Carbon dioxide detection in biological systems

CO₂ is produced during aerobic respiration. Normally, levels of CO₂ in the blood are tightly regulated but pCO₂ can rise (hypercapnia, pCO₂>45mmHg) in patients with lung diseases e.g. COPD. Hypercapnia is a risk factor in COPD but may be of benefit in the context of destructive inflammation through suppression of pro-inflammatory signalling.

Recently, we used an RNA-seq approach to determine the transcriptomic response of monocytes to hypercapnia (10% CO₂, 4hrs) under pH -buffered conditions. We identified a cluster of 254 genes to be CO₂ responsive under basal conditions and in the presence of a pro-inflammatory stimulus (LPS). Interestingly, genes encoding for mitochondrial proteins were highly represented in this cluster and nearly all mitochondrial encoded genes e.g. MT-CO1, MT-ND4 were enhanced under conditions of buffered hypercapnia. Mitochondrial DNA content was not enhanced, but acylcarnitine species and genes associated with fatty acid metabolism e.g. ACADSs were increased in hypercapnia. Primary IL-4 polarised macrophages exposed to buffered hypercapnia also increased expression of genes associated with fatty acid metabolism eg CPT1a and reduced expression of genes associated with glycolysis e.g. ALDOA and inflammation TNF.   Taken together these data suggest that buffered hypercapnia is a robust metabolic stimulus that influences the inflammatory status of immune cells. Current work is directed towards dissecting the contribution of hypercapnia to immunometabolism in immune cells using integrated analysis of RNA-seq, LC-MS metabolomics as well as GC-MS stable isotope tracing data. These studies are important for understanding immune dysregulation in patients experiencing hypercapnia.

Dr Eoin Cummins, University College Dublin, Ireland

Dr Eoin Cummins, University College Dublin, Ireland

Dr Cummins holds a BSc in Pharmacology and a PhD in molecular medicine. He is currently an Associate Professor in Physiology in the School of Medicine, University College Dublin.

The Cummins lab focuses on the role of physiological gases eg Oxygen (O₂) and carbon dioxide (CO₂), in the context of health and disease. It is well known that areas of hypoxia (low O₂) exist in regions of altered metabolic activity, ischemia, inflammation and cancer. Similarly, in areas with depleted oxygen, cellular and tissue carbon dioxide levels can rise, resulting in hypercapnia. The concentrations of these physiological gases are known to impact on cellular signaling and modify proliferative, healing and immune/ inflammatory responses. Current work in the Cummins lab is focused on understanding the contribution of carbon dioxide to metabolic signalling in immune cells and skeletal muscle.

Approximately one-third of global carbon-fixation occurs in an overlooked algal organelle called the pyrenoid. The pyrenoid contains the CO₂-fixing enzyme Rubisco and enhances carbon-fixation by supplying Rubisco with a high concentration of CO₂. Recently we have gained novel insights into the molecular structure and biogenesis of this ecologically fundamental organelle by determining the spatial organization of pyrenoid proteins and showing that the pyrenoid is a liquid-liquid phase separated system. To expand our knowledge outside of a model alga we are now systematically characterizing pyrenoids across the eukaryotic tree of life including green algae, red algae, diatoms and macroalgae. Our data is providing insights into the convergent evolution of CO₂-fixing biomolecular condensates and guiding the engineering of a pyrenoid into higher plants with a goal to enhance crop carbon fixation efficiency.

Professor Luke Mackinder, University of York, UK

Professor Luke Mackinder, University of York, UK

Luke is a UKRI Future Leader Fellow and a 2018 SEB Presidents Medallist for Cell Biology. His lab is applying high-throughput, systems and synthetic biology in diverse algae to rapidly dissect, predict and build CO₂ fixing pathways. This data is being used to generate a blueprint for the engineering of efficient CO₂ uptake systems in plants, with the goal of improving photosynthesis and ultimately crop yields and biological based carbon capture systems. Luke did a Marie Curie Funded PhD at the GEOMAR-Helmholtz Centre for Ocean Research Kiel, Germany and the Marine Biological Association of the UK. Followed by a 4-year postdoc at the Carnegie Institute for Plant Sciences, Stanford, before starting his own group at York in 2016. The Mackinder Lab is currently funded by the BBSRC, a BBSRC/NSF-Bio partnership award, EPSRC and the Bill and Melinda Gates Foundation.

Carbon dioxide (CO₂) is a widespread biological gas that triggers adaptive responses within organisms from all domains of life. However, the basic biochemical mechanisms through which organisms’ sense and respond to CO₂ are poorly understood. One direct way that CO₂ can modulate biological systems is by direct modification of free amine groups within proteins, forming a carbamate (Prot-CO₂). This modification can serve as a dynamic switch to alter the structure and chemical properties at these protein sites. However, Prot-CO₂ is notoriously challenging to detect on proteins using traditional analytical methods. Therefore, this modification has only been well-characterized on a handful of proteins and its diverse roles in biology are largely unknown. Herein, we describe a robust chemical proteomic method to quantitatively detect Prot-CO₂ sites in proteins. Using this method, we identify several high-confidence sites within different proteomes and explore the biochemical regulation of high-interest sites. This proof-of-concept work sets the stage for exploring the role of this dynamic modification as a biochemical regulator of CO₂ sensing in diverse organisms.

Dr Dustin King, Simon Fraser University, Canada

Dr Dustin King, Simon Fraser University, Canada

Dr Dustin King completed his CIHR-funded PhD in 2016 in the lab of renowned structural biologist Professor Natalie Strynadka at University of British Columbia, studying the biochemical basis of antibiotic resistance mechanisms and earning the Governor General's Gold Medal for the top PhD thesis. He then conducted postdoctoral research at SFU in Professor David Vocadlo’s lab, where he pioneered methods to study labile post-translational modifications. His work, supported by CIHR and MSFHR/CLEAR awards, included developing a method to identify CO₂-mediated carboxylation sites in proteins, enhancing understanding of CO₂-sensing mechanisms. In July 2022, he became an assistant professor in Simon Fraser University's MBB department, focusing on how cells respond to reactive metabolites. These metabolites form unique covalent bonds with protein residues, acting as chemical switches to regulate protein function. His research group develops innovative chemical proteomics methods to uncover these sites and characterise their biochemical and cellular mechanisms.

Over the last 10 years a number of studies have provided evidence demonstrating that improving photosynthesis can result in improved yield. This talk will focus on improvements in the RuBP regeneration phase of the Calvin-Benson-Bassham cycle and also electron transport. I will summarise some of the results of successful studies using genetic engineering of steps in the photosynthetic process that led to increases in yield. These results provide clear evidence for the potential of increased photosynthesis to contribute to improving our crop plants.

Professor Christine Raines, University of Essex, UK

Professor Christine Raines, University of Essex, UK

Christine Raines graduated with a BSc (Hons) Agricultural Botany in 1982 and a PhD in Photosynthetic electron transport graduating in1986 form Glasgow University. Christine’s post-doctoral research started in late 1985 at the Institute of Plant Science Research, Cambridge, working on the molecular biology of C3 cycle. In 1988 Christine was appointed to a faculty position at the University of Essex being promoted to Professor in 2004. Christine was Head of the School of Life Sciences at Essex (2011-2017), Pro-Vice Chancellor Research (2017-2021) for the University of Essex and she held a number of external roles; Editor in Chief, Journal of Experimental Botany (2011- ), Chair of Plant Section, Society of Experimental Biology (2009- ) and SEB President (2017-2019).  

Christine’s research interests are in plant molecular physiology analysis of gene expression and production and analysis of transgenic plants. Currently Christine leads a research group focussed on improving photosynthesis by re-engineering the CO₂ assimilatory pathway (the Calvin Benson cycle) and electron transport.

Exerting control over fertility remains a global challenge. Indeed, nearly half of all pregnancies are unintended. For men, the only available choices are condoms and vasectomy. While there are numerous efforts to develop new contraceptives for men, most impact sperm development. We are developing a novel, on-demand approach to male contraception where a man takes a birth control pill right before sex, only when necessary. We identified Soluble adenylyl cyclase (sAC; ADCY10) is directly regulated by bicarbonate and functions as a CO₂/HCO₃-/pHi sensor. In mammals, sAC is most abundantly expressed in sperm, and it has been genetically confirmed as essential for male fertility in mice and humans. Specifically, we and others have shown that AC is necessary for sperm maturation, motility, and capacitation. We are developing a battery of inhibitors specifically targeting sAC. We demonstrated that a single dose of a safe, acutely-acting sAC inhibitor with long residence time renders male mice temporarily infertile. Overall, our studies identify sAC inhibitors as potential candidates for on-demand male contraceptives, offering in vivo proof-of-concept for previously unexplored paradigms in contraception, including single-dose on-demand contraception for males.

Dr Justine Fischoeder, Weill Cornell Medical College, USA

Dr Justine Fischoeder, Weill Cornell Medical College, USA

Dr Justine Fischoeder is a postdoctoral researcher at the Weill Cornell Medical College in New York City. She got her PhD in biology from RWTH Aachen University in Germany, where she was working on cell communication during spermatogenesis. She decided to pursue her career in reproductive sciences, working on sperm motility in the lab of Drs Lonny Levin and Jochen Buck in the Department of Pharmacology at Cornell, whom she recently joined. Drs Levin and Buck first collaborated to purify and clone the bicarbonate-regulated soluble adenylyl cyclase (sAC).  They eventually merged into a single laboratory studying sAC and other bicarbonate-regulated enzymes. The LevBuck lab is currently developing a male contraceptive by inhibiting sAC in sperm cells.

To assess the impact of climate change on wheat yield, it is important to reliably predict its photosynthetic acclimation to a combination of [CO₂] and temperature changes. This study aimed to assess which photosynthetic parameters were most influenced by elevated [CO₂] and growth temperature. To unravel acclimation of photosynthetic temperature response (An-T) and whether its parameters were dependent on growth [CO₂], growth temperature or leaf rank, plants were grown under a range of temperatures (18 to 37 °C), two [CO₂] (400 and 800 µbar) and assessed at two leaf ranks (leaf 4 and 6). Photosynthetic parameters most influenced by acclimation changes in An-T response and optimum temperature (Topt) were the activation energies of maximum Rubisco carboxylation (EVcmax) and of triose phosphate utilization (ETp), which increased 1.157 and 1.859 kJ mol-1, respectively, per °C increase in growth temperature. Changes depended on the most recently experienced growth temperature, regardless of [CO₂] or growth temperatures during earlier growth. Under increasing [CO₂] levels and growth temperature, as predicted in future climates, photosynthetic acclimation will be controlled by growth temperature and at higher [CO₂] limitation of An set by triose phosphate utilization may occur more often in wheat.

Dr Steven M Driever, Wageningen University, The Netherlands

Dr Steven M Driever, Wageningen University, The Netherlands

Steven Driever is an Assistant Professor in Crop Physiology at the Centre for Crop Systems Analysis at Wageningen University, the Netherlands. He obtained an MSc in Biology from Wageningen University and a PhD in Plant Physiology from the University of Essex, UK, where he specialised in photosynthesis. He held several post-doctoral positions at the University of Essex, Wageningen University and the University of Illinois, before he was appointed as an Assistant Professor at Wageningen University in 2019, with tenure from 2023. He has initiated a new joint photosynthesis laboratory where he has developed a range of new methods. In his group, experimentation and modelling are combined, to understand how photosynthesis interacts with its environment, from the chloroplast to the canopy level.

Carbon dioxide (CO₂), a primary product of oxidative metabolism, can be sensed by eukaryotic cells eliciting specific responses via specific signalling pathways. In humans, hypercapnia - defined as the elevation of CO₂ in the bloodstream and tissues - occurs in severe lung diseases. Hypercapnia is known to be associated with adverse outcomes in patients with chronic obstructive pulmonary disease (COPD). COPD is a progressive pulmonary disorder caused by structural remodelling of small peripheral airways which affects airway resistance and hyperreactivity. Dr Shigemura’s group has reported that elevated CO₂ acts as a signalling molecule that promotes airway hyperreactivity and constriction, and that hypercapnic patients had higher airway resistance in COPD. In recent work, the group found that high CO₂ induces hypertrophic changes in lung smooth muscle cells and activates fibroblasts to produce extracellular matrix proteins, which results in structural remodelling of the lung with smooth muscle hypertrophy and extracellular matrix deposition around the peri-bronchial and peri-vascular area. These data suggest that CO₂ accumulates in the shared interstitial space between the airways and the pulmonary vasculature (broncho-vascular bundle) during respiratory failure. Dr Shigemura will provide new evidence that hypercapnia causes adverse structural remodelling of the lung and may drive COPD pathogenesis and progression.

Dr Masahiko Shigemura, Northwestern University, USA

Dr Masahiko Shigemura, Northwestern University, USA

Masahiko Shigemura is a Research Assistant Professor of Surgery at Northwestern University (USA). He obtained his PhD in Medicine from Hokkaido University (Japan) and subsequently carried out post-doctoral research in the laboratory of Dr Jacob I Sznajder at Northwestern University.

His primary research focuses on airway smooth muscle contractility during hypercapnia, a topic with potential clinical relevance for patients with obstructive lung diseases such as chronic obstructive pulmonary disease (COPD). As a Research Assistant Professor, he has been expanding his scientific expertise and conducting research to address airway pathobiology in the context of hypercapnia and COPD.

Rubisco is responsible for the majority of inorganic carbon assimilation on Earth. To ensure efficient CO₂-fixation Cyanobacteria and many autotrophic Proteobacteria concentrate CO₂ in proteinaceous organelles called carboxysomes. This organelle encapsulates key enzymes for CO₂ fixation, Rubisco and carbonic anhydrase, and are the centrepiece of the bacterial CO₂ concentrating mechanism (CCM). In the CCM, actively accumulated bicarbonate diffuses into the carboxysome and is converted to CO₂ by carbonic anhydrase. This produces a high CO₂ concentration near Rubisco ensuring efficient carboxylation. It remains unknown exactly how this 250+ megadalton protein complex assembles with high fidelity inside cells. The self-assembly of the α-carboxysome is orchestrated by the intrinsically disordered scaffolding protein, CsoS2, which interacts with both Rubisco and carboxysomal shell proteins. But, it’s been unknown how CsoSCA, the carbonic anhydrase, is incorporated into the α-carboxysome.

Here, I will discuss our recent work on the structural basis of carbonic anhydrase encapsulation into α-carboxysomes from Halothiobacillus neapolitanus. We find that CsoSCA interacts directly with Rubisco via an intrinsically disordered N-terminal domain. A sub 2 Å single-particle cryo-EM structure of Rubisco in complex with this peptide reveals that CsoSCA binding is predominantly mediated by a network of hydrogen bonds. CsoSCA's binding site overlaps with that of CsoS2 but the two proteins utilise substantially different motifs and modes of binding, revealing a plasticity of the Rubisco binding site. Our results update the current model for carboxysome biogenesis and inform strategies for engineering CO₂ concentration mechanisms into crops and industrially relevant microorganisms for improved growth and yields.

Dr Cecilia Blikstad, Uppsala University, Sweden

Dr Cecilia Blikstad, Uppsala University, Sweden

Cecilia (Cissi) obtained her PhD in Biochemistry from Uppsala University working on mechanistic enzymology and directed evolution of alcohol dehydrogenases. She thereafter moved to University of California Berkeley for a postdoc. Here she studied biochemical aspects of bacterial carbon fixation in David Savage lab. She specifically focused on the carboxysome, a specialised organelle which encapsulates the key enzymes for CO₂-fixation, Rubisco and carbonic anhydrase. In 2021 she returned back to Uppsala to start her independent research group and continue the research on the cyanobacterial CO₂ concentration mechanism.