Evaluating the contribution of plant metabolic pathways in the light to the ATP:NADPH demand using a meta-analysis of isotopically non-stationary metabolic flux analyses.

Balancing the ATP: NADPH demand from plant metabolism with supply from photosynthesis is essential for preventing photodamage and operating efficiently, so understanding its drivers is important for integrating metabolism with the light reactions of photosynthesis and for bioengineering efforts that may radically change this demand. It is often assumed that the C3 cycle and photorespiration consume the largest amount of ATP and reductant in illuminated leaves and as a result mostly determine the ATP: NADPH demand. However, the quantitative extent to which other energy consuming metabolic processes contribute in large ways to overall ATP: NADPH demand remains unknown. Here, we used the metabolic flux networks of numerous recently published isotopically non-stationary metabolic flux analyses (INST-MFA) to evaluate flux through the C3 cycle, photorespiration, the oxidative pentose phosphate pathway, the tricarboxylic acid cycle, and starch/sucrose synthesis and characterize broad trends in the demand of energy across different pathways and compartments as well as in the overall ATP:NADPH demand. These data sets include a variety of species including Arabidopsis thaliana, Nicotiana tabacum, and Camelina sativa as well as varying environmental factors including high/low light, day length, and photorespiratory levels. Examining these datasets in aggregate reveals that ultimately the bulk of the energy flux occurred in the C3 cycle and photorespiration, however, the energy demand from these pathways did not determine the ATP: NADPH demand alone. Instead, a notable contribution was revealed from starch and sucrose synthesis which might counterbalance photorespiratory demand and result in fewer adjustments in mechanisms which balance the ATP deficit.

The role of photorespiration in preventing feedback regulation via ATP synthase in Nicotiana tabacum.

Photorespiration consumes substantial amounts of energy in the forms of adenosine triphosphate (ATP) and reductant making the pathway an important component in leaf energetics. Because of this high reductant demand, photorespiration is proposed to act as a photoprotective electron sink. However, photorespiration consumes more ATP relative to reductant than the C3 cycle meaning increased flux disproportionally increases ATP demand relative to reductant. Here we explore how energetic consumption from photorespiration impacts the flexibility of the light reactions in nicotiana tabacum. Specifically, we demonstrate that decreased photosynthetic efficiency (ϕII ) at low photorespiratory flux was related to feedback regulation at the chloroplast ATP synthase. Additionally, decreased ϕII at high photorespiratory flux resulted in the accumulation of photoinhibition at photosystem II centers. These results are contrary to the proposed role of photorespiration as a photoprotective electron sink. Instead, our results suggest a novel role of ATP consumption from photorespiration in maintaining ATP synthase activity, with implications for maintaining energy balance and preventing photodamage that will be critical for plant engineering strategies.

Composite modeling of leaf shape along shoots discriminates Vitis species better than individual leaves.

PREMISE: Leaf morphology is dynamic, continuously deforming during leaf expansion and among leaves within a shoot. Here, we measured the leaf morphology of more than 200 grapevines (Vitis spp.) over four years and modeled changes in leaf shape along the shoot to determine whether a composite leaf shape comprising all the leaves from a single shoot can better capture the variation and predict species identity compared with individual leaves. METHODS: Using homologous universal landmarks found in grapevine leaves, we modeled various morphological features as polynomial functions of leaf nodes. The resulting functions were used to reconstruct modeled leaf shapes across the shoots, generating composite leaves that comprehensively capture the spectrum of leaf morphologies present. RESULTS: We found that composite leaves are better predictors of species identity than individual leaves from the same plant. We were able to use composite leaves to predict the species identity of previously unassigned grapevines, which were verified with genotyping. DISCUSSION: Observations of individual leaf shape fail to capture the true diversity between species. Composite leaf shape-an assemblage of modeled leaf snapshots across the shoot-is a better representation of the dynamic and essential shapes of leaves, in addition to serving as a better predictor of species identity than individual leaves.

Transport of Anthocyanins and other Flavonoids by the Arabidopsis ATP-Binding Cassette Transporter AtABCC2.

Flavonoids have important developmental, physiological, and ecological roles in plants and are primarily stored in the large central vacuole. Here we show that both an ATP-binding cassette (ABC) transporter(s) and an H+-antiporter(s) are involved in the uptake of cyanidin 3-O-glucoside (C3G) by Arabidopsis vacuolar membrane-enriched vesicles. We also demonstrate that vesicles isolated from yeast expressing the ABC protein AtABCC2 are capable of MgATP-dependent uptake of C3G and other anthocyanins. The uptake of C3G by AtABCC2 depended on the co-transport of glutathione (GSH). C3G was not altered during transport and a GSH conjugate was not formed. Vesicles from yeast expressing AtABCC2 also transported flavone and flavonol glucosides. We performed ligand docking studies to a homology model of AtABCC2 and probed the putative binding sites of C3G and GSH through site-directed mutagenesis and functional studies. These studies identified residues important for substrate recognition and transport activity in AtABCC2, and suggest that C3G and GSH bind closely, mutually enhancing each other's binding. In conclusion, we suggest that AtABCC2 along with possibly other ABCC proteins are involved in the vacuolar transport of anthocyanins and other flavonoids in the vegetative tissue of Arabidopsis.

Midtreatment Problems Implementing Evidence-based Interventions in Community Settings.

Multisystemic Therapy® (MST) and Functional Family Therapy (FFT) are two widely disseminated evidence-based family-based treatments for substance abusing and delinquent adolescents. This mixed-method study examined common implementation problems in midtreatment in MST and FFT. A convenience sample of experienced therapists (20 MST, 20 FFT) and supervisors (10 MST, 10 FFT) from dissemination sites across the United States participated in semistructured telephone interviews. Participants identified retrospectively serious midtreatment process problems they perceived as threats to treatment success. Coders extracted descriptions of problems from interview transcripts and coded them into 12 categories that fell into five major themes: engaging families in treatment; difficulties implementing strategies; family relational and communication problems; complications external to therapy; and youth problem behavior. Analyses examined caregiver, therapist, and youth variables as predictors of these common midtreatment problems and whether treatment outcomes varied depending on the type of problem, therapy model, and race/ethnic match of therapist and family. MST and FFT therapists and supervisors identified many similar problems. There were, however, model-specific differences consistent with differing features of the models (e.g., FFT participants identified more family relational problems and fewer follow-through problems than their MST counterparts). Results underscore the need to consider both common and specific factors in treatment process.

La terapia multisistémica (Multisystemic Therapy®, MST) y la terapia familiar funcional (Functional Family Therapy, FFT) son dos tratamientos factuales familiares ampliamente difundidos para adolescentes con problemas de abuso de sustancias y delincuencia. Este estudio de métodos combinados analizó los problemas de implementación comunes a mediados del tratamiento en la MST y la FFT. Una muestra de conveniencia de terapeutas experimentados (20 MST, 20 FFT) y supervisores (10 MST, 10 FFT) de centros de difusión de todo EE. UU. participaron en entrevistas telefónicas semiestructuradas. Los participantes reconocieron retrospectivamente graves problemas en el proceso a mediados del tratamiento que percibieron como amenazas para el éxito del tratamiento. Los codificadores extrajeron las descripciones de problemas de las transcripciones de las entrevistas y los codificaron en 12 categorías que comprendían cinco temas principales: interés de las familias por el tratamiento; dificultades para implementar estrategias; problemas de comunicación y relacionales con la familia; complicaciones ajenas a la terapia; y comportamiento problemático de los jóvenes. Los análisis examinaron las variables de cuidador, terapeuta y joven como predictores de estos problemas comunes a mediados del tratamiento y si los resultados del tratamiento variaron según el tipo de problema, el modelo de terapia y la coincidencia étnica/racial entre el terapeuta y la familia. Los terapeutas y los supervisores de la MST y la FFT detectaron varios problemas similares. Sin embargo, hubo diferencias específicas de los modelos que coincidieron con las distintas características de los modelos (p. ej.: los participantes de la FFT reconocieron más problemas de relaciones familiares y menos problemas para terminar el tratamiento que sus homólogos de la MST). Los resultados subrayan las necesidad de tener en cuenta tanto los factores comunes como específicos del proceso de tratamiento.