Biochemical Journal最新文献

Amyloidogenic proteins play a central role in a range of pathological conditions, yet their presence in thrombi has only recently been recognized. Whether computational prediction tools can identify amyloid- forming potential in thrombus proteomes remains unclear. AmyloGram is a computational tool that estimates amyloid-forming potential based on n-gram sequence encoding and random forest classification. Using AmyloGram, we analyzed 204 proteins in UniProt that were tagged by humans as amyloidogenic. We then applied the same approach to proteins identified in thrombi retrieved using mechanical thrombectomy from patients with cardioembolic and atherothrombotic stroke. In addition, we used AmyloGram to analyze the amyloidogenicity of 83,567 canonical human protein sequences. Among the UniProt-annotated 'amyloid' set, nearly all proteins received AmyloGram scores above 0.7, including 23 of the 24 human proteins. Even the lowest-scoring human protein, lysozyme (scoring 0.675), is known to form amyloid under certain conditions. In thrombi from both stroke subtypes in four different studies, all detected proteins (with a single exception) had AmyloGram scores above 0.7, suggesting a high likelihood of amyloid content. A majority of unannotated proteins also achieve AmyloGram scores exceeding 0.7. AmyloGram reliably identifies known amyloid-forming proteins and reveals that stroke thrombi are enriched for proteins with high amyloidogenic potential. These findings support the hypothesis that thrombus formation in stroke involves amyloid-related mechanisms and warrant further investigation using histological and functional validation.

RAS proteins have been studied for more than 40 years, mainly in the context of cancer. Given that RAS signaling promotes cell cycle progression, it is commonly assumed that its main function is to drive cell proliferation. In this review, we will, however, address the roles of RAS during cell differentiation, which is intertwined with cell division during organismal development and tissue homeostasis in the adult. Our analysis suggests a far-reaching and profound impact of RAS signaling in associated processes. During vertebrate embryonal development, the FGF-RAS-ERK signaling axis is involved as early as germ layer induction and embryonal patterning. Current evidence suggests that RAS fundamentally controls the balance between stem cells and their differentiated progeny. RAS signaling needs to be downmodulated to sustain pluripotent stem cells. Inhibition of RAS activity is also required to preserve adult stem cell quiescence. At the other end of the differentiation spectrum, a different kind of RAS inactivation by the GTPase-activating protein (GAP) neurofibromin 1 (NF1) appears central to permit terminal differentiation, e.g., of muscle tissue. This latter process is disabled in muscle-borne cancer and likely also in other cancer types. In the RAS-associated developmental diseases, the RASopathies, cell differentiation appears to be broadly perturbed throughout development. We suggest that loss of RAS pathway activity mainly manifests in the stem/progenitor cell compartment, whereas inhibition of NF1 GAP-mediated RAS inactivation blocks terminal differentiation. Given that defects accumulate during development, it is plausible to assume that only progressive pathological phenotypes of RASopathies offer a realistic chance for treatment, notably by repurposing RAS-MAPK pathway oncology drugs. Thus, the impact of RAS on cell differentiation represents, in our view, the common mechanistic denominator of cancer and RASopathies. We conclude by giving a perspective on how improving our insight into the functioning of RAS during cell differentiation could lead to the development of misdifferentiation-correcting drugs.

Type III CRISPR systems typically generate cyclic oligoadenylate second messengers such as cyclic tetra-adenylate (cA4) on detection of foreign RNA. These activate ancillary effector proteins which elicit a diverse range of immune responses. The Calp (CRISPR associated Lon protease) system elicits a transcriptional response to infection when CalpL (Calp Lon protease) binds cA4 in its SAVED (SMODS associated and fused to various effectors domain) sensor domain, resulting in filament formation and activation of the Lon protease domain, which cleaves the anti-Sigma factor CalpT, releasing the CalpS (Calp Sigma factor) for transcriptional remodelling. Here, we show that thermophilic viruses have appropriated the SAVED domain of CalpL as an anti-CRISPR, AcrIII-2 (second anti-CRISPR of type III systems), which they use to degrade cA4. AcrIII-2 dimers sandwich cA4, degrading it in a shared active site to short linear products, using a mechanism highly reminiscent of CalpL. This results in inhibition of a range of cA4 activated effectors in vitro. This is the first example of a virally encoded SAVED domain with ring nuclease activity, highlighting the complex interplay between viruses and cellular defences.

Folates are essential for all organisms. They are acquired either through de novo biosynthesis or from the diet. Yeast, fungi, and plants make their own folates, and it has not been clear if plasma membrane folate transporters exist in these organisms. Using a synthetic lethal screen in Saccharomyces cerevisiae, we observed that deletions in a gene encoding the previously identified glutathione (GSH) transporter, OPT1, exhibited severe growth defects with a disruption in folate biosynthesis. Uptake experiments confirmed that Opt1p/Hgt1p can transport folinic acid and the naturally abundant methyl tetrahydrofolate. As S. cerevisiae Opt1p was able to transport both folate and GSH, we used alanine-scanning mutants in the transmembrane domains of the channel pore to identify the residues required specifically for the uptake of folates and distinct from those required for GSH. We further examined the oligopeptide transporter (OPT) family of other organisms for the presence of folate transporters. In C. albicans, CaOPT1, the ortholog of S. cerevisiae OPT1, efficiently transported folate but not GSH, while the previously characterized GSH transporter, CaOPT7, could not transport folate. Aspergillus fumigatus has eight homologs of the OPT family, of which OptB and OptH transport folates. In the plant Arabidopsis thaliana, the Opt1 homologs AtOpt2, AtOpt4, and AtOpt6 transport folates. This discovery of folate transporters across fungi and plants fills a critical gap in our understanding of folate metabolism and can benefit the exploitation of these pathways in pathogenic fungi and in plants.