Conference Agenda

Rubisco is the central CO₂ fixing enzyme of the Calvin cycle and responsible for the vast majority of all CO₂ fixation on our planet today. In plants, Rubisco undergoes an elaborate set of steps involving the sequential action of at least 6 different dedicated folding and assembly chaperones to assemble into its enzymatically active form. This complexity evolved from much simpler Rubisco ancestors that functioned without any of these additional factors. In this talk I will summarize my lab’s work on retracing the evolution of Rubisco’s complex present-day assembly requirements. Using ancestral sequence reconstruction and the resurrection of billion-year-old Rubiscos, we are learning how this crucial enzyme gradually elaborated its structure and assembly mechanism. Some of these elaborations had history-changing effects on Rubisco’s catalytic properties, whereas others appear to be evolutionary accidents that simply became impossible to lose. This work is beginning to illuminate key events in Rubisco’s history leading up to and following the evolution of oxygenic photosynthesis, one of the most consequential events in the history of life on earth. It also raises the possibility of learning from evolution to re-simplify and improve the assemblies of agriculturally important Rubiscos.

Many microorganisms and upper eukaryotes produce specific protection structures, known as spores or cysts, or cuticular external processes, useful in nature to spark their resistance to adverse environmental conditions.[1] Diatoms, waterish photosynthetic microorganisms, which uptake inorganic silicates to produce highly porous biosilica shells, called frustules, are often exploited for producing biohybrid materials for applications in photonics, optoelectronics and biomaterial science.[2] Aquatic and soil bacteria extrude specific polysaccharides and proteinaceous materials to coat and protect single cells and mobilize metals from soil and water sources. Egg capsules of certain marine snails are protein structures with such interesting meso-porosity, able to protect snail larvae from UV, toxic chemicals and salt unbalances.All these structures can be in principle chemically decorated via common surface, green, bioorganic chemistry approaches aiming to incorporate functional organic molecules, phosphorescent or magnetic nanoparticles and pharmacological moieties.[3-4] In this abstract we will present a plethora of bioorganic methodologies to increase bioremediation potential of matrices biosynthesized by marine and soil microorganisms and eukaryotes. In details: i. potentiated diatoms via the production of a fully organic artificial melanin-like coating around single cells for bioremediation perspective [5], ii. eggs capsules from Sea Snail banded dye-murex functionalized with poly-phenolic chemical bulks to remove pharmaceutical pollutants, iii. soil microorganisms (Pseudomonas fluorescens, purple bacteria) immobilized on abiotic materials for bioelectronics and remediation perspectives. These bioorganic outcomes pave the way to the use of living microorganisms and their extracts in different scientific and applicative areas, such as biomedicine, cell-based technologies and living devices for bioremediation.

Aknowledgements: This work was financially supported by “Dipartimenti di Eccellenza-progetto MAR.V.E.L. (MARginal areas: Valorization of Ecosystem resources for fair and sustainable Livelihood, ICT-2018-20)”.

[1] M. Lo Presti, D. Vona, R. Ragni, S.R. Cicco, G.M. Farinola, MRS Comm. 11, 213–225 (2021). [2] R. Ragni,S.R. Cicco, D. Vona, and G.M. Farinola, Adv. Mater. 1704289,1-23 (2017). [3] G. Leone, G. De la Cruz Valbuena, S.R. Cicco, D. Vona, E. Altamura, R. Ragni, E. Molotokaite, M. Cecchin, S. Cazzaniga, M. Ballottari, C. D'Andrea, G. Lanzani, G. Sci. Rep. 11 (1), 5209 (2021). [4] S.R. Cicco, D. Vona, G. Leone., E. De Giglio, M. Bonifacio, S. Cometa, S. Fiore, F. Palumbo, R. Ragni, G.M. Farinola, Materials Science & Engineering C 104 109897 (2019). [5] D. Vona, S.R. Cicco, R. Ragni, C. Vicente-Garcia, G. Leone, M. Giangregorio, F. Palumbo, E. Altamura, G.M. Farinola, Photochem. Photobiol. Sci. 21, 949–958 (2022).

On Earth, oxygenic photosynthesis is the main process responsible for carbon input into ecosystems, transforming annually more than 100GT of CO₂ into biomass. During photosynthesis, sunlight is converted by the photosynthetic electron transport chain into ATP and NADPH which are used by the metabolism to transform CO₂ into biomass. Alternative electron pathways of photosynthesis have long been proposed to generate the additional ATP requirements of CO₂ fixation, but their relative physiological role and importance have remained elusive. Here, we dissect and quantify the contribution of cyclic, pseudo-cyclic, and chloroplast to mitochondria electron flows for their ability to sustain net photosynthesis in the microalga Chlamydomonas reinhardtii. We show that each pathway has the potential to energize substantial CO₂ fixation, can compensate for each other, and has very different efficiencies at doing it. By exploring their role and quantifying their importance in fluctuating light conditions, we unravel how the dynamic interplay of alternative pathways of photosynthesis maintains the cellular bioenergetic status of the cell. We will discuss how unraveling the bioenergetic landscape of cells can lead to biotechnological improvement of CO₂ capture.

Photosynthesis is the conversion of light energy into biological useful energy in the forms of ATP and NADPH for converting CO₂ to sugars. Light intensity can rapidly fluctuate, while inorganic carbon supply (e.g. CO₂) can also deplete and restrict how much light energy can be consumed for assimilating carbon. Since excess-absorbed light energy can be destructive through the production by chlorophyll of singlet oxygen, a highly reactive oxygen species (ROS), photosynthetic organisms must balance their metabolism with fluctuating environmental factors. Here, data will be presented on how the model unicellular alga, Chlamydomonas reinhardtii, copes with variable environmental conditions, with a focus on inter-organelle bioenergetics. We show that chloroplast-to-mitochondria electron flow is important during recovery from anoxia, whereby electrons are released by alternative oxidase. In contrast, hypoxia enhances electron flow out of the photosynthetic electron transfer system via flavodiiron proteins. In both cases, O₂ is fully reduced to H₂O and ROS production from over-reduced electron transfer systems is avoided. Maximal growth rates are highly dependent on CO₂ availability under high and fluctuating light regimes. This agreed not only with elevated photosynthetic efficiency, but also elevated respiratory rates, as revealed with oxygen flux analysis. With 2 % CO₂, primary metabolite profiling showed increased accumulation of photosynthetic (e.g. sugar phosphates) and respiratory (e.g. TCA cycle) metabolites, as well as amino acids and other organic acids, in comparison to 0.04 % CO₂. A high flux of TCA cycle intermediates occurs during fluctuating light regimes with 2 % CO₂, but not 0.04 % CO₂, again highlighting the contribution of mitochondria to rapid growth. Unrestricting CO₂ availability also increases H₂O₂ production rates, indicating that redox signalling could contribute to cellular proliferation. Overall, our results show how inter-organelle bioenergetics allow algae to cope with adverse environments, as well as enabling maximal growth under optimal conditions.