Furin is the third and best characterized member of the PCSK family of secretory convertases. It cleaves at basic amino acid motifs and recognizes a prototypical sequence R-X-(K/R)-R↓ [10]. The presence of this Furin(-like) motif in viral envelope glycoproteins constitutes a way to activate the fusion-dependant entry of several viruses such as HIV gp160, influenza hemagglutinin (Table 2), and some coronaviruses spike proteins [26,27].

In 1992, Furin was identified to be the cellular protease cleaving hemagglutinin (HA) of fowl plague/influenza virus (FPV; H7N1 A/FPV/Rostock/34), as well as the HIV glycoprotein gp160 into gp120 and gp41 [28,29,30]. The presence of a Furin cleavage sequence can shift the pathogenicity of certain viruses from non-virulent to virulent. For instance, Newcastle disease virus (NDV) fusion protein contains only a monobasic cleavage site, while the virulent NDV fusion protein possesses a polybasic sequence of amino acids that can be cleaved by Furin [31]. Similarly, the acquisition of a Furin cleavage site has been linked to the high pathogenicity of avian influenza [32], especially for the Hong Kong variant (H5N1 A/HK/97) [33,34] that has an insertion of RERR between the NTPQ and the Furin-site RKKR↓GL in HA (Table 2), resulting in a very efficient cleavage by more than one convertase, such as Furin, PC5 and PC7 [35]. Other pathogenic influenza virus do not possesses a Furin cleavage site and rely on trypsin-like proteases, e.g., HAT and TMPRSS2 [36]. The expression of these enzymes is restricted to the upper respiratory tract [37], while Furin is ubiquitously expressed, and the acquisition of a Furin(-like) site enhances the cellular tropism of influenza. However, the presence of a polybasic Furin(-like) cleavage site is not the only determining factor for cleavage by Furin, as the amino-acids adjacent to the Furin(-like) sequence are important too, as described in Table 2. Furthermore, the presence of oligosaccharides near the cleavage site can prevent processing [38], unless it is compensated by an increased number of basic residues as described in the non-pathogenic variant of influenza hemagglutinin A of H5N2 [39].

Virus	Glycoprotein	P8		P6		P4		P2	↓		P2′
HIV	gp160			V	Q	R	E	K	R	A	V
H7N1 A/FPV/Rostock/34	HA			K	K	R	E	K	R	G	L
Avian H5N8 TKY/IRE	HA			R	K	R	K	K	R	G	L
Avian H5N1 A/HK/97	HA	R	E	R	R	R	K	K	R	G	L
Avian H5N1 TKY/ENG	HA	N	T	P	Q	R	K	K	R	G	L
Human CMV	gB			H	N	R	T	K	R	S	T
Human MPV	F Protein			N	P	R	Q	S	R	F	V
Human RSV	F Protein			K	K	R	K	R	R	F	L
Dengue Virus (DENG2)	PrM			H	R	R	E	K	R	S	V
Ebola Virus	gp160			G	R	R	T	R	R	E	A
Chikungunya (CHIKV)	E3E2			P	R	R	Q	R	R	S	I
Zika Virus	PrM			A	R	R	S	R	R	A	V
SARS-CoV-2	S			S	P	R	R	A	R	S	V

Aside from glycoproteins from NDV and influenza hemagglutinin, HIV glycoprotein, as well as other retrovirus glycoproteins such as Rous sarcoma virus and murine leukemia virus are cleaved by Furin. Interestingly, these glycoproteins form trimers, and some viruses assemble with un-cleaved glycoproteins that remain either inactive or can be cleaved at the plasma membrane. The Furin-specific cleavage of gp160 in HIV-1 has been described as dispensable as other proprotein convertases [40] could substitute for Furin in some tissues, and plasmin could activate the gp160 at the plasma membrane. However, these hypotheses remain to be validated in vivo in humans.

Human metapneumovirus (hMPV) is a paramyxovirus responsible for acute respiratory tract infections which can result in hospitalization of both children and adults. We showed that blocking the activity of the protease-activated receptor 1 (PAR1), a G-coupled receptor, induced by inflammation, protects against hMPV infections [41]. Furthermore, Furin activates the envelope F-protein of hMPV by cleavage at NPRQSR₁₀₂↓FV (Table 2, where bold residues emphasise the critical P1 and P4 positions) [41]. Unexpectedly, PAR1 itself potently inhibits cellular Furin activity [42]. Indeed, PAR1 exhibits in its second luminal loop a Furin(-like) motif (PRSFLLR₄₆-NP) with a P1 and P6 Arg and is cleaved by PC5A and PACE4 but not by Furin. The presence of an Asn₄₇ at the P1′ site rather made it a potent Furin-inhibitor that binds its catalytic subunit and sequesters the [PAR1-Furin] complex in the trans-Golgi network (TGN), thereby inactivating Furin and preventing PAR1 from reaching the cell surface [42]. The overexpression of PAR1 has been described in the brain upon neuroinflammation and in patients with neurocognitive disorders associated with HIV infection (HAND) [42]. Interestingly, while PAR1 binds and inhibits Furin, PAR2 is cleaved by Furin at the motif RSSKGR₃₆↓SL, which also exhibits a P1 and P6 Arg but a favorable Ser at P1′ and Leu at P2′ (Table 2) [38]. Recently, another strategy of viral host-cell defense has been identified. Guanylate-binding proteins (GBPs) are interferon stimulated genes. Upon HIV infection, the cytosolic GBP2 and 5 expressions are induced, and these proteins can bind the cytosolic tail of Furin and prevent its trafficking beyond the cis/medial Golgi, thus keeping it in an inactive state bound to its inhibitory prodomain. This effectively inhibits the Furin processing of gp160 into gp120 and gp41 [25,43]. Thus, while PAR1 is the first secretory endogenous natural inhibitor of Furin that blocks its enzymatic activity and prevents its exit from the TGN, the cytosolic GBP2,5 also inhibits Furin activity by blocking it in the cis/medial Golgi in an inactive state.

Another family of viruses requiring Furin is the filoviridae. Marburg and Ebola are enveloped single negative strand RNA viruses of the filoviridae family. These viruses are highly pathogenic in humans causing hemorrhagic fever with a very high mortality rate. The glycoprotein of Marburg and Ebola (GPs) are cleaved by Furin into GP1 containing the receptor binding domain (RBD) and membrane-bound GP2 [44]. Pathogenic viruses of the family possess a Furin(-like) cleavage site, but experiments design to establish the role of Furin in pathogenicity indicated that Furin cleavage is not required for replication in cell culture [45] and disease severity in non-human primates [46]. Recent work identified a new Furin site in Ebola GP that requires N-glycosylation to be processed [47]. Interestingly, MARCH8 a member of the Membrane-associated RING-CH-type 8 (MARCH8) that has been described to have broad antiviral activity against several viruses, can inhibit Furin by forming a complex with the GP and Furin thereby sequestering it in the Golgi, and preventing the maturation of the viral particles [48]. This is the third example of endogenous Furin-inhibitors that sequester the enzyme in a subcellular Golgi compartment where it remains inactive.

The Flaviviruses envelope also requires Furin to get activated. The first evidence was reported for the Tick Borne Encephalitis virus. Flavivirus are enveloped positive strand RNA viruses. Their surface protein is composed of a heterodimer formed by prM and E [49]. The immature prM is cleaved by Furin at GSRTRR₂₀₅↓SV before the viral particles are released from the cell. The cleavage allows fusion and is required for infectivity. An acidic pH is required to promote a change of conformation of the heterodimer to facilitate Furin cleavage. While, the cleavage of prM into M is inefficient especially for Dengue virus (HRREKR₂₀₅↓SV) (Table 2), likely due to the presence of a Glu at P3 [21], Furin has been confirmed to be required for infectivity of Dengue virus in vitro by using LoVo cells that are Furin deficient [50]. Additionally, the role of Furin in the antibody-dependent-enhancement (ADE) of Dengue virus infection is intriguing. ADE is the mechanism by which the immunisation against Dengue could exacerbate the pathogenicity of Dengue in the context of a second infection. This has been explained by the fact that the binding of antibodies to an immature particle could enhance its infectivity. Interestingly, while Furin inhibition has been described to block anti-Envelope dependant enhancement [51], antibodies against prM (uncleaved by Furin) allow tissue entry of immature particles [52]. Recently, broadly neutralizing monoclonal antibodies were reported to protect against multiple tick-borne flaviviruses (TBFVs) [53], opening the way to the design of vaccines and antibody therapeutics against clinically relevant TBFVs.

Chikungunya virus (CHIKV) is a mosquito-transmitted α-virus that causes in humans an acute infection characterized by polyarthralgia, fever, myalgia, and headache. Since 2005 this virus has been responsible for an epidemic outbreak of unprecedented magnitude. By analogy with other α-viruses, it is thought that cellular proteases can process the viral precursor protein E3E2 to produce the receptor binding E2 protein that associates as a heterodimer with E1. Destabilization of the heterodimer by exposure to low pH allows viral fusion and infection. We demonstrated that membrane-bound Furin but also PC5B can process E3E2 from African CHIKV strains at the HRQRR₆₄₂↓ST site, whereas a CHIKV strain of Asian origin is cleaved at RRQRR₆₄₂↓SI (Table 2) by membranous and soluble Furin, PC5A, PC5B, and PACE4 but not by PC7 or SKI-1/S1P [54]. This cleavage was also observed in CHIKV-infected cells and could be blocked by Furin inhibitor decanoyl-RVKR-chloromethyl ketone. This inhibitor was compared with chloroquine for its ability to inhibit CHIKV spreading in myoblast cell cultures, a cell-type previously described as a natural target of this virus [54]. We observed an additive effect of dec-RVKR-cmk and chloroquine, supporting the concept that these two drugs act by essentially distinct mechanisms. The combinatory action of chloroquine and dec-RVKR-cmk led to almost total suppression of viral spread and yield [54].

Furin cleavage of viral glycoproteins of the Herpesviridae family have been reported for Herpes simplex virus 1 and 2 and Varicella Zoster (Table 1), contributing to the pathogenesis of the later in vivo [55,56]. Finally, while Furin cleaves many glycoproteins, its activity is required for other viral proteins such as for Hepatitis B virus (HBV) core antigen. For reviews see [21,25].

HIV-1 encodes four accessory gene products—Vpr, Vif, Vpu, and Nef—which are thought to collectively manipulate host cell biology to promote viral replication, persistence, and immune escape [57]. Increasing evidence suggests that extracellular Vpr could contribute to HIV pathogenesis through its effect on bystander cells. Soluble forms of Vpr have been detected in the sera and cerebrospinal fluids of HIV-1-infected patients, and in vitro studies have implicated extracellular Vpr as an effector of cellular responses, including G2 arrest, apoptosis, and induction of cytokines and chemokines production, presumably through its ability to transduce into multiple cell types. Thus, Vpr is a true virulence factor and is a potential and promising target in different strategies aiming to fight infected cells including latently HIV-infected cells [58]. While Furin is clearly implicated in the processing activation of multiple viral surface glycoproteins, including gp160 of HIV-1, other cell-surface associated convertases such as PC5A and PACE4 [9,59] seem to negatively regulate the activity of the HIV-1 by cleavage-inactivation of secreted Vpr at RQRR₈₈↓ [60]. PC-mediated processing of extracellular Vpr results in a truncated Vpr product that was defective for the induction of cell cycle arrest and apoptosis when expressed in human cells [60]. Thus, inhibitors of PC5/PACE4 may enhance HIV-1 virulence, and Furin-specific inhibitors would restrict HIV-1 infectivity, without affecting Vpr inactivation by PC5A/PACE4.

2.2. PCSK9 and Viral Infections