Transcriptomic and Biochemical Analysis Reveal Integrative Pathways Between Carbon and Nitrogen Metabolism in Guzmania monostachia (Bromeliaceae) Under Drought

Abstract

Most epiphytes are found in low-nutrient environments with an intermittent water supply. To deal with water limitation, many bromeliads perform crassulacean acid metabolism (CAM), such as Guzmania monostachia, which shifts from C3 to CAM and can recycle CO2 from the respiration while stomata remain closed during daytime and nighttime (CAM-idling mode). Since the absorbing leaf trichomes can be in contact with organic (urea) and inorganic nutrients (NO3−, NH4+) and the urea hydrolysis releases NH4+ and CO2, we hypothesized that urea can integrate the N and C metabolism during periods of severe drought. Under this condition, NH4+ can be assimilated into amino acids through glutamine synthetase (GS), while the CO2 can be pre-fixated by phosphoenolpyruvate carboxylase (PEPC). In this context, we evaluated the foliar transcriptome of G. monostachia to compare the relative gene expression of some genes involved with CAM and the N metabolism when bromeliads were submitted to 7days of drought. We also conducted a controlled experiment with an extended water deficit period (21days) in which bromeliads were cultivated in different N sources (urea, NH4+, and NO3−). Our transcriptome results demonstrated an increment in the expression of genes related to CAM, particularly those involved in the carboxylation metabolism (PEPC1, PPCK, and NAD-MDH), the movement of malate through vacuolar membrane (ALMT9), and the decarboxylation process (PEPCK). Urea stimulated the expression of PEPC1 and ALMT9, while Urease transcripts increased under water deficit. Under this same condition, GS1 gene expression increased, indicating that the NH4+ from urea hydrolysis can be assimilated in the cytosol. We suggest that the link between C and N metabolism occurred through the supply of carbon skeleton (2-oxoglutarate, 2-OG) by the cytosolic isocitrate dehydrogenase since the number of NADP-ICDH transcripts was also higher under drought conditions. These findings indicate that while urea hydrolysis provides NH4+ that can be consumed by glutamine synthetase-cytosolic/glutamate synthase (GS1/GOGAT) cycle, the CO2 can be used by CAM, maintaining photosynthetic efficiency even when most stomata remain closed 24h (CAM-idling) as in the case of a severe water deficit condition. Thus, we suggest that urea could be used by G. monostachia as a strategy to increase its survival under drought, integrating N and C metabolism.

Acid waters in tank bromeliads: Causes and potential consequences

“…Water may be the chief limiting factor for tropical epiphytes such as bromeliads (Zotz and Bader, 2009; Males, 2016), particularly as episodes of drought increase in frequency and duration due to global warming (Colwell et al, 2008; Corlett, 2016), but the relative scarcity of mineral nutrients in the canopy also requires special adaptations (Zotz, 2016). For example, aquaporins in the leaves of the bromeliads Vriesea gigantea and G. monostachia can take up urea and other organic forms of nitrogen from the tank (Matiz et al, 2019; Gonçalves and Mercier, 2021), and several species have been shown to obtain mineral N through their association with decomposing microbes and N‐fixing cyanobacteria (Bermudes and Benzing, 1991; Inselsbacher et al, 2007). Phosphorus may be even more limiting than N for tank bromeliads, which have numerous biochemical adaptations to improve P‐uptake and storage (Winkler and Zotz, 2009).…”

“…Perhaps the best‐studied adaptation to a limited and variable water supply is the wide range of water‐conserving CAM photosynthetic pathways shown in the group, from full CAM to weak CAM, including the C 3 –CAM intermediacy of the focal species in this study, G. monostachia (Smith et al, 1986 ; Maxwell et al, 1994 ; Crayn et al, 2015 ; Pikart et al, 2020 ). Adaptations to limited mineral nutrients include high‐affinity membrane transporters for nitrogen (Inselsbacher et al, 2007 ), phosphorus (Winkler and Zotz, 2009 ), and potassium (Winkler and Zotz, 2010 ), and the capacity to acquire organic sources of N such as urea (Matiz et al, 2019 ; Gonçalves and Mercier, 2021 ). The most fundamental adaptation to limitation of both water and nutrients may be the slow growth rate of most epiphytes, including tank bromeliads (Schmidt and Zotz, 2002 ).…”

“…We recognize that our focus on pH and water uptake sidesteps the rich and extensive literature on mineral nutrition in tank bromeliads (Pittendrigh, 1948 ; Benzing and Renfrow, 1974 ; Givnish et al, 2014 ) and that adaptations to water and nutrient limitation tend to occur together (Gonçalves and Mercier, 2021 ). In view of our results and in the more general context of resource limitation, we briefly discuss possible implications—for epiphytes as well as other species with foliar uptake of water and nutrients (Schreel and Steppe, 2020 )—of low pH for resource acquisition by tank bromeliads.…”

Defining Mechanisms of C3 to CAM Photosynthesis Transition toward Enhancing Crop Stress Resilience

“…In the leaves of another facultative bromeliad Guzmania monostachia, increases in the expression of CAM-related genes (PEPC1, PPCK, NAD-malate dehydrogenase, aluminum-activated malate transporter 9 (ALMT9), PEP carboxykinase (PEPCK)) and UREASE transcripts were shown under drought. And the role of integrating N and C metabolism of urea was suggested [78]. The CAM gene expression, antioxidant activities, and chlorophyll fluorescence were compared between a C 3 -CAM facultative species (Sedum album) and a C 4 -CAM facultative species (Portulaca oleracea) [79].…”