Role of the Protein Corona in the Colloidal Behavior of Microplastics.

Microparticles of polyethylene and polypropylene are largely found in aquatic environments because they are the most produced and persistent plastic materials. Once in biological media, they are covered by a layer of molecules, the so-called corona, mostly composed of proteins. A yeast protein extract from Saccharomyces cerevisiae was used as a protein system to observe interactions in complex biological media. Proteins, acting as surfactants and providing hydrophilic surfaces, allow the dispersion of highly hydrophobic particles in water and stabilize them. After 24 h, the microplastic quantity was up to 1 × 1011 particles per liter, whereas without protein, no particles remained in solution. Label-free imaging of the protein corona by synchrotron radiation deep UV fluorescence microscopy (SR-DUV) was performed. In situ images of the protein corona were obtained, and the adsorbed protein quantity, the coverage rate, and the corona heterogeneity were determined. The stability kinetics of the microplastic suspensions were measured by light transmission using a Turbiscan analyzer. Together, the microscopic and kinetics results demonstrate that the protein corona can very efficiently stabilize microplastics in solution provided that the protein corona quality is sufficient. Microplastic stability depends on different parameters such as the particle's intrinsic properties (size, density, hydrophobicity) and the protein corona formation that changes the particle wettability, electrostatic charge, and steric hindrance. By controlling these parameters with proteins, it becomes possible to keep microplastics in and out of solution, paving the way for applications in the field of microplastic pollution control and remediation.

A proteome scale study reveals how plastic surfaces and agitation promote protein aggregation.

Protein aggregation in biotherapeutics can reduce their activity and effectiveness. It may also promote immune reactions responsible for severe adverse effects. The impact of plastic materials on protein destabilization is not totally understood. Here, we propose to deconvolve the effects of material surface, air/liquid interface, and agitation to decipher their respective role in protein destabilization and aggregation. We analyzed the effect of polypropylene, TEFLON, glass and LOBIND surfaces on the stability of purified proteins (bovine serum albumin, hemoglobin and α-synuclein) and on a cell extract composed of 6000 soluble proteins during agitation (P = 0.1-1.2 W/kg). Proteomic analysis revealed that chaperonins, intrinsically disordered proteins and ribosomes were more sensitive to the combined effects of material surfaces and agitation while small metabolic oligomers could be protected in the same conditions. Protein loss observations coupled to Raman microscopy, dynamic light scattering and proteomic allowed us to propose a mechanistic model of protein destabilization by plastics. Our results suggest that protein loss is not primarily due to the nucleation of small aggregates in solution, but to the destabilization of proteins exposed to material surfaces and their subsequent aggregation at the sheared air/liquid interface, an effect that cannot be prevented by using LOBIND tubes. A guidance can be established on how to minimize these adverse effects. Remove one of the components of this combined stress - material, air (even partially), or agitation - and proteins will be preserved.

Protein Corona Composition of Silica Nanoparticles in Complex Media: Nanoparticle Size does not Matter.

Biomolecules, and particularly proteins, bind on nanoparticle (NP) surfaces to form the so-called protein corona. It is accepted that the corona drives the biological distribution and toxicity of NPs. Here, the corona composition and structure were studied using silica nanoparticles (SiNPs) of different sizes interacting with soluble yeast protein extracts. Adsorption isotherms showed that the amount of adsorbed proteins varied greatly upon NP size with large NPs having more adsorbed proteins per surface unit. The protein corona composition was studied using a large-scale label-free proteomic approach, combined with statistical and regression analyses. Most of the proteins adsorbed on the NPs were the same, regardless of the size of the NPs. To go beyond, the protein physicochemical parameters relevant for the adsorption were studied: electrostatic interactions and disordered regions are the main driving forces for the adsorption on SiNPs but polypeptide sequence length seems to be an important factor as well. This article demonstrates that curvature effects exhibited using model proteins are not determining factors for the corona composition on SiNPs, when dealing with complex biological media.

Glutathione is essential to preserve nuclear function and cell survival under oxidative stress.

Glutathione (GSH) is considered the most important redox buffer of the cell. To better characterize its essential function during oxidative stress conditions, we studied the physiological response of H2O2-treated yeast cells containing various amounts of GSH. We showed that the transcriptional response of GSH-depleted cells is severely impaired, despite an efficient nuclear accumulation of the transcription factor Yap1. Moreover, oxidative stress generates high genome instability in GSH-depleted cells, but does not activate the checkpoint kinase Rad53. Surprisingly, scarce amounts of intracellular GSH are sufficient to preserve cell viability under H2O2 treatment. In these cells, oxidative stress still causes the accumulation of oxidized proteins and the inactivation of the translational activity, but nuclear components and activities are protected against oxidative injury. We conclude that the essential role of GSH is to preserve nuclear function, allowing cell survival and growth resumption after oxidative stress release. We propose that cytosolic proteins are part of a protective machinery that shields the nucleus by scavenging reactive oxygen species before they can cross the nuclear membrane.

Repression of class I transcription by cadmium is mediated by the protein phosphatase 2A.

Toxic metals are part of our environment, and undue exposure to them leads to a variety of pathologies. In response, most organisms adapt their metabolism and have evolved systems to limit this toxicity and to acquire tolerance. Ribosome biosynthesis being central for protein synthesis, we analyzed in yeast the effects of a moderate concentration of cadmium (Cd(2+)) on Pol I transcription that represents >60% of the transcriptional activity of the cells. We show that Cd(2+) rapidly and drastically shuts down the expression of the 35S rRNA. Repression does not result from a poisoning of any of the components of the class I transcriptional machinery by Cd(2+), but rather involves a protein phosphatase 2A (PP2A)-dependent cellular signaling pathway that targets the formation/dissociation of the Pol I-Rrn3 complex. We also show that Pol I transcription is repressed by other toxic metals, such as Ag(+) and Hg(2+), which likewise perturb the Pol I-Rrn3 complex, but through PP2A-independent mechanisms. Taken together, our results point to a central role for the Pol I-Rrn3 complex as molecular switch for regulating Pol I transcription in response to toxic metals.

Endoplasmic reticulum is a major target of cadmium toxicity in yeast.

Cadmium (Cd(2+)) is a very toxic metal that causes DNA damage, oxidative stress and apoptosis. Despite many studies, the cellular and molecular mechanisms underlying its high toxicity are not clearly understood. We show here that very low doses of Cd(2+) cause ER stress in Saccharomyces cerevisiae as evidenced by the induction of the unfolded protein response (UPR) and the splicing of HAC1 mRNA. Furthermore, mutant strains (Delta ire1 and Delta hac1) unable to induce the UPR are hypersensitive to Cd(2+), but not to arsenite and mercury. The full functionality of the pathways involved in ER stress response is required for Cd(2+) tolerance. The data also suggest that Cd(2+)-induced ER stress and Cd(2+) toxicity are a direct consequence of Cd(2+) accumulation in the ER. Cd(2+) does not inhibit disulfide bond formation but perturbs calcium metabolism. In particular, Cd(2+) activates the calcium channel Cch1/Mid1, which also contributes to Cd(2+) entry into the cell. The results reinforce the interest of using yeast as a cellular model to study toxicity mechanisms in eukaryotic cells.