Biology subjects

Shakya, B.

Publications and source records attributed to Shakya, B..

4 recordsLinked to original sources

bioRxiv2024

Allosteric degraders induce CRL5ASB8 mediated degradation of XPO1

SummaryThe nuclear export receptor exportin 1 (XPO1/CRM1) is often overexpressed in cancer cells, leading to the mislocalization of numerous cancer-related protein cargoes1,2. Selinexor, a covalent XPO1 inhibitor, and other Selective Inhibitor of Nuclear Export (SINEs) restore proper nuclear localization by blocking XPO1-cargo binding2-7. SINEs also induce XPO1 degradation via the Cullin-RING E3 ubiquitin ligase (CRL) substrate receptor ASB87. Here we elucidate the mechanism underlying the high-affinity engagement of CRL5ASB8 with SINE-conjugated XPO1. Cryogenic electron microscopy (cryoEM) structures reveal that ASB8 binds to a cryptic site on XPO1, which becomes accessible only upon SINE conjugation. While molecular glue degraders typically interact with both CRL and the substrate8-10, SINEs bind to XPO1 without requiring interaction with ASB8 for efficient XPO1 degradation. Instead, an allosteric mechanism facilitates high affinity XPO1-ASB8 interaction, leading to XPO1 ubiquitination and degradation. ASB8-mediated degradation is also observed upon treatment of the endogenous itaconate derivate 4-octyl itaconate, which suggests a native mechanism that is inadvertently exploited by synthesized XPO1 inhibitors. This allosteric XPO1 degradation mechanism of SINE compounds expands the known modes of targeted protein degradation beyond the well-characterized molecular glue degraders and proteolysis targeting chimeras of CRL4.

cancer biology↗

bioRxiv2024

Phosphate-dependent nuclear export via a novel NES class recognized by exportin Msn5

Gene expression in response to environmental stimuli is dependent on nuclear localization of key signaling components, which can be tightly regulated by phosphorylation. This is exemplified by the phosphate-sensing transcription factor Pho4, which requires phosphorylation for nuclear export by the yeast exportin Msn5. Unlike the traditional hydrophobic nuclear export signal (NES) utilized by the Exportin-1/XPO1 system, cryogenic-electron microscopy structures reveal that Pho4 presents a novel, phosphorylated 35-residue NES that interacts with the concave surface of Msn5 through two Pho4 phospho-serines that align with two Msn5 basic patches, unveiling a previously unknown mechanism of phosphate-specific recognition. Furthermore, the discovery that unliganded Msn5 is autoinhibited explains the positive cooperativity of Pho4/Ran-binding and proposes a mechanism for Pho4s release in the cytoplasm. These findings advance our understanding of the diversity of signals that drive nuclear export and how cargo phosphorylation is crucial in regulating nuclear transport and controlling cellular signaling pathways.

biophysics↗

bioRxiv2020

Erythrocyte CD55 mediates the internalization of Plasmodium falciparum parasites

Invasion of human erythrocytes by the malaria parasite Plasmodium falciparum is a multi-step process. Previously, a forward genetic screen for P. falciparum host factors identified erythrocyte CD55 as essential for invasion, but its specific role and how it interfaces with the other factors that mediate this complex process are unknown. Using CRISPR-Cas9 editing, antibody-based inhibition, and live cell imaging, here we show that CD55 is specifically required for parasite internalization. Pre-invasion kinetics, erythrocyte deformability, and echinocytosis were not influenced by CD55, but entry was inhibited when CD55 was blocked or absent. Visualization of parasites attached to CD55-null erythrocytes point to a role for CD55 in progression of the moving junction. Our findings demonstrate that CD55 acts after discharge of the parasites rhoptry organelles, and plays a unique role relative to all other invasion receptors. As the requirement for CD55 is strain-transcendent, these results suggest that CD55 or its interacting partners may hold potential as therapeutic targets for malaria.

microbiology↗

bioRxiv2019

A common polymorphism in the druggable ion channel PIEZO1 is associated with protection from severe malaria

Malaria caused by the Apicomplexan parasite Plasmodium falciparum has served as a strong evolutionary force throughout human history, selecting for red blood cell polymorphisms that confer innate protection against severe disease. Recently, gain-of-function mutations in the mechanosensitive ion channel PIEZO1 were shown to ameliorate Plasmodium parasite growth, blood-brain barrier dysfunction, and mortality in a mouse model of malaria. In humans, the gain-of-function allele PIEZO1 E756del is highly prevalent and enriched in Africans, raising the possibility that it is under positive selection due to malaria. Here we used a case-control study design to test for an association between PIEZO1 E756del and malaria severity among children in Gabon. We found that the E756del variant is strongly associated with protection against severe malaria in heterozygotes, independent of the protection conferred by the sickle cell trait (hemoglobin AS). In vitro experiments using donor red blood cells failed to find an effect of E756del on parasite growth, suggesting this variant confers a mild channel defect and/or that its protective effect may be mediated by other tissue types in vivo. Nonetheless, we show that Yoda1, a small molecule agonist of PIEZO1, has potent antimalarial activity in both E756del and wild-type red blood cells. Our findings demonstrate that PIEZO1 is an important innate determinant of malaria susceptibility in humans and holds potential as druggable host target for malaria control.

microbiology↗