Journal of General Virology (1995), 76, 1801-1806.
1801
Printed in Great Britain
The complete nucleotide sequence of the RNA 3 of lilac ring mottle
ilarvirus
S. W. Scott* and Xin G e t
Department of Plant Pathology and Physiology, Clernson University, Clemson, SC 29634-0377, USA
The nucleotide sequence of lilac ring motile ilarvirus
(LRMV) RNA 3 consists of 2287 nucleotides and
contains two open reading frames (ORF). The first
encodes a putative translation product of 285 amino
acids (Mr 31308) and the second encodes a putative
translation product of 206 amino acids (Mr 22 751). The
3' terminal nucleotides can be folded into a loop
structure similar to models proposed for other
ilarviruses, although the last four nucleotides are UCGC
not AUGC. The absence of the terminal AUGC motif in
both LRMV and two isolates of apple mosaic ilarvirus
(ApMV) provides circumstantial evidence which
confirms the importance of AUGC motifs upstream of
the terminal AUGC in the protein binding function
associated with these models. Although the 3' terminal
structure of LRMV exhibits similarities to that of ApMV,
comparison of the putative translation products of the
two ORFs with similar products for other ilarviruses
showed greatest identity with citrus leaf rugose (CiLRV)
and citrus variegation ( C W ) ilarviruses both of which
are members of subgroup 2 of this genus. Thus it is
proposed that LRMV be reassigned to subgroup 2 rather
than remaining in its current subgroup, 7, or being
reassigned to subgroup 3 which contains ApMV.
Lilac ring mottle virus (LRMV) was first described and
characterized by van der Meer et al. (1976). It shares
some of the properties of the members of the genus
Ilarvirus, namely: four RNAs with M r values of 1.2, 1"1,
0-9 and 0-4x 10~ and irregularly shaped isometric
particles with some heterogeneity in size. The virus is
strongly immunogenic but in serological tests did not
exhibit any relationship with prunus necrotic ringspot
(PNRSV), tobacco streak (TSV), elm mottle (EMV) and
apple mosaic (ApMV) ilarviruses (van der Meer &
Huttinga, 1979).
Zuidema & Jaspars (1984) have proposed a model for
the secondary structure of the 3' terminal regions of TSV
and the closely related alfalfa mosaic virus (A1MV). In
this model, hairpin (stem-loop) structures are stabilized
by the presence of AUGC motifs with the terminal four
nucleotides being AUGC. This secondary structure was
proposed as being involved in the 'genome activation'
common to both ilarviruses and A1MV in which the
presence of either RNA 4 or the coat protein of the virus
is required to initiate infection (van Vloten-Doting,
1975). In a subsequent report the 97 nucleotides at the 3'
terminus of LRMV were described and shown to form a
structure similar to that found in TSV and A1MV (Bol et
al., 1985). However, the four nucleotides at the 3'
terminus of LRMV were ACGC rather than AUGC.
Two recent reports have examined extensively the
secondary structure at the 3' terminus of A1MV and its
involvement in protein binding. Houser-Scott et al.
(1994) concluded that the coat proteins of both A1MV
and ilarviruses ' recognize invariant AUGC sequences in
the context of conserved structural elements'. Moreover,
it appeared that it is the penultimate AUGC motif (nt
865-868 in RNA 4) which is essential for protein binding
in A1MV. Reusken et al. (1994) identified 'a minimum of
two specific binding sites for CP (coat protein) near the
3' end of RNA 3 '. Site 1 was proximal to the 3' terminus
and included two AUGC motifs but not the terminal
AUGC. The AUGC motifs in this site therefore
corresponded to the penultimate and antepenultimate
A U G C motifs in the Houser-Scott model. Site 2 was
upstream of site 1 and also contained two AUGC motifs.
Mutation of individual AUGC motifs to AGGC had
different effects on the infectivity of RNA 3. Mutation of
the terminal AUGC had little effect, whereas, mutation
of the penultimate site strongly reduced infectivity and
mutation of the antepenultimate site abolished infectivity. Mutation of the AUGC motifs in site 2 also had
little effect on infectivity of RNA 3.
In comparing the A1MV sequence with published
sequences of ilarviruses [TSV - Cornelissen et al. (1984);
* Author for correspondence. Fax + 1 803 656 0274.
I" Present address: Department of Biological Sciences, University of
South Carolina, Columbia, SC 29208, USA.
Sequence data from this article have been deposited with the
EMBL/GenBauk databases under accession no. U17391.
0001-3028 © 1995 SGM
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CAUGGGAGAAAUUI/C CGGGGUGUUACGUGACAACAUCGUUGC
M
G
E
I
S
G
V
L
R
D
N
I
V
A
CUUUCAGAGGUUGUGUACCUGUCGAAGCAAAGAUUUCAAAGGGAGGAGCU
F
R
G
C
V
P
V
E
A
K
I
S
K
G
G
A
401
451
GAAGAAGAAGGUCAGAAAAUUGAUCAAUGUUGAC
K
K
K
V
R
K
L
I
N
V
D
UUUGUUACGUUC C CAGGAUIKTUGCGUUCUACCUCGUGUAGCGAUUCAUGU
C
Y
V
P
R
I
L
R
S
T
S
C
S
D
S
C
UUCUUAUUGAAUAAAGCUACACUTJGAGAAGAUUC CCCUUGGAG~CC
F
L
L
N
K" A
T
L
E
K
I
P
L
G
V
F
UCUAAAUGAGAUGUUUUUCGUUCGUACUGGUUGGCCUAGAUCUUUGAUGA
L
N
E
M
F
F
V
R
T
G
W
P
R
S
L
CUAAGGACGUGAUAGAUGGGAAAGGUCUUUGCCUUACACAUCAAAUCAUU
K
D
V
I
D
G
K
G
L
C
L
T
H
Q
I
I
G C U C C U A C U U U A C C G G U C G G A U G U U C C G C A G G U C G U I / G G U U A C CUULUgUG
A
P
T
L
P
V
G
C
S
A
G
R
W
L
P
F
W
GGAAGAAGACUUCGGUCUCAAGAUGACGUACCAAAAGGAUGUGC
E
E
D
F
G
L
K
M
T
Y
Q
K
D
V
C ULK/UAAGA C C A A A U C U G A G U C U U U G G U G A A G G A U G CGAU-UUC U G A A G C A
F
K
T
K
S
E
S
L
V
K
D
A
I
S
E
A
GUG CGAGAUUCGUUAAUGGCUGGACUUGUC
V
R
D
S
L
M
A
G
L
V
551
601
651
701
751
801
851
901
951
T
C CAUCA
I
T
UGUUGAUCGAUUIIUACCACUCAAA.AGAGUGAGGCCA.AUGIA.AGAUGUUGGA
L
I
D
F
T
T
Q
K
S
E
A
N
E
D
V
G
1051
1100
1050
1000
950
900
850
800
750
700
650
600
550
500
450
400
200
250
300
350
50
100
150
GUACUCCD-O-JGGGAGUACUUCGACGAACUAUGUCUAUUACCAGACCUUCG
Y
S
F
G
S
T
S
T
N
Y
V
Y
Y
Q
T
F
1551
GACACAGGUUC CUCUCGUC CAAAUAUUGUGGACUUGAGAAGGUUUUUC C C
H
R
F
L
S
S
K
Y' C
G
L
E
K
V
F
P
G CUGGGACAGUGGCUUCUGAUCUCAAUGGAUC C CAUGCUAUUI/UAUGGGU
A
G
T
V
A
S
D
L
N
G
S
H
A
I
L
W
V
AAUUGAUG CAG CCUUUCC CAGUACGAUGAACG CUAAUAGUGGUAUCAAGG
I
D
A
A
F
P
S
T
M
N
A
N
S
G
I
K
V
U U C A U U C CA[K/UGGUGG C A G A C U G C C A I i A C U G C C U C C A A U G A A G C C C C C G
H
S
I
W
W
Q
T
A
K
L
P
P
M
K
P
P
CAGAAUUUCUUACAAUGUGAGAAAUGACCACACUCGGGGAGUCGUUGUCU
Q
N
F
L
Q
C
E
K
*
UGUUUGUGGGACCUCUUGGUCUCUGCAGACUUAUACUUAUCUUGAAGAUU
UUUCUCUCCGAGAUGAAiK~CCUGCUAUGUGGAGCAGA~GAAAUCGGA
CUGA[K/UGUCCGUCAUUCAAA~C
CUCUGGGGUGUACUCAGUACC CUUG
UI/I/UGGAUGAGUI/UGUAUI/CC G A C U G U U U G U U U C U C C C A G U C G A U G C C U G
U~JGGUGUUUACAGACGAGUAUGGUAUUGACCAUAUGUUC CUAUUCUCUCU
CUCAGGAGAGGAGAAUAGAUGCCUC CAAAGGAGUCGC
1751
1801
1851
1901
1951
2001
2051
2101
2151
2201
2251
UGAUGAAG CUUUCGAUUCUAC CGCGCUCA-AAGAGC CUGAAGCACCGAAUC
D
E
A
F
D
S
T
A
L
K
E
P
E
A
P
N
R
U A U U C U G U U C U G A U U G G CUl/UAC C G U U A A A G C U G A U G G U ~ J A U C C C G G C U U
Y
S
V
L
I
G
F
T
V
K
A
D
G
Y
P
G
F
1651
1701
AGGCUGAACUGAAGAAGUUUCCAGAUUUAAUACACGCUAACACAGUGGUG
A
E
L
K
K
F
P
D
L
I
H
A
N
T
V
E
A
1601
V
UALrUCUCUACCGGUUCAAUUUCCGGGUUCUGAGUGGCAUAAGALrU-UCCGG
Y
S
L
P
V
Q
F
P
G
S
E
W
H
K
I
S
G
G
1501
CUAUGCAGGCGACUCAGcCuAuGAUUGGUUCCAUCCCUGUUGCUUUGGGA
M
Q
A
T
Q
P
M
I G
S
I
P
V
A
L
1451
N
GAGUAUTJGUGCGUAUGCGGCUUUGCCG CGCGGCAAUACGCGLrUGUAUGCC
GUAGUGUGUCUAUACUUCUl/ACUGACUCAAUCGUCAACAUGAAUG CUCAA
G G G A A U G C C U U C U G U C A G C U U U G U G G U G G U G C U A A A G U C C CGAAUI/UGGG
AAGAAGAUGUCAAC CGCCGUGGUGCGGGUUCUACC CCGCGGCGUGGGAAA
M
S
T
A
V
V
R
V
L
P
R
G
V
G
K
1201
1251
1301
1351
ACCCACCCAACGUUCACGUAACUUCGCUGCACAACGGCAGCGGAGUAACG
P
T
Q
R
S
R
N
F
A
A
Q
R
Q
R
S
GUCAGUUGUUGAGGGGCGUGG CACCGGAAAUUCGGC CCAAACAC CGUUAU
S
V
V
E
G
R
G
T
G
N
S
A
Q
T
P
L
*
1151
1401
UCCACUCUCC CGCGUGUUI/UGUUGCAAUCGAACGGAGCUAGUGUUGUUGA
S
T
L
P
R
V
L
L
Q
S
N G
A
S
V
V
E
1101
2050
2100
2150
2200
2250
2287
2000
1950
1900
1850
1800
1750
1700
1650
1600
1550
1500
1450
1250
1300
1350
1400
1200
1150
Fig. 1. Complete nuclotide sequence of LRMV. Putative translation products for the two O R F s are shown beneath the sequence. The repeated nucleotides in the 5' U T R are double
underlined. • - - - • indicates the location of the primer (nt 127-146) that was used to determine the 5' terminus of the sequence. A U G C motifs that occur in the 3' U T R are in bold
type and underlined.
G C U I / U C U G A U U C G A A G U U G U U G U C A C C G A U U C C CGGGAAU-UCAC C C G A A G
L
S
D
S
K
L
L
S
P
I
P
G
N
S
P
E
V
1001
CAGAAUUCUUCUCAAGUUCU
Q
N
S
S
Q
v
L
P
M
P
CAUCGUAACAUAUAUC
H
R
N
I
Y
L
CAAG CCUGGGAUCU CUGUU CAAAGGAUUUCUCAGUGUUUG CAACAUCUUG
Q
A
W
D
L
C
S
K
D
F
S
V
F
A
T
S
W
501
CUCUACGA
S
T
T
U A C U G U G U G G A A U I / U G G A A U A U A U A U CAC.A.C.A.G.A.A.U.A.C.A.C.U.U.C. .C.C.G U•G U C G
AAUCUCGACAGGAGAUAUAUACCACGUG CUUCUCAC CCAAAUCGA~.AGUG
AGGCAGUGAGAUACGUGAAUAUAUACGAGCUAGUC CUI/AAAAUCGCUAGG
AUUAGUUACCGAAGUUUCUUCGAAGGAAUACCAACAUAUCGAUCAUGGCU
M
A
CUCACUACAUUCAAGAAGAUCACCUACGAAGGAAAAGAUUGGGACUCCCU
L
T
T
F
K
K
I
T
Y
E
G
K
D
W
D
S
L
151
201
251
301
351
GGAUUC C~GAGCAUAC
CGAAUAUAUUCACUGUUUUACUAGACGA
UACGCUAGUUCUGAAGUUUGUUAACUUGGCGACAGAACUUUUGUUC
CCUC
UAC CUUGU CAACU CGGCGGAUACUGAGGACGAAUI/U CAU CGCUAG CGUGU
1
51
i01
c%
Short communication
prune dwarf virus (PDV) - Bachman et al. (1994); citrus
leaf rugose virus ( C i L R V ) - G e & Scott (1994)] and
unpublished sequence data from this laboratory for the
ilarviruses citrus variegation virus (CVV), EMV, spinach
latent virus (SLV) and Parietaria mottle (ParMV) virus,
it became apparent that similar structures with an
AUGC motif at the 3' terminus and a second AUGC
motif (10-14 nt upstream) existed in all cases (HouserScott et al., 1994). The small fragment of sequence of
LRMV (Bol et al., 1985) in which a terminal AUGC was
lacking was a notable exception to this. Subsequently,
two sequences for ApMV were published in which
AUGC motifs that stabilize the stem-loops are present
but the four terminal nucleotides were AAGC (S~inchezNavarro & P~illas, 1994) and GAGG (Alrefai et al.,1994).
In this paper we describe the complete sequence of the
RNA 3 of LRMV and confirm that a 3' terminal
structure similar to that described for other ilarviruses
and A1MV can be formed. The absence of a terminal
AUGC motif might suggest that this sequence most
closely resembles that of ApMV. However, comparisons
of the putative translation products of the two ORFs of
the RNA 3 with similar products of other ilarviruses
show that LRMV is most closely related to the subgroup
2 ilarviruses.
LRMV and antisera to the virus were the kind gift of
Dr D.Z. Maat, Wageningen, The Netherlands. The
virus was grown in Chenopodium quinoa and purified
according to van der Meer et al. (1976). Details of the
procedures used for cloning and sequencing have been
described previously (Ge & Scott, 1994; Scott & Ge,
1995). In essence, RNA was extracted from purified
virions, polyadenylated, and cDNA was synthesized
using oligo(dT)12_ls. The cDNA was cloned into the
EcoRV site of pBluescript II SK(+) (pSK) plasmid
vector (Stratagene). DNA was sequenced using an
automated DNA 373A sequencer.
A single large clone, pLRMV-7, of approx. 2 kb was
obtained and was sequenced completely in both
directions using a combination of subcloning and
synthetic oligonucleotide primers designed from the
internal sequence. The 5' end of the sequence was
determined directly from the RNA using the oligonucleotide 5' GCTAGCGATGAAATTCGTCC 3' as a
primer.
Fragments of sequence were assembled using the
software program GeneJockey and sequences were
aligned and analysed by using the GAP, PILEUP and
PRETTY procedures of the GCG software package
(version 7).
The complete sequence of LRMV RNA 3 (Fig. 1) is
2287 nt in length and contains two ORFs. These code for
putative translation products of 285 amino acids (Mr
31308) and 206 amino acids (M r 22751), respectively.
1803
Table I. Percentage identity (similarity) among
deduced translation products of either ORF 1
(movement protein) or ORF 2 (coat protein) of RNA 3
of L R M V , CVV, CiLRV, PDV, T S V and A l M V
LRMV
ORF 1
ORF 2
TSV
CiLRV
CVV
PDV
ApMV 1
ApMV z
26"8 (44.1)
45-9 (656)
49.2 (64.6)
24-4 (44.8)
23.2 (45.9)
42.4 (57.0)
39.1 (55.0)
22-2 (45,4)
22.5 (44.0)*
19.7 (40.4)*
A1MV
24"0 (455)
21-9 (42'8)
* These comparisons were made using the translation products from
the sequence for the RNA 4 of these viruses published by IS~inchezNavarro & P~illas (1994) and ~Alrefai et al. (1994).
The 5' untranslated region (UTR), the intergenic region
and the 3' UTR are 344 nt, 153 nt and 309 nt in length,
respectively. The 3' UTR contains five AUGC motifs the
last of which occurs at positions 2269-2272. Comparing
the terminal 97 nucleotides of our sequence for LRMV
with that of Bol et al. (I985) indicated only two
differences: A instead of U at position 2284 and G
instead of A at position 2232. The terminal nucleotides
of the 3' UTR of Our sequence can be folded to form a
stem-loop structure that exhibits only a minor difference
from the structure previously proposed for the 3'
terminus of LRMV by Bol et al. The original structure
has a bulge on the 3' arm of the second loop whereas in
our structure a bulge occurs on the 5' arm of the same
loop.
Comparisons of the putative translation products of
both ORF 1 and ORF 2 with similar products from other
ilarviruses and A1MV (Table 1) indicate that LRMV
shows the greatest identity (39-49 %) for both ORFs
with CiLRV and CVV. With other ilarviruses a n d
A1MV, the identities are between 19 % and 27 %. When
the products of ORF 1 (putative movement protein) for
LRMV and the two citrus viruses were aligned and a
consensus sequence generated (Fig. 2) there were regions
of as many as nine amino acids which were conserved in
all three proteins. Adopting a similar procedure for the
products of ORF 2 (coat protein) showed no such
conserved areas. However, if a simple multiple sequence
alignment were made using the procedure PILEUP then
areas where three or four amino acids were conserved in
all three proteins could be found (Fig. 3). These areas
tended to occur most frequently in the middle of the
proteins.
The sequence of the RNA 3 of LRMV shares many of
the characteristics reported for other ilarviruses. The 5'
and 3' UTRs and the intergenic region are all comparable
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LRMV
CiLRV
CW
LRMV
CiLRV
CVV
LRMV
CiLRV
CVV
LRMV
CiLRV
CVV
LRMV
CiLRV
CW
LRMV
CiLRV
CW
LVQNSSQVLL
LLERTMNAQR
LLERTLNAAK
LLERT-NA--
201
EAVRDSLMAG
NDVAASLMNS
NDVAASLMNS
NDVAASLMNS
292
AQTPL* ......
K G R G Q N T P T P *.
HNTPK* ......
I~fE. . . . . . D
RA ....... S G--PE-MPDF
SDSKLLSPIP
RAID.VGLVS
RACTTGLISS
250
TTQKSEANED
TTEIVKVTKD
TIEELPESDS
NEE-VRDAIS
GNSPEVLIDF
SGEEQMMPDF
GEQPE.MPDF
KDV-A .... R
EDFGLPMVYO
200
SESLVKDAIS
NEESVRDAIS
NEETVRDAIS
CiLRV
CW
LRMV
CiLRV
CW
LRMV
228
PGIKPPPNFL VCEMDDK*
PGVKPPSNFL VVEE*...
PPMKPPQNP 5 QCEK*...
201
TKFYVSTVPL
TKFYVSTTPL
HSIWWQTAKL
Fig. 3
RQLVLPPGST
KQQVFPTGTT
LEKVFPAGTV
151
IRVKAAKYCA
IRVKRGKYCA
HRFLSSKY=~
VWEFDAA..P
VWDFDTA..P
LWVIDAAFPS
200
AAGTANIISV
ATGAVHEISI
TMNANSG~KV
150
NATGPVAPNR
NATGPVAPDR
ALKEPEAPNR
101
TELGKIKSL. HHTTKVYSVM YGFTCKADGY
S E L S K I R T L . I{DTTKVYSVM I G F V C K S D G Y
AE__LLK~FPD~I ~ S V L
IGFTVKADGY
CiLRV
CVV
LRMV
VADLRDNYNF
VSEVKANWNL
ASDLNGSHAI
I00
SGFSFPDAWG SAKIAYASMR
SGYSFPDRWG SGTIAYMTLR
SGY,~__FGST.S T N Y V Y ~ Q T F E
51
QPQRMVVGSM PIDTITWKS..FPGEQWHEF
QPQQLLVGSM PTNLPTWKS..FPGEQWIIEV
~ATQPMIG~_SI ~ V A L G Y S L P V Q F ~ S E W ~ d K I
CiLRV
CW
LRMV
AGFMDNFDSA
AGFMDGFDVN
PGFDEAFDST
5O
R R P T N R S R N W A Q G .... QRS
RQPTARSRQW AQGLANVRRS
GKP___~RS____~F _AAQRQRSNAM
1
MA_NRSNAIEV N G V W Y N R A D N A P V A N A R . G R
..MSGNAIEI NGRWYQPAPN SAPTRGRGGR
................. MST AVVRVLPRGV
CiLRV
CVV
LRMV
Fig. 3. Comparison of the putative translation products of ORF 2 of LRMV, CiLRV and CVV. The sequences were aligned using the PILEUP procedure. Amino acids which occur
in all three proteins are in bold type and underlined.
Fig. 2. Comparison of the putative translation products of ORF 1 of LRMV, CiLRV and CVV. The individual sequences were adjusted in length using the GAP procedure and then
aligned and the consensus sequence produced using the PRETTY procedure. Amino acids in the consensus sequence which occur in all three proteins are in bold type and underlined.
Fig. 2
V V E . . . . . . _S - - T P . . . . . . . .
SVGRWIPFWE
FIAPTLPPDC
KDVPITFKTK
KDVTADMAIR
KDVAASLSLR
EDFGLKMTYQ
EDFGLPMVYQ
EDFGLPMVYQ
-G-TAN ..............
SAGRWLPFWE
SVGRWIPFWE
SVGRWIPFWE
151
IIAPTLPVGC
FIAPTLPPDC
FIAPTLPPDC
-HKGLFL~SIS
N]fMFILKTGW P R S L R V K D V L
VVEGRGTGNS
GRYEDLSTAS
WETAMEAQS
EKKPVGVLPM
RCML-NAATM
150
DGKGLCLTHQ
AHKGLFLTHS
NHKGIFLTHS
PRSLMTKDVI
PRSLRVKDIL
PRSLRVKDVL
NEMFFVRTGW
NKMFILKTGW
NKMFILKTGW
i00
RILRSTSCSD
RILSDTACTD
RILHDTSCTD
251
VGSTLPRVLL QSNGASWES
RID2LANTDNT F G K V I . . S G D
GGMTANATGE KKSTVVSGNG
EKIPLGVFPL
EKKPVGLLPM
EKKPVGVLPM
I-IRI~IYLIYIP R I L - D T S C T D
K-VKKMINVS
DFSVFA~"KWN
---AWELVSK
101
SCFLLNKATL
RCMLINAATM
RCMLVNAATM
HRNIYLCYVP
HRNIYLIYIP
HRNIYLIYIP
KKVRKLINVD
KMVKKMINVS
RFTKKMINVS
DFSVFATSWK
DFSVLATKWN
DFSVFATKWN
51
GAQAWDLCSK
NS.AWELVSK
S..AWELVSK
-P-E_A-I-K-
VADSSTFRGC
M A L S - FK- IS VE- KI)F- S L M N E V C G V M R E H
50
VPVEAKISKG
LSAKEALIKA
QPKEALIKAN
IVASTTFRGC
VADSST..SG
VADSSTFRGC
YEGKDWDSLM
VEHKDFLSLM
VEQKDFTSLM
GEISGVLRDN
NEVCGVMREH
NEVCGVMREH
1
MALTTFKKIT
MALSSFKAIS
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Short communication
in size to those described for the R N A 3 of TSV
(Cornelissen et al., 1984), PDV (Bachman et al., 1994),
and CiLRV and CVV (Scott & Ge, 1995). The sizes of
the putative translation products of the two ORFs are
also in good agreement with those reported for other
ilarviruses. In the 5' UTR, repeats of eight nucleotides
( G A A U A U A U ) occur. Three repeats of 15 nucleotides
have been reported in the 5' U T R of PDV (Bachman et
al., 1994) and three or four repeats of a 27-30 nucleotide
sequence have been reported in the 5' UTRs of the
strains of AIMV which have been sequenced (Langereis
et al., 1986). In contrast TSV, CiLRV and CVV contain
no repeated sequences in the 5' U T R longer than five
nucleotides (Cornelissen et al., 1984; Scott & Ge, 1995).
The repeats in PDV and A1MV are U-rich and share
52 % identity (Bachman et al., 1994). However, although
the repeats in L R M V are not U-rich they do bear a slight
resemblance to the repeated sequences in the other two
viruses.
Clearly the structures at the 3' terminus of L R M V are
similar to those reported for other ilarviruses. However,
as the last four nucleotides at the 3" terminus are U C G C
not A U G C , L R M V differs from the majority of
ilarviruses for which sequence data are available
(Houser-Scott et al., 1994) and most closely resembles
the sequences for ApMV (S~inchez-Navarro & Pfillas,
1994; Alrefai et al., 1994) in this respect.
Huttinga & Mosch (1976) demonstrated that L R M V
requires the presence of either coat protein or R N A 4
in order that infection can occur. However, there is no
report in the literature of L R M V protein being able to
cross-activate other ilarviruses. Gonsalves & Fulton
(1977) showed that an isolate o f ApMV (referred to by
them as rose mosaic virus) was coat protein-dependent
and could also cross-activate PNRSV. Thus the necessity
for protein binding in these situations, plus the absence
o f the terminal A U G C motif in L R M V and the two
isolates of ApMV, provide indirect confirmation that it is
the A U G C motifs upstream of the Y-terminal A U G C
which are essential for protein binding (Houser-Scott
et al., 1994; Reusken et al., 1994).
While the similarities in the Y-terminal structures
would suggest that L R M V is closely related to ApMV
the relationships of the movement protein and the coat
protein would not support this. The subgroupings in the
genus Ilarvirus are based on serological relationships
(Francki et al., 1991) and while there is a clear
relationship between CiLRV, CVV and EMV (subgroup
2) and between ApMV and PNRSV (subgroup 3) there
is no relationship between L R M V and any of these
viruses (van der Meer & Huttinga, 1979). Neither have
we been able to demonstrate such a relationship between
either CiLRV or CVV and L R M V despite repeated
serological assays. However, this may not be surprising
1805
when it is considered that the putative coat proteins of
CVV and CiLRV share 65 % identity (Scott & Ge, 1995)
whereas L R M V shares 42.4% and 39"1% identity with
the coat proteins of CiLRV and CVV, respectively.
On the basis of the identity between the putative
translation products of the two ORFs of L R M V and the
two citrus viruses, L R M V should probably be considered
to be a member of the subgroup 2 of the ilarviruses
rather than being placed in a distinct group. If this is
accepted then the recently reported Fragaria chiloensis
ilarvirus (Spiegel et al., 1993) should also be placed in
subgroup 2 as a serological relationship exists between it
and LRMV.
This is technical contribution no. 4016 of the South Carolina
Agricultural Experiment Station. S.W.S gratefully acknowledges the
use of facilities at SCRI, Invergowrie, UK to grow and purify LRMV
while on sabbatical in 1993.
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