Inhibition of Soluble Recombinant Furin by Human Proteinase Inhibitor 8

Dahlen, Jeffrey R.; Jean, François; Thomas, Gary; Foster, Donald C.; Kisiel, Walter

doi:10.1074/jbc.273.4.1851

Cited by 87 publications

(73 citation statements)

References 36 publications

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“…Major limitations of these agents include either cell cytotoxicity and/or poor cellular permeability and targeting. Other approaches used recombinant-based inhibitors (19)(20)(21)(22). These strategies are based on the expression of proteins that contain a furin-like recognition sequence (RXXR) within the inhibitor binding region of either human a 1 -antitrypsin (22), a 2 -macroglobulin (21), proteinase-8 (20), or the turkey ovomucoid third domain (19).…”

Section: +mentioning

confidence: 99%

“…Other approaches used recombinant-based inhibitors (19)(20)(21)(22). These strategies are based on the expression of proteins that contain a furin-like recognition sequence (RXXR) within the inhibitor binding region of either human a 1 -antitrypsin (22), a 2 -macroglobulin (21), proteinase-8 (20), or the turkey ovomucoid third domain (19). Although reasonably effective, the inability of these recombinant proteins to inhibit selectively furin and not other proprotein convertases could be problematic.…”

Section: +mentioning

confidence: 99%

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Human Carcinoma Cell Growth and Invasiveness Is Impaired by the Propeptide of the Ubiquitous Proprotein Convertase Furin

et al. 2005

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Furin, a potent proprotein convertase involved in activation of several cancer-related substrates, is synthesized as an inactive zymogen, thus minimizing the occurrence of premature enzymatic activity that would lead to inappropriate protein activation or degradation. This natural inhibitory mechanism is based on the presence of an inactivating prosegment at the NH 2 terminal of the zymogen. After initial autocatalytic cleavage, the prosegment remains tightly associated with the convertase until it reaches the trans-Golgi network where the dissociation of the prosegment and activation of furin occurs. We hypothesized that the inhibitory properties of the preprosegment of furin (ppFur) could be beneficial if ectopically expressed in tumor cells. Transfection of four human head and neck squamous cell carcinoma cell lines with the complete ppFur cDNA sequence (pIRES-EGFP-ppFur) or with the empty expression vector (pIRES-EGFP) was done. The inhibitory effect was evaluated using in vivo tumorigenicity, invasion, anchorage-independent growth in soft agar, and proliferation assays, as well as by investigating impairment of furin substrates processing. Following transfection of ppFur, a significant reduction in cell proliferation, tumorigenicity, and invasiveness was observed in vitro and in vivo. These biological changes are directly related to the inhibition of furin-mediated activation of crucial cancer-related substrates, such as membrane type 1 matrix metalloproteinase, transforming growth factor-B, insulin-like growth factor-1 receptor, and vascular endothelial growth factor-C. PpFur expression in head and neck squamous cell carcinoma cell lines showed a mechanistic link between furin inhibition, decreased substrate processing, cell proliferation, and invasive ability. These findings suggest that furin inhibition is a feasible approach to ameliorate and even abolish the malignant phenotype of various malignancies. (Cancer Res 2005; 65(10): 4162-71)

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confidence: 99%

Section: +mentioning

confidence: 99%

Human Carcinoma Cell Growth and Invasiveness Is Impaired by the Propeptide of the Ubiquitous Proprotein Convertase Furin

et al. 2005

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“…Although the serpins were initially characterized as inhibitors of chymotrypsin-like extracellular serine proteases, in principle they can inhibit any protease that forms a covalent intermediate with the substrate. Examples of serpin-mediated regulation of intracellular processing events include the inhibition of the serine and thiol proteases of neuroendocrine cells (Hwang et al, 1999(Hwang et al, ,2002Fell et al, 2002), as well as the inhibition of SPCs (Dahlen et al, 1998;Dufour et al, 1998;Jean et al, 1998;Tsuji et al, 1999).…”

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confidence: 99%

DrosophilaSerpin 4 Functions as a Neuroserpin-Like Inhibitor of Subtilisin-Like Proprotein Convertases

Osterwalder

Kuhnen²,

Leiserson³

et al. 2004

J. Neurosci.

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The proteolytic processing of neuropeptide precursors is believed to be regulated by serine proteinase inhibitors, or serpins. Here we describe the molecular cloning and functional expression of a novel member of the serpin family, Serine protease inhibitor 4 (Spn4), that we propose is involved in the regulation of peptide maturation in Drosophila. The Spn4 gene encodes at least two different serpin proteins, generated by alternate splicing of the last coding exon. The closest vertebrate homolog to Spn4 is neuroserpin. Like neuroserpin, one of the Spn4 proteins (Spn4.1) features a unique C-terminal extension, reminiscent of an endoplasmic reticulum (ER) retention signal; however, Spn4.1 and neuroserpin have divergent reactive site loops, with Spn4.1 showing a generic recognition site for furin/SPC1, the founding member of the intracellularly active family of subtilisin-like proprotein convertases (SPCs). In vitro, Spn4.1 forms SDS-stable complexes with the SPC furin and directly inhibits it. When Spn4.1 is overexpressed in specific peptidergic cells of Drosophila larvae, the animals exhibit a phenotype consistent with disrupted neuropeptide processing. This observation, together with the unique combination of an ER-retention signal, a target sequence for SPCs in the reactive site loop, and the in vitro inhibitory activity against furin, strongly suggests that Spn4.1 is an intracellular regulator of SPCs.

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“…The various successful approaches include: active site-directed chloromethyl ketone inhibitors (12,13), reversible peptide-based inhibitors (14 -17), plant derivatives (18), and several engineered variants of protein-based inhibitors that possess a furin-like motif. These include ␣2-macroglobulin (␣2-MF) (19), ␣ 1 -antitrypsin (␣ 1 -AT) Portland (␣ 1 -PDX) (20 -22), proteinase inhibitor 8 (PI8) (23), the turkey ovomucoid third domain (24), and eglin C (25,26). However, these effective inhibitors directed against the basic amino acid-specific members lack selectivity.…”

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Development of Protein-based Inhibitors of the Proprotein of Convertase SKI-1/S1P

Pullikotil

Vincent

Nichol

et al. 2004

Journal of Biological Chemistry

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Processing of membrane-bound transcription factors such as sterol regulatory element-binding proteins (SREBPs) and the ER-stress response factor ATF6, and glycoproteins of some hemorrhagic fever viruses are initiated by the proprotein convertase SKI-1/S1P. So far, no cellular protein-based inhibitor of the hydrophobicamino acid specific SKI-1 is known. The prosegment of the basic-amino acid specific convertases (e.g. furin and PC5) or ␣ 1 -PDX, a variant of ␣ 1 -antitrypsin (␣ 1 -AT) exhibiting an RIPR 358 sequence at the reactive site loop, were shown to potently inhibit these secretory proteinases. Accordingly, we tested the SKI-1-inhibitory potential of various point mutants of either the 198 amino acid preprosegment of SKI-1-(1-198) or ␣ 1 -AT. Transient transfections data showed that, out of numerous mutants studied, the R134E prosegment mutant or the ␣ 1 -AT reactive site loop variants RRVL 358 , RRYL 358 and RRIL 358 are the best specific cellular inhibitors of SKI-1. The observed inhibition of the processing of endogenous SREBP-2, exogenous ATF6 and a PDGF-A (RRLL 86 ) variant were >55% and reach ϳ80% in stable transfectants. We also show that SKI-1 forms SDS-stable complexes with these ␣ 1 -AT variants, but not with wild-type ␣ 1 -AT or ␣ 1 -PDX. Finally, these inhibitors were also shown to affect the processing and stability of the Crimean-Congo hemorrhagic fever virus glycoprotein.Proteins and peptides that are biologically active are often generated by intracellular limited proteolysis of inactive precursors. The mammalian proprotein convertases (PCs) 1 of the secretory pathway are calcium-dependent serine proteinases related to bacterial subtilisin that cleave various precursors at the general consensus motif (K/R)(X) n (K/R)2, where n ϭ 0, 2, 4, or 6 and X is any amino acid (1-3). The PC family counts seven basic amino acid-specific kexin-like convertases: furin, PC1/3, PC2, PC4, PACE4, PC5/6, and PC7/LPC (4). The eighth member is the recently discovered pyrolysin-like SKI-1/S1P that cleaves at the consensus motif (R/K)X(hydrophobic)Z2, where Z is variable (5), while the last member (6, 7) NARC-1 cleaves the sequence VFAQ 153 2 in its prosegment (8, 9). 2 More PCs contain an N-terminal signal sequence, followed by a prosegment, a catalytic domain and a P-domain. In addition, PCs possess a C-terminal segment that varies between the different members. The critical role of PCs in the proteolytic maturation of multiple proproteins, their implication in various pathologies (1, 10, 11), and their unidentified specific and/or redundant functions, make them attractive targets for the development of potent and selective inhibitors. The various successful approaches include: active site-directed chloromethyl ketone inhibitors (12, 13), reversible peptide-based inhibitors (14 -17), plant derivatives (18), and several engineered variants of protein-based inhibitors that possess a furin-like motif. These include ␣2-macroglobulin (␣2-MF) (19), ␣ 1 -antitrypsin (␣ 1 -AT) Portland (␣ 1 -PDX) (20 -22), proteinase i...

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Inhibition of Soluble Recombinant Furin by Human Proteinase Inhibitor 8

Cited by 87 publications

References 36 publications

Human Carcinoma Cell Growth and Invasiveness Is Impaired by the Propeptide of the Ubiquitous Proprotein Convertase Furin

Human Carcinoma Cell Growth and Invasiveness Is Impaired by the Propeptide of the Ubiquitous Proprotein Convertase Furin

DrosophilaSerpin 4 Functions as a Neuroserpin-Like Inhibitor of Subtilisin-Like Proprotein Convertases

Development of Protein-based Inhibitors of the Proprotein of Convertase SKI-1/S1P

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