Plants act as a crucial interface between humans and their environment. The wide use of nanoparticles (NPs) has raised great concerns about their potential impacts on crop health and food safety, leading to an emerging research theme about the interaction between plants and NPs. However, up to this day even the basic issues concerning the eventual fate and characteristics of NPs after internalization are not clearly delineated due to the lack of a well-established technique for the quantitative analysis of NPs in plant tissues. We endeavored to combine a quantitative approach for NP analysis in plant tissues with TEM to localize the NPs. After using an enzymatic digestion to release the NPs from plant matrices, single particle-inductively coupled plasma-mass spectrometry (SP-ICP-MS) is employed to determine the size distribution of silver nanoparticles (Ag NPs) in tissues of the model plant Arabidopsis thaliana after exposure to 10 nm Ag NPs. Our results show that Macerozyme R-10 treatment can release Ag NPs from Arabidopsis plants without changing the size of Ag NPs. The characteristics of Ag NPs obtained by SP-ICP-MS in both roots and shoots are in agreement with our transmission electron micrographs, demonstrating that the combination of an enzymatic digestion procedure with SP-ICP-MS is a powerful technique for quantitative determination of NPs in plant tissues. Our data reveal that Ag NPs tend to accumulate predominantly in the apoplast of root tissues whereby a minor portion is transported to shoot tissues. Furthermore, the fact that the measured size distribution of Ag NPs in plant tissue is centered at around 20.70 nm, which is larger than the initial 12.84 nm NP diameter, strongly implies that many internalized Ag NPs do not exist as intact individual particles anymore but are aggregated and/or biotransformed in the plant instead.

CO2 enters the biosphere via the slow, oxygen‐sensitive carboxylase, Rubisco. To compensate, most microalgae saturate Rubisco with its substrate gas through a carbon dioxide concentrating mechanism. This strategy frequently involves compartmentalization of the enzyme in the pyrenoid, a non‐membrane enclosed compartment of the chloroplast stroma. Recently, tremendous advances have been achieved concerning the structure, physical properties, composition and in vitro reconstitution of the pyrenoid matrix from the green alga Chlamydomonas reinhardtii. The discovery of the intrinsically disordered multivalent Rubisco linker protein EPYC1 provided a biochemical framework to explain the subsequent finding that the pyrenoid resembles a liquid droplet in vivo. Reconstitution of the corresponding liquid‐liquid phase separation using pure Rubisco and EPYC1 allowed a detailed characterization of this process. Finally, a large high‐quality dataset of pyrenoidal protein‐protein interactions inclusive of spatial information provides ample substrate for rapid further functional dissection of the pyrenoid. Integrating and extending recent advances will inform synthetic biology efforts towards enhancing plant photosynthesis as well as contribute a versatile model towards experimentally dissecting the biochemistry of enzyme‐containing membraneless organelles.

Nature’s vital, but notoriously inefficient, CO2-fixing enzyme Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) often limits the growth of photosynthetic organisms including crop species. Form I Rubiscos comprise eight catalytic large- and eight auxiliary small-subunits and can be classified into two distinct lineages – “red” and “green”. While red-type Rubiscos (Form IC and ID) are found in Rhodophytes, their secondary symbionts and certain proteobacteria, green-type Rubiscos (Form IA and IB) exist in terrestrial plants, Chlorophytes, cyanobacteria and other proteobacteria. Eukaryotic red-type Rubiscos exhibit desirable kinetic properties, namely high specificity and high catalytic efficiency, with certain isoforms outperforming green-type Rubiscos. However, it is not yet possible to functionally express a high-performing red-type Rubisco in chloroplasts to boost photosynthetic carbon assimilation in green plants. Understanding the molecular and evolutionary basis for divergence between red- and green-type Rubiscos could help us to harness the superior CO2-fixing power of red-type Rubiscos. Here we review our current understanding about red-type Rubisco distribution, biogenesis and sequence-structure and present opportunities and challenges to utilising red-type Rubisco kinetics towards crop improvements.

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vi 4.4.1 Partitioning of PtRubisco is sub-stoichiometric and concentration dependent .