase paralogous, NYC1 and NOL ( Yu et al, 2021 ). Next, magnesium was released from Chl a by the STAY-GREEN protein (SGR) to yield pheophytin a ( Shimoda et al, 2016 ).…”

LpNOL‐knockdown suppression of heat‐induced leaf senescence in perennial ryegrass involving regulation of amino acid and organic acid metabolism

“…The molecular mechanisms of LpNOL regulating leaf senescence and controlling the functional stay-green phenotype in perennial ryegrass were reported in a previous study (Yu et al, 2021). While the mechanisms by which NOL regulates leaf senescence are yet not completely understood, NOL-mediated leaf senescence has been associated with positive regulation of Chl b degradation due to its role as Chl b reductase (Sato et al, 2010;Yu et al, 2021). In this study, metabolic profiling of perennial ryegrass with LpNOL knockdown found that five metabolites (valine, malic acid, threonic acid, shikimic acid, and chlorogenic acid) were uniquely upregulated in LpNOL-knockdown plants exposed to heat stress, and six metabolites (aspartic acid, glutamic acid, 5-oxoproline, senescence could be related to the activation of amino acid and organic acid metabolism (Figure 9) in addition to its previously reported function in the catalytic breakdown of Chl degradation (Yu et al, 2021).…”

“…The enhancement of Chl content, F v /F m , Pn, and cell membrane stability suggest that knockdown of LpNOL suppressed heat-induced leaf senescence. The molecular mechanisms of LpNOL regulating leaf senescence and controlling the functional stay-green phenotype in perennial ryegrass were reported in a previous study (Yu et al, 2021). While the mechanisms by which NOL regulates leaf senescence are yet not completely understood, NOL-mediated leaf senescence has been associated with positive regulation of Chl b degradation due to its role as Chl b reductase (Sato et al, 2010;Yu et al, 2021).…”

“…While the mechanisms by which NOL regulates leaf senescence are yet not completely understood, NOL-mediated leaf senescence has been associated with positive regulation of Chl b degradation due to its role as Chl b reductase (Sato et al, 2010;Yu et al, 2021). In this study, metabolic profiling of perennial ryegrass with LpNOL knockdown found that five metabolites (valine, malic acid, threonic acid, shikimic acid, and chlorogenic acid) were uniquely upregulated in LpNOL-knockdown plants exposed to heat stress, and six metabolites (aspartic acid, glutamic acid, 5-oxoproline, senescence could be related to the activation of amino acid and organic acid metabolism (Figure 9) in addition to its previously reported function in the catalytic breakdown of Chl degradation (Yu et al, 2021). For an orderly discussion of metabolic regulation of LpNOL, relating to heat-induced leaf senescence, those metabolites (amino acids and organic acids) with increased abundance in LpNOL, knockdown lines exposed to heat stress and the associated metabolic pathways (respiration, secondary metabolism, antioxidant metabolism, and protein synthesis) are illustrated in Figure 9 and discussed below.…”

Overexpression of 7-hydroxymethyl Chlorophyll a Reductase from Cucumber in Tobacco Accelerates Dark-Induced Chlorophyll Degradation

“…Then, 7-HMChl a is transferred to Chl a by 7-hydroxymethyl Chl a reductase (HCAR) [ 14 ]. However, the Chl cycle is more than a reciprocal conversion of Chl a and Chl b ; it also plays an important role in the degradation of light-harvesting Chl a / b protein complex of photosystem II (LHCII) [ 15 , 16 ]. Arabidopsis SGR1 physically interacts with the Chl catabolic enzymes (CCEs) and LHCII, forming a multi-protein complex that is likely important for rapid detoxification of Chl catabolic intermediates in senescing chloroplasts [ 17 , 18 ].…”