Poxvirus Membrane-bound G Protein-coupled Receptor Homologs

6 Pages·0.067 MB·English

by  McFadden G., Moyer R.

Poxvirus Membrane-
bound G Protein-coupled
Receptor Homologs
Grant McFadden1,* and Richard Moyer2
1The John P. Robarts Research Institute and Department of Microbiology and Immunology,
The University of Western Ontario, 1400 Western Road, London, Ontario, N6G 2V4, Canada
2Department of Molecular Genetics and Microbiology, University of Florida College of Medicine,
PO Box 100266, Gainesville, FL 32610-0266, USA
*corresponding author tel: [phone], fax: [phone], e-mail: [email protected]
DOI: 10.1006/rwcy.2000.14017.
SUMMARY to the number of intervening amino acids between
the first two cysteines. Both the classification of
chemokines and their receptors have been reviewed
To date, virus-encoded homologs of G protein-
(Oppenheim et al., 1991; Schall and Bacon, 1994;
coupled receptors (GPCRs) have been discovered
Murphy, 1994; Barker and Monk, 1997). Chemokine
only in members of the poxvirus and herpesvirus
action is most frequently mediated through inter-
families. The herpesvirus members have been more
action with appropriate G protein-coupled receptors
extensively characterized than the poxvirus members,
(GPCRs). While GPCRs have not been detected
but all were initially discovered by database searches
to date in the commonly studied members of the
of predicted open reading frame sequences deduced
Orthopoxvirus genus such as vaccinia, ectromelia, or
during DNA sequencing studies of the various viral
cowpox virus, sequence analysis has revealed poten-
DNAgenomes.Unlike theherpesvirus examples(e.g.
tial chemokine GPCRs within the genomes of swine-
Kaposi’s sarcoma-associated GPCR), the poxvirus
pox virus (the prototypic member of the Suipoxvirus
GPRC homologs have not yet been analyzed for
genus) (Massung et al., 1993) and sheep pox virus, a
biological activity.
memberoftheCapripoxvirusgenus(Caoetal.,1995).
Poxvirusgenomesarelinearterminatingwithregions
comprising inverted terminal repetitions (ITRs).
BACKGROUND
Genes within the terminal repetitions are diploid.
Swinepox virus and sheep pox ORF designations are
Discovery
based on genomic HindIII restriction patterns where
the largest fragment is designated as ‘A’. An analysis
Chemokines are major regulatory components of the of swinepox virus sequence reported two related
immune system, being involved in trafficking, local- ORFs, one being a truncated ORF (C3L) at the
ization, and activation of various leukocyte popula- junction of the left inverted ITR and the left-most
tions. Collectively, chemokines can be classified unique region of the genome. A full-length version of
according to the relative positions of the first pair the same open reading frame (ORF K2R) was found
of a conserved motif of cysteine residues. The desig- at the junction of the right ITR and the right-most
nationsareC,CC,CXC,andCX3C,where‘X’refers unique region of the genome. Most likely, the
2110 Grant McFadden and Richard Moyer
incomplete C3L ORF represents an artifact derived virus-encoded chemokine receptors is the US28 gene
from a variant virus, grown in cell culture, where in of human cytomegalovirus. This protein both binds
the absence of any selection, a partial deletion of the chemokines (Neote et al., 1993) and induces a rise in
gene, mirrored in both copies of the ORF has intracellular calcium after binding of the appropriate
occurred. chemokines (Gao and Murphy, 1994).
The sheep pox ORF (Cao et al., 1995) is located In the absence of data demonstrating functionality
within the HindIII Q2 fragment of the KC-1 strain for the poxvirus proteins, one can only speculate as
and corresponds to ORF 3L (Gershon and Black, to function based on the relatively high homology
1987).LiketheswinepoxvirusORF,theQ2fragment exhibitedtocertainCCchemokinereceptors(Table1).
of the KC-1 strain is located near the right-most That prediction would be that both poxvirus GPCRs
terminal extreme of the sheep pox virus genome, a do indeed bind to chemoattractants, most likely che-
locationtypical ofnonessential genesdevotedtocon- mokines of the CC class. Although superficially, all
trolling the host response to the infection. While the structural features required for signaling follow-
structuralfeaturesofthetwoORFsdiscussedhereare ing receptor–ligand engagement appear to be present
consistent with these two encoded proteins func- in the poxvirus GPCRs, an equally likely scenario,
tioning as GPCRs, functionality has not yet been basedonotherpoxvirusexamplesisfortheseproteins
demonstrated. to function as nonsignaling, ligand sinks.
Structure
GENE
The poxvirus GPCR homologs share all the features Accession numbers
typical of both cellular and viral GPCRs which are
depicted graphically in Figure 1. These features
Sheep pox virus ORF3L: S78201
include: (1) an extracellular N-terminus, (2) an intra-
Swinepox ORF K2R (complete ORF): L21031
cellular C-terminus, (3) seven (cid:11)-helical transmem-
Swinepox C3L (truncated ORF): L22013
branedomains,whichareorientedperpendicularlyto
the plasma membrane, (4) three intracellular and
threeextracellularhydrophilicconnectingloops,(5)a PROTEIN
disulfide bond linking cysteine residues in the first
and second extracellular loops, and (6) the presence
Accession numbers
of proline residues in transmembranedomains II, IV,
V, VI, and VII.
GenBank:
Capripox virus: Q86917
Main activities and
Swinepox virus: Q08520
pathophysiological roles Human CC (CCR8): P51685
Rhesus monkey CC: AAC72403
Equine herpesvirus 2 receptor: S55594
Clearly, the encoding of both chemokines and
Human CXC chemokine receptor (CXCR2): P25025
chemokine receptors by viruses indicates the impor-
Herpesvirus saimiri GPCR: Q01035
tanceofmodulatingchemokineactivityduringcertain
HHV8: AAB51506
viral infections. Indeed, in the case of Epstein–Barr
EBV: P32249
virus, which does not encode a GPCR, the virus
CMV (US28): P09704
inducessynthesisofacellularGPCRduringinfection
(Birkenbach et al., 1993; Schweickart et al., 1994),
presumably to accomplish a similar purpose. Sequence
Viral-encoded GPCRs are relatively prevalent
amongst the herpesvirus, examples being found in
The complete protein sequence of the sheep pox and
human herpesvirus 8 (HHV8) (Guo et al., 1997),
swinepox virus ORFs is shown in Figure 1.
herpesvirus saimiri (Nicholas et al., 1992; Ahuja and
Murphy,1993),humancytomegalovirus(US28)(Chee
Description of protein
et al., 1990), and equine herpesvirus 2 (Telford et al.,
1995)(ORFE1,whichispresentintwocopiesdueto
its location within the terminal repeat region of the The poxvirus-encoded proteins are slightly larger in
viral chromosome). A useful functional paradigm for size than the typical GPCR, but are nevertheless,
Poxvirus Membrane-bound G Protein-coupled Receptor Homologs 2111
Figure1 Alignmentofputativepoxvirus-encodedGprotein-coupledreceptors(GPCRs)withthoseofviralandcellular
origin.Eachproteincontainsatypicalseventransmembranemotifsignature,indicatedabovethesequencesinwhichthey
occur. EachGPCRalso containsaconservedproline residuewithin transmembranedomainsII,IV,V, VI,andVIIand
two conserved cysteine residues, one in the extracellular region between transmembrane domain II and III, the second
within the extracellular region between transmembrane regions IV and V. Typically, these two cysteines are linked in a
disulfide bridge. Both the proline and cysteine residues are indicated as shaded residues within the consensus sequence.
EachproteinalsocontainsthehighlyconservedmotifDRYLAIVHAattheendofthethirdtransmembranedomain.The
DRYLAIVHAmotifisnotuniversallypresent,beingabsentintheCMVUS28protein.AnotherfeatureofGPCRsisthe
presenceofphosphorylationsites,typicallyserineresiduesintheC-terminalportionofthemoleculeandglycosylationsites
withintheextracellularN-terminalmostregionofthemolecule.TheproteinsdepictedarethoseofCPV(capripoxvirus),
SPV(swinepoxvirus),HuCC(humanCCchemokinereceptor),MnCC(rhesusmonkeyCCchemokinereceptor),EQHV
(equineherpesvirus2GPCR).Thenumberofaminoacidsintheproteinisgiven.Thealignmentsandconsensussequence
were derived using the PRETTY program of the Genetics Computer Group Package, Madison, Wisconsin.
1 70
CAPRI MNYTLSTVSS ATMYNSSSNI TTIATTIIST ILSTISTNQN NVTTPSTYEN TTTISNYTTA YNTTYYSDDY
SPV ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~MTSPT NSTMLTYTTN NYYDDDYYEY STITDYYNTI
EQHV ~~~~~~~~~~ ~~~~~~~~~~ ~~MATTSATS TVNTSSLATT MTTNFTSLLT SVVTTIASLV PSTNSSEDYY
MOUSE ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~MDYTME PNV.TMTD.Y
MONKEY ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~MDYTLD PSMTTMTDYY
HUMAN ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~~~~~~~ ~~~~MDYTLD LSVTTVTDYY
CONSEN ---------- ---------- ---------- ---------- ---------- ---------- ----------
I II
71 140
CAPRI DDYEVS...I VDIPHCDDGV DTTSFGLITL YSTIFFLGLF GN.IIVLTVL RKYKIKTIQD MFLLNLTLSD
SPV NNDITSSSVI KAFDNNCTFL EDTKYHIIVI HIILFLLGSI GNIFVV.SLI AFKRNKSITD IYILNLSMSD
EQHV DDLDDVDYEE SAPCYKSDTT RLAAQVVPAL YLLVFLFGLL GNILVVIIVI RYMKIKNLTN MLLLNLAISD
MOUSE ..YPDF...F TAPCDAEFLL RGSMLYLAIL YCVLFVLGLL GNSLVILVLV GCKKLRSITD IYLLNLAASD
MONKEY ..YPDS...L SSPCDGELIQ RNDKLLLAVF YCLLFVFSLL GNSLVILVLV VCKKLRNITD IYLLNLALSD
HUMAN ..YPDI...F SSPCDAELIQ TNGKLLLAVF YCLLFVFSLL GNSLVILVLV VCKKLRSITD VYLLNLALSD
CONSEN ---------- ---------- ---------- ----F----- GN-------- ---------- ---LNL--SD
III
141 210
CAPRI LIFVLVFPFN LYDS.IAKQW SLGDCLCKFK AMFYFVGFYN SMSFITLMSI DRYLAVVHPV KSMPIRTKRY
SPV CIFVFQIPFI VY..SKLDQW IFGNILCKIM SVLYYVGFFS NMFIITLMSI DRYFAIVHPI KRQPYRTKRI
EQHV LLFLLTLPFW MHYIGMYHDW TFGISLCKLL RGVCYMSLYS QVFCIILLTV DRYLAVVYAV TALRFRTVTC
MOUSE LLFVLSIPFQ THNL.L.DQW VFGTAMCKVV SGLYYIGFFS SMFFITLMSV DRYLAIVHAV YAIKVRTASV
MONKEY LLFVFSFPFQ TYYQ.L.DQW VFGTVMCKVV SGFYYIGFYS SMFFITLMSV DRYLAVVHAV YAIKVRTIRM
HUMAN LLFVFSFPFQ TYYL.L.DQW VFGTVMCKVV SGFYYIGFYS SMFFITLMSV DRYLAVVHAV YALKVRTIRM
CONSEN --F----PF- ---------W --G---CK-- ---------- ----I-L--- DRY-A-V--- -----RT---
IV V
211 280
CAPRI GI.VLSMVVW IVSTIESFPI MLFYET..KK VYGITYCHVF YND.NAKIW. KLFINFEINI FGMIIPLTIL
SPV GIL.MCCSAW LLSLILSSPV SKLYENIPHM SKDIYQCTLT NENDSIIAFI KRLMQIEITI LGFLIPIIIF
EQHV GIVT.CVCTW FLAGLLSLPE FFFHGH..QD DNGRVQCDPY YPEMSTNVW. RRAHVAKVIM LSLILPLLIM
MOUSE G.TALSLTVW LAAVTATIPL MVFYQV..AS EDGMLQCFQF YEE.QSLRW. KLFTHFEINA LGLLLPFAIL
MONKEY GTTTLSLLVW LTAIMATIPL LVFYQV..AS EDGVLQCYSF YNQ.QTLKW. KIFTNFEMNI LGLLIPFTIF
HUMAN G.TTLCLAVW LTAIMATIPL LVFYQV..AS EDGVLQCYSF YNQ.QTLKW. KIFTNFKMNI LGLLIPFTIF
CONSEN G--------W --------P- ---------- ------C--- ---------- ---------- -----P--I-
VI
281 350
CAPRI LYCYYKILNT LKTSQTKNK. KAIKMVFLIV ICSVLFLLPF SVTVFVS... SLYLLNVFSG CMALRFVNLA
SPV VYCYYRIFTT VVRLRNRRKY KSIKIVLMIV VCSLICWIPL YIVLMIATIV SLYTSNIFRH LCLYLNLAYA
EQHV AVCYYVIIRR LLRRPSKKKY KAIRLIFVIM VAYFVFWRPY NVALFLT... TFHATLLNLQ CALSSNLDMA
MOUSE LFCYVRILQQ LRGCLNHNRT RAIKLVLTVV IVSLLFWVPF NVALFLT... SLHDLHILDG CATRQRLALA
MONKEY MFCYIKILHQ LKRCQNHNKT KAIRLVLIVV IASLLFWVPF NVVLFLT... SLHSMHILDG CSISQQLNYA
HUMAN MFCYILILHQ LKRCQNHNKT KAIRLVLIVV IASLLFWVPF NVVLFLT... SLHSMHILDG CSISQQLTYA
CONSEN --CY--I--- ---------- --I------- --------P- ---------- ---------- ---------A
2112 Grant McFadden and Richard Moyer
Figure 1 (Continued)
VII
351 420
CAPRI VHVAEIVSLC HCFINPLIYA FCSREFTKKL LRLRTTSSAG SISIG*~~~~ ~~~~~~~~~~ ~~~~~~~~~~
SPV ITFSETISLA RCCINPIIYT LIGEHVRSRI SSICSCIYRD NRIRKKLFSR KSSSSSNII* ~~~~~~~~~~
EQHV LLITKTVAYT HCCINPVIYA FVGEKFRRHL YHFFHTYVAI YLCKYIPFLS GDGEGKEGPT RI*~~~~~~~
MOUSE IHVTEVISFT HCCVNPVIYA FIGEKFKKHL MDVFQKSCSH IFLYLGRQMP VGALERQLSS BQRSSHSSTL
MONKEY THVTEIISFT HCCVNPVIYA FVGEKFKKHL SEIFQKSCSH IFIYLGRQMP RESCEKSSSC QQHSFRSSSI
HUMAN THVTEIISFT HCCVNPVIYA FVGEKFKKHL SEIFQKSCSQ IFNYLGRQMP RESCEKSSSC QQHSSRSSSV
CONSEN ---------- -C--NP-IY- ---------- ---------- ------~~~~ ~~~~~~~~~~ ~~~~~~~~~~

421
CAPRI ~~~~~
SPV ~~~~~
EQHV ~~~~~
MOUSE DDIL*
MONKEY DYIL*
HUMAN DYIL*
CONSEN ~~~~~
Table 1 Percent identity between poxvirus, various herpesvirus, and cellular-encoded GPCRs
Source % Identity
CPV SPV Hu Mn EQHV Hu IL-8 HSV HHV8 EBV CMV
CC CC CXC
CPV – 37.6 44.8 45.9 31.3 29.4 21.2 17.4 27.6 29.5
SPV 37.6 – 38.7 38.7 30.9 32.1 20.2 17.8 24.4 27.4
Hu CC 44.8 38.7 – 94.3 39.1 37.4 19.1 21.9 25.1 29.8
Mn CC 45.9 38.7 94.3 – 38.7 36.4 18.7 22.9 25.1 30.8
EQHV 31.3 31.0 39.1 38.7 – 34.5 22.8 22.6 27.5 31.8
Hu IL-8 29.4 32.1 37.4 36.4 34.5 – 30.8 29.6 27.6 32.9
HSV 21.2 20.3 19.1 18.7 22.8 30.8 – 35.0 31.2 32.8
HHV8 17.4 17.8 22.0 23.0 22.6 29.6 35.0 – 19.6 21.6
EBV 27.7 24.4 25.1 25.1 27.5 27.6 31.2 19.6 – 27.4
CMV 29.5 27.4 29.8 30.8 31.8 32.9 23.5 21.6 27.4 –
CPV,capripoxvirus(381aa);SPV,swinepoxvirus(370aa);HuCC,humanCCchemokinereceptor(CCR8)(355aa);
MnCC,rhesusmonkeyCCchemokinereceptor(356aa);EQHV,equineherpesvirus2receptor(383aa);HuIL-8,humanCXC
chemokinereceptor(CXCR2)(360aa);HSV,herpesvirussaimiriGPCR(331aa);HHV8,humanherpesvirus8GPCR(352aa);
EBV,cellularGPCRinducedbyEpstein–Barrvirusinfection(361aa);CMV,cytomegalovirusGPCR(US28)(345aa).Identities
werecalculatedusingtheBESTFITprogram(GeneticsComputerGroupPackage,Madison,Wisconsin).
structurally typical of GPCRs (see Figure 1). Begin- extracellular(E1–E3)domains.Extensionofsequence
ning with a glycosylated extracellular N-terminal beyond the seventh transmembrane domain defines
domain,theserpentineseventransmembranedomains an intracellular cytoplasmic tail of the protein
definethreealternatingintracellular(I1–I3)andthree (Murphy, 1994; Barker and Monk, 1997).
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