Szulc, N. | bioRxiv Science

bioRxiv2022

Exploring a diverse world of effector domains and amyloid signaling motifs in fungal NLR proteins

NLR proteins are intracellular receptors constituting a conserved component of the innate immune system of multicellular organisms. In fungi, NLRs are characterized by high diversity of architectures and presence of amyloid signaling. Here, we explore the diverse world of effector and signaling domains of fungal NLRs using state-of-the-art bioinformatic methods including MMseqs2 for fast clustering, probabilistic context-free grammars for sequence analysis, and AlphaFold2 deep neural networks for structure prediction. In addition to substantially improving the overall annotation, especially in basidiomycetes, the study identifies novel domains and reveals the structural similarity of MLKL-related HeLo- and Goodbye-like domains forming the most abundant superfamily of fungal NLR effectors. Moreover, compared to previous studies, we found several times more amyloid motifs, including novel families, and validated aggregating and prion-forming properties of the most abundant of them in vitro and in vivo. Also, through an extensive in silico search, the NLR-associated amyloid signaling is for the first time identified in basidiomycetes. The emerging picture highlights similarities and differences in the NLR architectures and amyloid signaling in ascomycetes, basidiomycetes and other branches of life.

bioinformatics↗

bioRxiv2020

On the permeability of cell membranes subjected to lipid oxidation

The formation of transient hydrophilic pores in their membranes is a well-recognized mechanism of permeabilization of cells exposed to high-intensity electric pulses. However, the formation of such pores alone is not able to explain all aspects of the so-called electroporation phenomenon. In particular, the reasons for the sustained permeability of cell membranes, which persist long after the pulses application, remain elusive. The complete resealing of the cell membranes takes indeed orders of magnitude longer than the time of electropore closure as reported from molecular modelling investigations. A possible alternative mechanism to explain the observed long-lived permeability of cell membranes, lipid peroxidation, has been previously suggested but the theoretical investigations of membrane lesions, containing excess amounts of hydroperoxides, have shown that the conductivities of such lesions were not high enough to reasonably explain the entire range of experimental measurements. Here, we expand on these studies and investigate the permeability of cell membrane lesions that underwent secondary oxidation. Molecular dynamics simulations and free energy calculations on lipid bilayers in different states show that such lesions provide a better model for post-pulsed permeable and conductive electropermeabilized cells. These results are further discussed in context of sonoporation and ferroptosis, respectively a procedure and a phenomena, among others, in which alike electroporation substantial lipid oxidation might be triggered. HighlightsO_LIThe contribution of secondary lipids oxidation to the permeabilization of model membranes is quantitatively assessed C_LIO_LISmall patches of secondary lipids oxidation cause formation long-lived pores in lipid bilayers. C_LIO_LIThe cholesterol content of membranes enhances the life-time of the formed pores. C_LIO_LIA single pore accounts for the measured post-pulse electropermeabilization of cells. C_LIO_LIThe diffusion of the secondary oxidation lipids, even after pores closure leads to permeability of lipid membrane. C_LI

biophysics↗