Dark respiration rates are not determined by differences in mitochondrial capacity, abundance and ultrastructure in C4 leaves

Our understanding of the regulation of respiration in C plants, where mitochondria play different roles in the different types of C photosynthetic pathway, remains limited. We examined how leaf dark respiration rates (R), in the presence and absence of added malate, vary in monocots representing the three classical biochemical types of C photosynthesis (NADP-ME, NAD-ME and PCK) using intact leaves and extracted bundle sheath strands. In particular, we explored to what extent R are associated with mitochondrial number, volume and ultrastructure. We found that the respiratory response of NAD-ME and PCK type bundle sheath strands to added malate was associated with differences in mitochondrial number, volume, and/or ultrastructure, while NADP-ME type bundle sheath strands did not respond to malate addition. In general, mitochondrial traits reflected the contributions mitochondria make to photosynthesis in the three C types. However, despite the obvious differences in mitochondrial traits, no clear correlation was observed between these traits and R. We suggest that R is primarily driven by cellular maintenance demands and not mitochondrial composition per se, in a manner that is somewhat independent of mitochondrial organic acid cycling in the light.

Role of hydrogen sulfide in the regulation of respiration, blood flow and bile secretory function of the liver

In acute experiments on laboratory rats, intra-portal administration of L-cysteine (20 mg/kg), the precursor of hydrogen sulfide synthesis, stimulated oxygen consumption of liver by 38.6% and reduced oxygen tension by 37.1%. Activation of tissue respiration occurred due to the strengthening of oxygen-dependent synthetic processes in liver, in particular those associated with mitochondrial enzyme-catalysed bile acid biosynthesis through the acidic pathway. The concentrations of taurocholic acid and mixtures of taurodeoxycholic and taurohenodeoxycholic acids increased by 10.3 and 17.9%, respectively, compared to the initial levels. In addition, the level of free cholesterol was decreased by 33.9% and esterification processes were intensified, as indicated by an increase in the concentration of esterified cholesterol by 22.6% in the bile of rats. The latter was to some extent confirmed by a decrease in the level of free bile acids (by 15.8%) involved in the biosynthesis of cholesterol esters and intensification of tissue respiration in the liver. L-cysteine dilated intrahepatic vessels, resulting in a significant decrease of the systemic blood pressure and blood pressure in the portal vein by 17.6 and 24.5%, respectively. L-cysteine increased the rate of local blood flow in the liver and blood supply by 28.2 and 24.4%, respectively. Blockade of cystathionine-γ-lyase by DL-propargylglycine (11 mg/kg) significantly inhibited the L-cysteine-induced tissue respiration and bile acid biosynthesis in the liver. Administration of DL-propargylglycine resulted in constriction of blood vessels of the liver and, as a consequence, to an increased blood pressure and a decreased blood flow rate in tissue. Our data point to an involvement of hydrogen sulfide in the regulation of liver tissue respiration and bile secretory function.

Unveiling a key role of oxaloacetate-glutamate interaction in regulation of respiration and ROS generation in nonsynaptic brain mitochondria using a kinetic model

Glutamate plays diverse roles in neuronal cells, affecting cell energetics and reactive oxygen species (ROS) generation. These roles are especially vital for neuronal cells, which deal with high amounts of glutamate as a neurotransmitter. Our analysis explored neuronal glutamate implication in cellular energy metabolism and ROS generation, using a kinetic model that simulates electron transport details in respiratory complexes, linked ROS generation and metabolic reactions. The analysis focused on the fact that glutamate attenuates complex II inhibition by oxaloacetate, stimulating the latter’s transformation into aspartate. Such a mechanism of complex II activation by glutamate could cause almost complete reduction of ubiquinone and deficiency of oxidized form (Q), which closes the main stream of electron transport and opens a way to massive ROS generating transfer in complex III from semiquinone radicals to molecular oxygen. In this way, under low workload, glutamate triggers the respiratory chain (RC) into a different steady state characterized by high ROS generation rate. The observed stepwise dependence of ROS generation on glutamate concentration experimentally validated this prediction. However, glutamate’s attenuation of oxaloacetate’s inhibition accelerates electron transport under high workload. Glutamate-oxaloacetate interaction in complex II regulation underlies the observed effects of uncouplers and inhibitors and acceleration of Ca2+ uptake. Thus, this theoretical analysis uncovered the previously unknown roles of oxaloacetate as a regulator of ROS generation and glutamate as a modifier of this regulation. The model predicted that this mechanism of complex II activation by glutamate might be operative in situ and responsible for excitotoxicity. Spatial-time gradients of synthesized hydrogen peroxide concentration, calculated in the reaction-diffusion model with convection under a non-uniform local approximation of nervous tissue, have shown that overproduction of H2O2 in a cell causes excess of its level in neighbor cells.

Wt1 Positive dB4 Neurons in the Hindbrain Are Crucial for Respiration

Central pattern generator (CPG) networks coordinate the generation of rhythmic activity such as locomotion and respiration. Their development is driven by various transcription factors, one of which is the Wilms tumor protein (Wt1). It is present in dI6 neurons of the mouse spinal cord, and involved in the coordination of locomotion. Here we report about the presence of Wt1 in neurons of the caudoventral medulla oblongata and their impact on respiration. By employing immunohistofluorescence staining, we were able to characterize these Wt1 positive (+) cells as dB4 neurons. The temporal occurrence of Wt1 suggests a role for this transcription factor in the differentiation of dB4 neurons during embryonic and postnatal development. Conditional knockout of Wt1 in these cells caused an altered population size of V0 neurons already in the developing hindbrain, leading to a decline in the respiration rate in the adults. Thereby, we confirmed and extended the previously proposed similarity between dB4 neurons in the hindbrain and dI6 neurons of the spinal cord, in terms of development and function. Ablation of Wt1+ dB4 neurons resulted in the death of neonates due to the inability to initiate respiration, suggesting a vital role for Wt1+ dB4 neurons in breathing. These results expand the role of Wt1 in the CNS and show that, in addition to its function in differentiation of dI6 neurons, it also contributes to the development of dB4 neurons in the hindbrain that are critically involved in the regulation of respiration.

Shifting paradigms and novel players in Cys-based redox regulation and ROS signaling in plants - and where to go next

Cys-based redox regulation was long regarded a major adjustment mechanism of photosynthesis and metabolism in plants, but in the recent years, its scope has broadened to most fundamental processes of plant life. Drivers of the recent surge in new insights into plant redox regulation have been the availability of the genome-scale information combined with technological advances such as quantitative redox proteomics and in vivo biosensing. Several unexpected findings have started to shift paradigms of redox regulation. Here, we elaborate on a selection of recent advancements, and pinpoint emerging areas and questions of redox biology in plants. We highlight the significance of (1) proactive H2O2 generation, (2) the chloroplast as a unique redox site, (3) specificity in thioredoxin complexity, (4) how to oxidize redox switches, (5) governance principles of the redox network, (6) glutathione peroxidase-like proteins, (7) ferroptosis, (8) oxidative protein folding in the ER for phytohormonal regulation, (9) the apoplast as an unchartered redox frontier, (10) redox regulation of respiration, (11) redox transitions in seed germination and (12) the mitochondria as potential new players in reductive stress safeguarding. Our emerging understanding in plants may serve as a blueprint to scrutinize principles of reactive oxygen and Cys-based redox regulation across organisms.

The RNA helicase DDX5 supports mitochondrial function in small cell lung cancer

DEAD-box helicase 5 (DDX5) is a founding member of the DEAD-box RNA helicase family, a group of enzymes that regulate ribonucleoprotein formation and function in every aspect of RNA metabolism, ranging from synthesis to decay. Our laboratory previously found that DDX5 is involved in energy homeostasis, a process that is altered in many cancers. Small cell lung cancer (SCLC) is an understudied cancer type for which effective treatments are currently unavailable. Using an array of methods, including short hairpin RNA–mediated gene silencing, RNA and ChIP sequencing analyses, and metabolite profiling, we show here that DDX5 is overexpressed in SCLC cell lines and that its down-regulation results in various metabolic and cellular alterations. Depletion of DDX5 resulted in reduced growth and mitochondrial dysfunction in the chemoresistant SCLC cell line H69AR. The latter was evidenced by down-regulation of genes involved in oxidative phosphorylation and by impaired oxygen consumption. Interestingly, DDX5 depletion specifically reduced intracellular succinate, a TCA cycle intermediate that serves as a direct electron donor to mitochondrial complex II. We propose that the oncogenic role of DDX5, at least in part, manifests as up-regulation of respiration supporting the energy demands of cancer cells.

Wt1 positive neurons in the hindbrain are essential for respiration

Neuronal networks commonly referred to as central pattern generator (CPG) networks coordinate the generation of rhythmic activity like locomotion and respiration. These networks are proposed to exhibit a high degree of homology in their development. Their establishment is influenced by a variety of transcription factors. One of them is the Wilms tumor protein Wt1 that is present in dI6 neurons of the ventral spinal cord, which are involved in the coordination of locomotion. Here we report about the so far undescribed presence of Wt1 in neurons of the caudoventral medulla oblongata and their impact on respiration. By performing marker analyses, we were able to characterize these Wt1 positive (+) cells as dB4 neurons. The temporal pattern of Wt1 occurrence suggests a role for Wt1 in the differentiation of dB4 neurons during embryonic and postnatal development. Conditional knockout of Wt1 in these cells caused an altered population size of V0 neurons already in the developing hindbrain leading to a decline in the respiration rate in the adults. Thereby, we confirmed and extended the so far proposed homology between neurons of the dB4 domain in the hindbrain and dI6 neurons of the spinal cord in terms of development and function. Ablation of Wt1+ dB4 neurons resulted in the death of neonates due to the inability to initiate respiration suggesting a vital role for Wt1+ dB4 neurons in breathing. These results extend the role of Wt1 in the CNS and show that in addition to its function in differentiation of dI6 neurons it also contributes to the development of dB4 neurons in the hindbrain that are critically involved in the regulation of respiration.