Sakai, Yosuke

Tokushima University Institutional Repository

fmicb-07-01655 October 14, 2016 Time: 13:1 # 1PERSPECTIVEpublished: 18 October 2016doi: 10.3389/fmicb.2016.01655Edited by:Akihide Ryo,Yokohama City University, JapanReviewed by:Hiroaki Takeuchi,Tokyo Medical and Dental University,JapanAyumi Kudoh,Yokohama City University, Japan*Correspondence:Akio Adachiadachi@tokushima-u.ac.jpMasako Nomaguchinomaguchi@tokushima-u.ac.jpSpecialty section:This article was submitted toVirology,a section of the journalFrontiers in MicrobiologyReceived: 23 August 2016Accepted: 04 October 2016Published: 18 October 2016Citation:Sakai Y, Doi N, Miyazaki Y, Adachi Aand Nomaguchi M (2016)Phylogenetic Insights intothe Functional Relationship betweenPrimate Lentiviral ReverseTranscriptase and Accessory ProteinsVpx/Vpr. Front. Microbiol. 7:1655.doi: 10.3389/fmicb.2016.01655Phylogenetic Insights into theFunctional Relationship betweenPrimate Lentiviral ReverseTranscriptase and AccessoryProteins Vpx/VprYosuke Sakai1, Naoya Doi1, Yasuyuki Miyazaki2, Akio Adachi1* and Masako Nomaguchi1*1 Department of Microbiology, Tokushima University Graduate School of Medical Science, Tokushima, Japan, 2 Departmentof Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, Tokyo, JapanThe efficiency of reverse transcription to synthesize viral DNA in infected cellsgreatly influences replication kinetics of retroviruses. However, viral replication in non-dividing cells such as resting T cells and terminally differentiated macrophages ispotently and kinetically restricted by a host antiviral factor designated SAMHD1(sterile alpha motif and HD-domain containing protein 1). SAMHD1 reduces cellulardeoxynucleoside triphosphate (dNTP) pools and affects viral reverse transcription step.Human immunodeficiency virus type 2 (HIV-2) and some simian immunodeficiencyviruses (SIVs) have Vpx or Vpr to efficiently degrade SAMHD1. Interestingly, the reversetranscriptase (RT) derived from HIV-1 that encodes no anti-SAMHD1 proteins hasbeen previously demonstrated to uniquely exhibit a high enzymatic activity. It is thusnot irrational to assume that some viruses may have acquired or lost the specificRT property to better adapt themselves to the low dNTP environments confrontedin non-dividing cells. This adaptation process may probably be correlated with theSAMHD1-antagonizing ability by viruses. In this report, we asked whether such adaptiveevents can be inferable from Vpx/Vpr and RT phylogenetic trees overlaid with SAMHD1-degrading capacity of Vpx/Vpr and with kinetic characteristics of RT. Resultant twotrees showed substantially similar clustering patterns, and therefore suggested that theproperties of RT and Vpx/Vpr can be linked. In other words, HIV/SIVs may possess theirown RT proteins to adequately react to various dNTP circumstances in target cells.Keywords: HIV, SIV, Vpx, Vpr, RT, SAMHD1, dNTPINTRODUCTIONAccessory proteins of various human immunodeficiency viruses/simian immunodeficiency viruses(HIV/SIVs) are believed to be essential for optimal viral replication, persistence, and pathogenicityin vivo (Matheson et al., 2016). Vpx was least well studied among the five accessory proteins(Vif, Vpx, Vpr, Vpu, and Nef) for its functional role and the underlying molecular basis untilthe recent identification of cellular anti-viral factor SAMHD1 as a target for Vpx (Hrecka et al.,2011; Laguette et al., 2011). Extensive studies since then have generated a series of criticalFrontiers in Microbiology | www.frontiersin.org 1 October 2016 | Volume 7 | Article 1655fmicb-07-01655 October 14, 2016 Time: 13:1 # 2Sakai et al. Phylogenetic Insights into RT and Vpx/Vprfindings to understand the interaction of Vpx and SAMHD1(Goldstone et al., 2011; Hrecka et al., 2011; Laguette et al., 2011,2012; Baldauf et al., 2012; Descours et al., 2012; Lahouassa et al.,2012; Lim et al., 2012; St Gelais et al., 2012; Lenzi et al., 2014,2015): (1) all Vpx and some Vpr proteins derived from variousHIV/SIVs target SAMHD1 for proteasomal degradation; (2)SAMHD1 reduces cellular deoxynucleoside triphosphate (dNTP)pools to a level similar to that observed in non-dividing myeloidand resting T cells; (3) HIV-1 reverse transcriptase (RT) showsa high binding affinity to dNTPs relative to those from otherlentiviruses. Of note here, HIV-1 can replicate in macrophages tosome extent, and lacks SAMHD1-degrading activity. Thus, HIV-1 appears to be unique among primate lentiviruses to act againstSAMHD1.On the basis of the experimental results summarized above,it would hold true to hypothesize that some specific viruses haveadapted themselves to better fit the physiological environmentsin non-dividing cells. Therefore, we here have performedphylogenetic analyses using distinct Vpx/Vpr and RT proteinsfrom viruses with/without anti-SAMHD1 activity. To thisend, we phylogenetically examined the target viral proteins(Vpx/Vpr and RT with no sequence ambiguity) derived fromthree types of diverse SIVs based on their genome featuresrelating to vpx, vpr, and vpu genes (Fujita et al., 2010; Sakaiet al., 2016) as follows. (1) “Prototype viruses” carrying vprgene analyzed here were SIVagm (isolated from the Africangreen monkey), SIVmnd-1 (mandrill), SIVlst (l’Hoest’s monkey),SIVsun (sun-tailed monkey), SIVsyk (Sykes’ monkey), SIVdeb(DeBrazza’s monkey), SIVtal (talapoin monkey), SIVasc (red-tailed guenon), SIVcol (colobus monkey), SIVwrc (westernred colobus), and SIVolc (olive colobus). (2) “HIV-1 typeviruses” carrying vpr and vpu genes analyzed here were,SIVcpz (chimpanzee), SIVgor (gorilla), SIVgsn (greater spot-nosed monkey), SIVmon (mona monkey), SIVmus (mustachedmonkey), and SIVden (Dent’s monkey). (3) “HIV-2 type viruses”carrying vpx and vpr genes analyzed here were SIVsmm(sooty mangabey monkey), SIVmac (macaque monkey), SIVmne(pig-tailed macaque), SIVstm (stump-tailed macaque), SIV-rcm (red-capped mangabey), SIVmnd-2 (mandrill), and SIVdrl(drill monkey).PHYLOGENY OF VPX/VPR PROTEINSDERIVED FROM VIRUSESWITH/WITHOUT SAMHD1-DEGRADINGACTIVITYEarly studies have shown that Vpx is essential for HIV-2 andSIVmac replication in primary non-dividing cells (Fujita et al.,2010; Schaller et al., 2014). Subsequent studies have revealedthat Vpx enhances viral DNA synthesis by inducing proteasomaldegradation of an anti-viral factor in those cells (Fujita et al.,2010; Schaller et al., 2014). It is now established that the anti-viral factor SAMHD1 abundantly present in the non-dividingcells is degraded by Vpx from HIV-2 type viruses and Vpr fromsome prototype viruses (Laguette et al., 2012; Lim et al., 2012).There are no Vpr proteins with SAMHD1-degrading activityderived from HIV-1 and HIV-2 type viruses except for Vprfrom SIVmus (HIV-1 type) (Laguette et al., 2012; Lim et al.,2012).In order to easily see the genetic background for the resultsdescribed above (Laguette et al., 2012; Lim et al., 2012), wealso inferred a bootstrap phylogenetic tree of Vpx/Vpr proteinsfrom diverse HIV/SIVs by the neighbor-joining method aspreviously described (Sakai et al., 2016), and the proteinswith/without SAMHD1-degrading activity were highlighted byblue and red letters, respectively. Viruses without Vpx (prototypeand HIV-1 type viruses) were analyzed for their Vpr proteins,and viruses with both Vpx and Vpr (HIV-2 type viruses)were examined for their Vpx proteins. As shown in Figure 1,Vpx/Vpr proteins from viruses with SAMHD1-degrading activity(viral groups: HIV-2, SIVsmm/mac/mne/stm, SIVrcm/mnd-2/drl, SIVagm, SIVmus/gsn/den/mon, and SIVdeb/syk/tal/asc)clearly formed different clusters from those by Vpr proteins fromviruses without SAMHD1-degrading activity (viral groups: HIV-1, SIVcpz/gor, SIVmnd-1/lst/sun, and SIVolc/wrc/col). No virusstrains without SAMHD1-degrading activity were found in theclusters with the activity and the opposite was true. These resultssuggested that HIV/SIVs with/without anti-SAMHD1 activitydiverged at some time point in the past.PHYLOGENY OF RT PROTEINS DERIVEDFROM VIRUSES WITH/WITHOUTSAMHD1-DEGRADING ACTIVITYHuman immunodeficiency virus type 1 does not encode anyanti-SAMHD1 proteins as described above, which appears to bedisadvantageous to the virus for efficient replication in targetcells, especially in non-dividing myeloid and resting T cells.Nonetheless, HIV-1 grows well in humans and is markedlypathogenic for humans. While the mechanism(s) underlyingfor this viral property remains elusive, one could postulate, asa likely possibility, that the high growth ability of HIV-1 isattributable to its relatively high RT activity (Schaller et al., 2014).Indeed, it has been reported that HIV-1 RT is more active insynthesizing viral DNA than the other viral RT proteins derivedfrom viruses with SAMHD1-degrading ability (Lenzi et al., 2014,2015).We therefore constructed a phylogenetic tree of various RTproteins from viruses with/without anti-SAMHD1 activity inorder to predict their evolutional positions as described above(Sakai et al., 2016), and the result was shown in Figure 2.RT proteins from viruses without anti-SAMHD1 activity(viral groups: HIV-1, SIVcpz/gor, SIVmnd-1/lst/sun, andSIVolc/wrc/col) and those from viruses with the activity (viralgroups: SIVrcm/mnd-2/drl, SIVagm, SIVmus/gsn/den/mon,SIVdeb/syk/tal/asc, HIV-2, and SIVsmm/mac/mne/stm)separately formed clusters as observed in the Vpx/Vpr tree(Figure 2). Importantly, viruses with a high RT activity (Lenziet al., 2014, 2015) were confined to the HIV-1 cluster withoutSAMHD1-degrading ability. In contrast, viruses with a lowRT activity (Lenzi et al., 2014, 2015) were found in variousFrontiers in Microbiology | www.frontiersin.org 2 October 2016 | Volume 7 | Article 1655fmicb-07-01655 October 14, 2016 Time: 13:1 # 3Sakai et al. Phylogenetic Insights into RT and Vpx/VprFIGURE 1 | Phylogeny of Vpx/Vpr proteins derived from various human immunodeficiency viruses/simian immunodeficiency viruses (HIV/SIVs).Phylogenetic tree based on diverse Vpx/Vpr proteins was constructed as previously described (Sakai et al., 2016), and is shown together with SAMHD1 (sterile alphamotif and HD-domain containing protein 1)-degrading activity. Viral clones with/without SAMHD1-degrading activity (Laguette et al., 2012; Lim et al., 2012) areindicated by blue and red letters, respectively. Note that Vpr proteins from HIV-2 type viruses (HIV-2, SIVsmm/mac/mne/stm, and SIVrcm/mnd-2/drl) are excludedfrom this tree for simplicity and clarity. Branches were calculated from 1000 bootstrap replicates, and the bootstrap values are labeled on the major branches. Scalebar represents the genetic distance. For virus designations, origins, and types, see the text (Introduction). Amino acid sequences were obtained from the HIVsequence database at Los Alamos National Laboratory (http://www.hiv.lanl.gov) or from the GenBank (http://www.ncbi.nlm.nih.gov).virus clusters with SAMHD1-degrading ability (SIVagm, HIV-2,SIVmac, and SIVmne). While not included in this phylogenetictree due to the sequence unavailability to us, two more HIV-1strains (92RW and 93IN) and two other HIV-2/SIVmne strains(Rod10 and 170) were reported to have RT proteins with a highand low enzymatic activity, respectively (Lenzi et al., 2014).Together with the result in Figure 1, our phylogenetic tree here isconsistent with a hypothesis that the ability of Vpx/Vpr proteinsfrom various HIV/SIVs to degrade SAMHD1 and the differentenzymatic activity of RT proteins from various HIV/SIVs areintimately linked.CONCLUDING REMARKSPrimate lentiviral diversification is frequently accompanied byfunctional alterations of viral proteins required for efficient viralreplication in different cellular environments, which can provideFrontiers in Microbiology | www.frontiersin.org 3 October 2016 | Volume 7 | Article 1655fmicb-07-01655 October 14, 2016 Time: 13:1 # 4Sakai et al. Phylogenetic Insights into RT and Vpx/VprFIGURE 2 | Phylogeny of reverse transcriptase (RT) proteins derived from various HIV/SIVs. Phylogenetic tree based on diverse RT proteins wasconstructed as previously described (Sakai et al., 2016), and is shown together with SAMHD1-degrading and RT activities. Viral clones with/withoutSAMHD1-degrading activity (Laguette et al., 2012; Lim et al., 2012) are indicated by blue and red letters, respectively. In addition, viral clones experimentally testedfor RT activity (Lenzi et al., 2014, 2015) are indicated by boldface type letters. Note that RT proteins from the HIV-1 group show a higher enzymatic activity relative tothose from the other viral groups examined (Lenzi et al., 2014, 2015). Branches were calculated from 1000 bootstrap replicates, and the bootstrap values are labeledon the major branches. Scale bar represents the genetic distance. For virus designations, origins, and types, see the text (Introduction). Amino acid sequences wereobtained from the HIV sequence database at Los Alamos National Laboratory (http://www.hiv.lanl.gov) or from the GenBank (http://www.ncbi.nlm.nih.gov).virologically essential informations to understand the inter-/intra-species transmission and adaptation processes. In thisreport, we have described a potential link between the SAMHD1-degrading activity of Vpx/Vpr proteins and the enzymatic activityof RT proteins. Our phylogenetic analyses (Figures 1 and 2) areconsistent with a scenario that HIV-1 without anti-SAMHD1activity may already have or have acquired RT with enhancedenzymatic activity. However, because viral strains experimentallyFrontiers in Microbiology | www.frontiersin.org 4 October 2016 | Volume 7 | Article 1655fmicb-07-01655 October 14, 2016 Time: 13:1 # 5Sakai et al. Phylogenetic Insights into RT and Vpx/Vprexamined for their anti-SAMHD1 and RT activities throughbiochemical and molecular biological approaches are limited sofar, it would be too early to draw a clear conclusion on this issue.Extensive experimental studies are required to obtain decisiveresults.AUTHOR CONTRIBUTIONSYS, AA, and MN designed the research. YS performed thephylogenetic analysis. YS, ND, YM, AA, and MN analyzed anddiscussed the results. YS, AA, and MN wrote the manuscript.FUNDINGThis study is supported in part by a research grant from the JapanSociety for the Promotion of Science (JSPS KAKENHI: GrantNumber JP26293104).ACKNOWLEDGMENTWe thank Ms. Kazuko Yoshida (Department of Microbiology,Tokushima University Graduate School of Medical Science,Tokushima, Japan) for her editorial assistance.REFERENCESBaldauf, H. M., Pan, X., Erikson, E., Schmidt, S., Daddacha, W., Burggraf, M., et al.(2012). SAMHD1 restricts HIV-1 infection in resting CD4+ T cells. Nat. 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Retrovirology 9:105. doi: 10.1186/1742-4690-9-105Conflict of Interest Statement: The authors declare that the research wasconducted in the absence of any commercial or financial relationships that couldbe construed as a potential conflict of interest.The reviewer AK and handling Editor declared their shared affiliation, and thehandling Editor states that the process nevertheless met the standards of a fair andobjective review.Copyright © 2016 Sakai, Doi, Miyazaki, Adachi and Nomaguchi. This is an open-access article distributed under the terms of the Creative Commons AttributionLicense (CC BY). The use, distribution or reproduction in other forums is permitted,provided the original author(s) or licensor are credited and that the originalpublication in this journal is cited, in accordance with accepted academic practice.No use, distribution or reproduction is permitted which does not comply with theseterms.Frontiers in Microbiology | www.frontiersin.org 5 October 2016 | Volume 7 | Article 1655

霊長類レンチウイルスの分岐・進化：アクセサリー蛋白質Vpx/Vprに基づく系統樹解析

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霊長類レンチウイルスの分岐・進化：アクセサリー蛋白質Vpx/Vprに基づく系統樹解析

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