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LDLR, LRP1, and Megalin redundantly participate in the uptake of Clostridium novyi alpha-toxin.

ABSTRACT: Clostridium novyi alpha-toxin (Tcnα) is a potent exotoxin that induces severe symptoms including gas gangrene, myositis, necrotic hepatitis, and sepsis. Tcnα binds to sulfated glycosaminoglycans (sGAG) for cell-surface attachment and utilizes low-density lipoprotein receptor (LDLR) for rapid entry. However, it was also shown that Tcnα may use alternative entry receptors other than LDLR. Here, we define that LRP1 and Megalin can also facilitate the cellular entry of Tcnα by employing reconstitutive LDLR family proteins. LDLR, LRP1, and Megalin recognize Tcnα via their ligand-binding domains (also known as LDL receptor type A repeats). Notably, LDLR and LRP1 have contrasting expression levels in many different cells, thus the dominant entry receptor for Tcnα could be cell-type dependent. These findings together increase our knowledge of the Tcnα actions and further help to understand the pathogenesis of C. novyi infection-associated diseases.

SUBMITTER: Zhou Y

PROVIDER: S-EPMC9445046 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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LDLR, LRP1, and Megalin redundantly participate in the uptake of Clostridium novyi alpha-toxin.

Zhou Yao Y Li Danyang D Li Diyin D Chen Aizhong A He Liuqing L Luo Jianhua J Tao Liang L

Communications biology 20220905 1

Clostridium novyi alpha-toxin (Tcnα) is a potent exotoxin that induces severe symptoms including gas gangrene, myositis, necrotic hepatitis, and sepsis. Tcnα binds to sulfated glycosaminoglycans (sGAG) for cell-surface attachment and utilizes low-density lipoprotein receptor (LDLR) for rapid entry. However, it was also shown that Tcnα may use alternative entry receptors other than LDLR. Here, we define that LRP1 and Megalin can also facilitate the cellular entry of Tcnα by employing reconstituti ...[more]

PMID: 36064583

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Project description:BackgroundThe aggregation and spread of α-synuclein (α-Syn) protein and related neuronal toxicity are the key pathological features of Parkinson's disease (PD) and Lewy body dementia (LBD). Studies have shown that pathological species of α-Syn and tau can spread in a prion-like manner between neurons, although these two proteins have distinct pathological roles and contribute to different neurodegenerative diseases. It is reported that the low-density lipoprotein receptor-related protein 1 (LRP1) regulates the spread of tau proteins; however, the molecular regulatory mechanisms of α-Syn uptake and spread, and whether it is also regulated by LRP1, remain poorly understood.MethodsWe established LRP1 knockout (LRP1-KO) human induced pluripotent stem cells (iPSCs) isogenic lines using a CRISPR/Cas9 strategy and generated iPSC-derived neurons (iPSNs) to test the role of LRP1 in α-Syn uptake. We treated the iPSNs with fluorescently labeled α-Syn protein and measured the internalization of α-Syn using flow cytometry. Three forms of α-Syn species were tested: monomers, oligomers, and pre-formed fibrils (PFFs). To examine whether the lysine residues of α-Syn are involved in LRP1-mediated uptake, we capped the amines of lysines on α-Syn with sulfo-NHS acetate and then measured the internalization. We also tested whether the N-terminus of α-Syn is critical for LRP1-mediated internalization. Lastly, we investigated the role of Lrp1 in regulating α-Syn spread with a neuronal Lrp1 conditional knockout (Lrp1-nKO) mouse model. We generated adeno-associated viruses (AAVs) that allowed for distinguishing the α-Syn expression versus spread and injected them into the hippocampus of six-month-old Lrp1-nKO mice and the littermate wild type (WT) controls. The spread of α-Syn was evaluated three months after the injection.ResultsWe found that the uptake of both monomeric and oligomeric α-Syn was significantly reduced in iPSNs with LRP1-KO compared with the WT controls. The uptake of α-Syn PFFs was also inhibited in LRP1-KO iPSNs, albeit to a much lesser extent compared to α-Syn monomers and oligomers. The blocking of lysine residues on α-Syn effectively decreased the uptake of α-Syn in iPSNs and the N-terminus of α-Syn was critical for LRP1-mediated α-Syn uptake. Finally, in the Lrp1-nKO mice, the spread of α-Syn was significantly reduced compared with the WT littermates.ConclusionsWe identified LRP1 as a key regulator of α-Syn neuronal uptake, as well as an important mediator of α-Syn spread in the brain. This study provides new knowledge on the physiological and pathological role of LRP1 in α-Syn trafficking and pathology, offering insight for the treatment of synucleinopathies.

Dataset Information

LDLR, LRP1, and Megalin redundantly participate in the uptake of Clostridium novyi alpha-toxin.

Publications

LDLR, LRP1, and Megalin redundantly participate in the uptake of Clostridium novyi alpha-toxin.

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