Peeling back the layers of crassulacean acid metabolism: functional differentiation between Kalanchoë fedtschenkoi epidermis and mesophyll proteomes

Abstract

Summary Crassulacean acid metabolism (CAM) is a specialized mode of photosynthesis that offers the potential to engineer improved water‐use efficiency (WUE) and drought resilience in C3 plants while sustaining productivity in the hotter and drier climates that are predicted for much of the world. CAM species show an inverted pattern of stomatal opening and closing across the diel cycle, which conserves water and provides a means of maintaining growth in hot, water‐limited environments. Recent genome sequencing of the constitutive model CAM species Kalanchoë fedtschenkoi provides a platform for elucidating the ensemble of proteins that link photosynthetic metabolism with stomatal movement, and that protect CAM plants from harsh environmental conditions. We describe a large‐scale proteomics analysis to characterize and compare proteins, as well as diel changes in their abundance in guard cell‐enriched epidermis and mesophyll cells from leaves of K. fedtschenkoi. Proteins implicated in processes that encompass respiration, the transport of water and CO2, stomatal regulation, and CAM biochemistry are highlighted and discussed. Diel rescheduling of guard cell starch turnover in K. fedtschenkoi compared with that observed in Arabidopsis is reported and tissue‐specific localization in the epidermis and mesophyll of isozymes implicated in starch and malate turnover are discussed in line with the contrasting roles for these metabolites within the CAM mesophyll and stomatal complex. These data reveal the proteins and the biological processes enriched in each layer and provide key information for studies aiming to adapt plants to hot and dry environments by modifying leaf physiology for improved plant sustainability.

How to resolve the enigma of diurnal malate remobilisation from the vacuole in plants with crassulacean acid metabolism?

“…These data indicate post-translational control over malate decarboxylation that is subject to metabolic regulation by CAM-associated metabolites, possibly malate. Diel changes in phosphopeptide abundance of PPDK protein in K. fedtschenkoi were reported by Abraham et al (2020) and are consistent with phosphorylation/inactivation of PPDK during the night in this CAM species. Phosphorylation of PPDK in C 4 plants is catalysed by PPDK regulatory protein (PPDK-RP; Chastain et al, 2011).…”

“…Diel changes in phosphopeptide abundance of PPDK protein in K. fedtschenkoi were reported by Abraham et al . (2020) and are consistent with phosphorylation/inactivation of PPDK during the night in this CAM species. Phosphorylation of PPDK in C 4 plants is catalysed by PPDK regulatory protein (PPDK‐RP; Chastain et al ., 2011).…”

“…Phosphorylation of PPDK in C 4 plants is catalysed by PPDK regulatory protein (PPDK‐RP; Chastain et al ., 2011). An orthologue of PPDK‐RP in K. fedtschenkoi showed increased protein abundance at the end of the photoperiod and for most of the dark period; this would be consistent with inactivation of PPDK when malate is accumulating in the vacuole, rather than being processed via decarboxylation (Abraham et al ., 2020). Moreover, in the inducible CAM plant M. crystallinum PPDK‐RP transcript abundance was upregulated when switching to CAM mode (Lim et al ., 2019).…”

“…Subsequent analyses of the transcriptome (Yang et al ., 2017) and proteome (Abraham et al ., 2020) of K. fedtschenkoi identified diel changes in transcript and protein abundance of kf NAD ME1. Protein abundance of kf NAD ME1 peaked in the middle of the photoperiod at a time when the rate of malate decarboxylation was maximal, suggesting a level of transcriptional regulation over the daytime breakdown of malate via ME (Abraham et al ., 2020). ME liberates CO 2 from malate to accommodate further refixation by RuBisCo in the mesophyll via the Calvin–Benson cycle (Fig.…”

“…Light intensity seems to be a critical factor in orchestrating diurnal malate metabolism and different potential rate-limiting steps have been suggested: (1) efflux from the vacuole; (2) liberation of CO 2 in the cytosol by decarboxylation; or (3) assimilation of the liberated CO 2 via RuBisCo in the chloroplast (L€ uttge, 2004). Driven by its growing eco-agricultural importance, the field of CAM research is now advancing quickly and important technical breakthroughs in the past decade have allowed an unprecedented exploration of CAM genomes, transcriptomes, proteomes and metabolomes (Ming et al, 2015;Yang et al, 2017;Ceusters et al, 2019;Abraham et al, 2020). In combination with rapidly progressing computational modelling approaches (Shameer et al, 2018;Chomthong & Griffiths, 2020) Factors influencing the light-dependent assimilation of CO 2 via RuBisCo and its relationship to the onset and subsequent rate of malate mobilisation during the photoperiod in CAM plants are still not fully resolved.…”