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Cleavage of RRRRR and RRRRA motifs by rFurin and rKLK8

ABSTRACT: The project aims to determine where recombinant Furin or KLK8 can cleave a motif found at the cleavage site of Activin-A. A mutation at the fifth arginine was also tested in a R310A peptide to look at the effect of this mutation on the cleavage by these two proteases.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Manon Bulliard

LAB HEAD: Daniel Constam

PROVIDER: PXD060918 | Pride | 2025-02-24

REPOSITORIES: Pride

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			Action	DRS
	MB_220614_QEx_Sample_01.raw	Raw
	MB_220614_QEx_Sample_05.raw	Raw
	MB_220614_QEx_Sample_06.raw	Raw
	MB_221110_Qex_01.raw	Raw
	MB_221110_Qex_06.raw	Raw

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SARS-CoV-2 S protein ACE2 interaction reveals novel allosteric targets

Project description:SARS-CoV-2 spike protein binding to receptor ACE2 allosterically enhances furin proteolysisat distal S1/S2 cleavage sites

2021-08-09 | PXD023138 | Pride

Project description:Empty follicle syndrome caused by non-cleavable mutation of ZP2 furin cleavage site

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Sequential Glycosylations at the Multibasic Cleavage Site of SARS-CoV-2 Spike Protein Regulate Viral Activation, Assembly, and Infection

Project description:The multibasic furin cleavage site at the S1/S2 boundary of the spike protein (S protein) is a hallmark of SARS-CoV-2 and plays an crucial role in viral infection. O-glycosylation near the furin site catalyzed by host cell glycosyltransferases can theoretically hinder spike protein processing and impede viral infection, but so far such a hypothesis has not been tested with authentic viruses. The mechanism underlying furin activation also remains poorly understood. In this study, we have discovered that GalNAc-T3 and T7 jointly initiate clustered O-glycosylations in the multibasic S1/S2 boundary region. These O-glycosylations inhibit furin processing of the spike protein and surprisingly suppress the incorporation of S protein into virus-like-particles (VLPs). Mechanistic analysis revealed that the assembly of spike protein into VLPs relies on protein-protein interaction between the furin-cleaved S protein and a double aspartic motif on the membrane protein of SARS-CoV-2, suggesting a novel mechanism for furin activation of the S protein. Interestingly, a point mutation at P681, found in the alpha and delta variants of SARS-CoV-2, confers resistance to glycosylation by GalNAc-T3 and T7, thereby diminishing the inhibitory effect against furin processing. However, an additional mutation at N679 in the most recent omicron variant reverses this resistance, rendering it susceptible to glycosylation in vitro and sensitive to the expression of GalNAc-T3 and T7 in human lung cells. Together, our findings suggest a glycosylation-based defense mechanism employed by host cells against SARS-CoV-2 and unveil the intricate interplay between the host and pathogen at this critical “battlefield” as the virus initially evades and currently succumbs to host cell glycosylation..

2024-03-21 | PXD049110 | Pride

SARS-CoV-2 stock sequencing for infection of nonhuman primates

Project description:SARS-CoV-2 USA/WA1 strain stock was sequenced to determine whether the furin cleavage site was intact.

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Loss of furin in β cells induces an mTORC1-ATF4 anabolic pathway that leads to β cell dysfunction

Project description:ABSTRACT: Furin is a proprotein convertase (PC) responsible for proteolytic activation of a wide array of precursor proteins within the secretory pathway. It maps to the PRC1 locus, a type 2 diabetes susceptibility locus, yet its specific role in pancreatic β cells is largely unknown. The aim of this study was to determine the role of furin in glucose homeostasis. We show that furin is highly expressed in human islets, while PCs that potentially could provide redundancy are expressed at considerably lower levels. β cell-specific furin knockout (βfurKO) mice are glucose intolerant, due to smaller islets with lower insulin content and abnormal dense core secretory granule morphology. RNA expression analysis and differential proteomics on βfurKO islets revealed activation of Activating Transcription Factor 4 (ATF4), which was mediated by mammalian target of rapamycin C1 (mTORC1). βfurKO cells show impaired cleavage of the accessory V-ATPase subunit Ac45, and by blocking this pump in β cells the mTORC1 pathway is activated. Furthermore, βfurKO cells show lack of insulin receptor cleavage and impaired response to insulin. Taken together, these results suggest a model of mTORC1-ATF4 hyperactivation in β cells lacking furin, which causes β cell dysfunction. METHOD: Data obtained for the β cell specific knockout (RIP-Cre+/- furin flox/flox) and control (furin flox/flox) samples were compared to investigate the effect of furin loss on the mRNA profile of islets. Although this method does not directly provide furin targets, the notion of differential gene expression might elucidate affected cellular processes, potentially leading to upstream furin substrates.

2020-05-12 | GSE150312 | GEO

Genomic determinants of Furin cleavage in diverse European SARS-related bat coronaviruses

Project description:Furin cleavage in European SARS-related bat coronaviruses

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Furin-mediated cleavage of zona pellucida proteins maintains oocyte integrity and supports oocyte survival

Project description:Zona pellucida (ZP) proteins, which play an important role in oocyte development, undergo post-translational regulation through furin-mediated cleavage. However, our understanding of the functional significance of furin-mediated cleavage of ZP proteins in female reproduction remains limited. Here, using mouse models with disrupted furin cleavage sites in ZP1, ZP2, and ZP3, we found that loss of the furin site in ZP2 caused female infertility associated with empty follicle syndrome (EFS), characterized by the absence of oocyte retrieval after ovarian hyperstimulation. In contrast, female mice carrying cleavage-resistant variants at the furin sites of ZP1 and ZP3 exhibited defective zona pellucidae in a subset of oocytes, leading to reduced fecundity. Mechanistically, disruption of the furin cleavage site in ZP2 impaired the transmembrane transport of the non-cleaved ZP2 protein and subsequently reduced the levels of SNARE proteins, ultimately triggering oocyte apoptosis through activation of the p53 and PI3K signaling pathways. Collectively, we uncovered the essential role of furin-mediated cleavage of ZP proteins in female fertility and provided new mechanistic insights into the pathogenesis of EFS. These findings open new avenues for investigating the contribution of post-translational modifications to female reproduction and for developing potential therapeutic strategies to treat female infertility.

2025-10-29 | PXD070013 |

RNA-Seq data from hamster lung hila and peripheries infected with BANAL-20-236, WK-521, and WK-521 dFCS

Project description:Experimental golden hamsters were infected with BANAL-20-236, WK-521 (a parental strain of SARS-CoV-2), and WK-521 dFCS (a WK-521 without furin cleavage site [FCS]) to investigate virological properties and host response in vivo.

2024-05-20 | GSE239893 | GEO

NGF and proNGF activate functionally distinct mRNAs in PC12 cells: an early gene expression profiling

Project description:The biological activities of NGF and of its precursor proNGF are quite distinct, due to different receptor binding profiles, but nothing is known about how proNGF regulates gene expression. We performed experiments to address this question, by verifying whether a proNGF specific transcriptional signature, distinct from that of NGF, could be identified. To this aim, we studied gene expression regulation by proNGF and NGF in PC12 cells incubated for 1 and 4 hours with recombinant NGF and proNGF, in its wild-type or in a furin-cleavage resistant form. mRNA expression profiles were analyzed by whole genome microarrays at these early time points, in order to identify specific profiles of NGF and proNGF. Three-conditions experiment: PC12 cells were treated with NGF, wild type proNGF precursor (proNGF-WT), furin resistant mutated proNGF precursor (proNGF-KR) for 1 and 4 hours. Two biological replicates were performed for each condition. RNA was extracted after 1 hour and 4 hours of treatment for each treatment.

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Project description:3 strains of SARS-CoV-2 were created to the function of the QTQTN motif adjacent to the Furin cleavage site on Spike protein: WT (QTQTN), Mutant 1 (QTQVN), Mutant 2 (QVQVN).

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