viruses Review Emerging Role of PYHIN Proteins as Antiviral Restriction Factors Matteo Bosso and Frank Kirchhoff * Institute of Molecular Virology, Ulm University Medical Center, 89081 Ulm, Germany; [email protected] * Correspondence: frank.kirchhoff@uni-ulm.de; Tel.: +49-731-50065150 Academic Editor: Sébastien Nisole Received: 26 November 2020; Accepted: 16 December 2020; Published: 18 December 2020 Abstract: Innate immune sensors and restriction factors are cellular proteins that synergize to build an effective first line of defense against viral infections. Innate sensors are usually constitutively expressed and capable of detecting pathogen-associated molecular patterns (PAMPs) via specific pattern recognition receptors (PRRs) to stimulate the immune response. Restriction factors are frequently upregulated by interferons (IFNs) and may inhibit viral pathogens at essentially any stage of their replication cycle. Members of the Pyrin and hematopoietic interferon-inducible nuclear (HIN) domain (PYHIN) family have initially been recognized as important sensors of foreign nucleic acids and activators of the inflammasome and the IFN response. Accumulating evidence shows, however, that at least three of the four members of the human PYHIN family restrict viral pathogens independently of viral sensing and innate immune activation. In this review, we provide an overview on the role of human PYHIN proteins in the innate antiviral immune defense and on viral countermeasures. Keywords: PYHIN; DNA sensing; restriction factors; viral counteraction; immune evasion 1. Introduction Viruses strictly rely on their host cells for replication and spread. However, although viral pathogens are capable of exploiting numerous cellular factors and pathways, the cell does not provide a friendly environment. As a consequence of countless past encounters with viral pathogens, mammalian cells have evolved sensors of foreign invaders that alert and activate a large variety of antiviral effector proteins [1–5]. Pyrin and hematopoietic interferon-inducible nuclear (HIN) domain-containing (PYHIN) protein family members have initially been recognized as novel types of pattern recognition receptors (PRRs) and proposed to trigger innate immune responses and inflammasome activation upon detection of pathogen-derived DNA [6–8]. However, most of the evidence comes from numerous studies on the PYHIN protein AIM2 (Absent In Melanoma 2), a cytoplasmic sensor of double-stranded DNAs [9–12]. In contrast to AIM2, however, the remaining human PYHIN proteins are predominantly localized in the nucleus and accumulating evidence suggests that they exert antiviral effects by suppressing viral transcription rather than by sensing viral DNAs. This review aims at summarizing recent findings that support a role of human PYHIN proteins as antiviral restriction factors. 2. The PYHIN Protein Family PYHIN proteins are characterized by two functional domains: an N-terminal pyrin domain (PYD) and at least one C-terminal hematopoietic interferon-inducible nuclear protein with a 200-amino-acid repeat domain (HIN200) (Figure1). The PYD is part of the bigger superfamily of death domains (DD) characterized by an alpha-helical-based folding, promoting homo- or hetero-typic interactions with other PYD-containing proteins. PYD-PYD interactions regulate a variety of cellular processes, Viruses 2020, 12, 1464; doi:10.3390/v12121464 www.mdpi.com/journal/viruses Viruses 2020, 12, x FOR PEER REVIEW 2 of 18 Virusesdomains2020, 12(DD), 1464 characterized by an alpha-helical-based folding, promoting homo- or hetero-typic2 of 17 interactions with other PYD-containing proteins. PYD-PYD interactions regulate a variety of cellular rangingprocesses, from ranging inflammation from inflammation and immunity and to apoptosisimmunity andto apoptosis cell cycle [and13]. Thecell HINcycle domain[13]. The is onlyHIN founddomain in is PYHIN only found family in members PYHIN family and promotes members DNA-binding and promotes in DNA-binding a non-sequence in specifica non-sequence fashion viaspecific tandem fashion oligonucleotide via tandem/oligosaccharide-binding oligonucleotide/oligosaccharide-binding (OB) folds [14,15]. Sequence(OB) folds independent [14,15]. Sequence DNA bindingindependent is achieved DNA bybinding electrostatic is achieved interactions by electrostati betweenc specific interactions side chains between of positively specific side charged chains HIN of domainpositively amino charged acid residuesHIN domain and theamino phosphate acid residues groups and in thethe DNAphosphate backbone groups [16 ].in HINthe DNA domains backbone have been[16]. classifiedHIN domains in three have subfamilies, been classified designated in three -A,subfamilies, -B, and -C, designated based on -A, the -B, amino and acidic-C, based sequence on the followingamino acidic a conserved sequence MFHATVAT following a conserved motif [17]. MFHATVAT motif [17]. Figure 1. The human pyrin and hematopoietic interferon-inducible nuclear (HIN) domain (PYHIN) proteinFigure family.1. The human Schematic pyrin organization and hematopoietic of human interfer PYHINon-inducible proteins. nuclear Each PYHIN(HIN) domain family member(PYHIN) possessesprotein family. an N-terminal Schematic pyrin organization domain (PYD) of andhuma onen orPYHIN more HINproteins. domains, Each classified PYHIN as family HIN A, member HIN B andpossesses HIN C. an With N-terminal the exception pyrin ofdomain AIM2, (PYD) all PYHIN and on proteinse or more harbor HIN at domains, least one nuclearclassified localization as HIN A, signalHIN B (NLS). and HIN C. With the exception of AIM2, all PYHIN proteins harbor at least one nuclear localization signal (NLS). PYHIN coding genes are exclusively found in mammals and their numbers range from 1 in horses to upPYHIN to 13 in coding mice [18 genes,19]. Humans are exclusively encode fourfound PYHIN in mammals proteins and (Figure their1): numbersγ-IFN-Inducible range from protein 1 in 16horses (IFI16), to up IFN-Inducible to 13 in mice protein [18,19]. X Humans (IFIX) also encode known four as PyrinPYHIN and proteins HIN domain-containing (Figure 1): γ-IFN-Inducible protein 1 (PYHIN1),protein 16 Myeloid(IFI16), IFN-Inducibl Nuclear Differentiatione protein X Antigen (IFIX) also (MNDA), known and as Pyrin Absent and In HIN Melanoma domain-containing 2 (AIM2) [7]. MNDAprotein was1 (PYHIN1), the first human Myeloid PYHIN Nuclear family Differentiat memberion that Antigen was discovered (MNDA), and and is foundAbsent in In the Melanoma nucleus of 2 myeloid(AIM2) [7]. cells MNDA [20]. The was di fftheerentiation first human stage- PYHIN and lineage-specificfamily member expression that was discovered of MNDA and [21] is suggested found in athe potential nucleus role of myeloid in transcriptional cells [20]. regulation The differentiation of myeloid stage- cell di andfferentiation. lineage-specific In this expression context, MNDA of MNDA has been[21] suggested shown to interacta potential with role nucleolin in transcriptional and nucleophosmin regulation [22 of,23 myeloid], as well cell as differentiation. to bind and enhance In this DNA-bindingcontext, MNDA affi hasnity been of the shown transcription to interact factor with Ying nucleolin Yang 1and (YY1) nucleophosmin [22]. Furthermore, [22,23], MNDA as well itself as to hasbind been and proposed enhance to DNA-binding directly regulate affinity transcription of the intranscription monocytes [24factor]. Ying Yang 1 (YY1) [22]. Furthermore,The second MNDA PYHIN itself family has memberbeen proposed to be discovered to directly was regulate IFI16 [transcription25]. IFI16 is expressed in monocytes in various [24]. cell typesThe andsecond tissues. PYHIN Initial family studies member focused to onbe thediscovered role of IFI16 was inIFI16 modulating [25]. IFI16 transcription is expressed and in thevarious cell cycle.cell types IFI16 and interacts tissues. with Initial the studies tumor suppressorfocused on p53the [role26] andof IFI16 increases in modulating its transcriptional transcription activity and [27 the]. Morecell cycle. recent IFI16 findings interacts indicate with that the IFI16tumor is suppressor required for p53 optimal [26] and RNA incr polymeraseeases its transcriptional type II (RNA activity pol II) binding[27]. More to the recent promoters findings of interferonindicate that (IFN)- IFI16α and is required IFN-stimulated for optimal genes RNA (ISGs) polymerase [28]. In addition, type II it(RNA has beenpol II) shown binding that IFI16to the acts promoters as a transcriptional of interferon repressor (IFN)- thatα and inhibits IFN-stimulated a minimal promoter genes (ISGs) containing [28]. anIn intactaddition, Sp1 bindingit has been site [shown29,30]. Thesethat IFI16 early acts studies as a suggestedtranscriptional that the repressor inhibitory that activity inhibits is dependenta minimal onpromoter the HIN containing domains [29 an] andintact that Sp1 IFI16 binding may compete site [29,30]. with Sp1These for DNAearly bindingstudies [suggested30]. IFI16 wasthat also the showninhibitory to directly activity interact is dependent with the on p53 the and HIN c-myc domains gene promoters[29] and that [31 ]IFI16 and tomay promote compete cell with cycle Sp1 arrest for andDNA induction binding of[30]. cellular IFI16 was senescence also shown [32]. to Finally, directly it interact has been with reported the p53 that and IFI16 c-myc interacts gene promoters with the breast[31] and cancer to promote type-1 susceptibility cell cycle arrest protein and (BRCA1) induction and of