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Journal of Virology, August 1999, p. 6500-6505, Vol. 73, No. 8
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
A Putative Cell Surface Receptor for
Anemia-Inducing Feline Leukemia Virus Subgroup C Is a Member of a
Transporter Superfamily
Chetankumar S.
Tailor,1,*
Brian J.
Willett,2 and
David
Kabat1,*
Department of Biochemistry and Molecular
Biology, Oregon Health Sciences University, Portland, Oregon
97201-3098,1 and Department of
Veterinary Pathology, University of Glasgow, Glasgow G61 1QH, United
Kingdom2
Received 12 February 1999/Accepted 27 April 1999
 |
ABSTRACT |
Domestic cats infected with the horizontally transmitted feline
leukemia virus subgroup A (FeLV-A) often produce mutants (termed FeLV-C) that bind to a distinct cell surface receptor and cause severe
aplastic anemia in vivo and erythroblast destruction in bone marrow
cultures. The major determinant for FeLV-C-induced anemia has been
mapped to a small region of the surface envelope glycoprotein that is
responsible for its receptor binding specificity. Thus, erythroblast
destruction may directly or indirectly result from FeLV-C binding to
its receptor. To address these issues, we functionally cloned a
putative cell surface receptor for FeLV-C (FLVCR) by using a human
T-lymphocyte cDNA library in a retroviral vector. Expression of the
2.0-kbp FLVCR cDNA in naturally resistant Swiss mouse fibroblasts and
Chinese hamster ovary cells caused substantial susceptibility to FeLV-C
but no change in susceptibilities to FeLV-B and other retroviruses. The
predicted FLVCR protein contains 555 amino acids and 12 hydrophobic
potential membrane-spanning sequences. Database searches indicated that
FLVCR is a member of the major-facilitator superfamily of transporters
and implied that it may transport an organic anion. RNA blot analyses
showed that FLVCR mRNA is expressed in multiple hematopoietic lineages rather than specifically in erythroblasts. These results suggest that
the targeted destruction of erythroblasts by FeLV-C may derive from
their greater sensitivity to this virus rather than from a preferential
susceptibility to infection.
| |
INTRODUCTION |
Feline leukemia viruses (FeLVs)
cause prevalent contagious infections of domestic cats that result in
proliferative, degenerative, and immunosuppressive disorders (16,
17, 32). The viruses recovered from infected cats often contain
mixtures of components that utilize distinct cell surface receptors and
have been classified into the A, B, and C interference groups
(44). FeLV-A strains are horizontally transmitted in saliva,
preferentially infect cat cells, and often cause a slowly developing
immunodeficiency and T-cell lymphoma (16, 17, 32, 36). In
contrast, FeLV subgroups B and C have broader host ranges (17, 21,
32) and are formed from FeLV-A in infected cats, with FeLV-B
being formed by recombination with endogenously inherited retroviral sequences (32, 37, 45, 46) and FeLV-C apparently being formed by mutations (32, 42). Although the receptor genes for FeLV-A and FeLV-C have not been cloned or unambiguously identified, FeLV-A has been shown to bind to a 70-kDa protein that has been proposed to be the cell surface receptor (13). The receptor for FeLV-B, however, has been identified as the sodium-dependent phosphate symporter Pit1 (50), which is also used by gibbon ape leukemia virus (33) and 10A1 murine leukemia virus
(30, 54).
FeLV-C formation in cats has been tightly associated with a fatal pure
erythrocyte aplasia, also known as erythroid aplasia or aplastic anemia
(1, 10, 20, 32, 35, 40-42). This disease, which is specific
for the erythroid lineage and is similar to human aplastic anemia
(9, 23), involves depletion of erythroid CFU (CFU-E) and
burst-forming units (BFU-E) in vivo, and it can be induced in bone
marrow cultures by FeLV-C infections (1, 35, 43, 51).
Studies of cats infected with FeLV-A/FeLV-C chimeras and site-directed
mutants have demonstrated that aplastic anemia is determined
predominantly by a small variable region, vr1, in the envelope
glycoprotein of FeLV-C that also determines the receptor binding
specificity of the glycoprotein (4, 42). These findings
suggest that the aplastic anemia may result from FeLV-C binding to its
receptor, either on erythroblasts or on other cells that control the
bone marrow microenvironment (1, 27, 28). By analogy to
other retroviruses (4, 19, 22, 52), the FeLV-C envelope
glycoprotein might perturb the normal function of its receptor,
resulting in cell killing or in changes in cytokine production. The
observation that FeLV-C can infect other hematopoietic cells, including
myeloid and lymphoid cells (7), implies that erythroblasts
may be either exceptionally sensitive to secondary sequelae of
infection or critically dependent on the normal function of the FeLV-C receptor.
We have addressed these issues by functionally cloning and
characterizing a putative cell surface receptor for FeLV-C (FLVCR). This was done with a human T-lymphocyte cDNA library in a retroviral vector by procedures that have recently been used to clone cDNAs that
encode coreceptors for simian immunodeficiency viruses (8) and receptors for xenotropic and polytropic murine leukemia viruses (3, 48, 55) and for simian type D retroviruses (39,
49). This method has major advantages, in part because full
library representation occurs within relatively small cultures of
naturally resistant murine NIH 3T3 fibroblasts and because each cell in the culture expresses only a small number (usually one or fewer) of
human cDNAs. The cells that expressed the FeLV-C receptor were selected
by infection, and the FLVCR cDNA was cloned from the cellular DNA by
PCR. Our results suggest that FLVCR is a member of a large superfamily
of transporters and that it is most closely related to evolutionary
branches of this superfamily that function to transport the organic
anion(s). These results have important implications for our
understanding of FeLV-induced diseases.
| |
MATERIALS AND METHODS |
Cells and viruses.
Mouse NIH 3T3, human TE671, and Chinese
hamster ovary (CHO) cells were used as target cells for infection.
TECeB15, TELCeB6 (6), and Phoenix-Eco (provided by Garry
Nolan, Stanford University, Stanford, Calif.) are packaging cell lines
that produce replication-defective retrovirus. TECeB15 and TELCeB6
cells do not contain retroviral envelope genes and therefore produce
noninfectious virus. CHO cells were maintained in Dulbecco's modified
alpha medium supplemented with 10% fetal bovine serum (FBS), and
Phoenix-Eco cells were maintained in Dulbecco's minimal essential
medium supplemented with high glucose and 10% FBS. All other cell
lines were maintained in Dulbecco's minimal essential medium
supplemented with low glucose and 10% FBS.
lacZ(FeLV-B), lacZ(A-MLV), and lacZ(RD114) producer cells were provided
by Yasuhiro Takeuchi (Institute of Cancer Research, London, United
Kingdom). lacZ(FeLV-C) pseudotype virus was generated by transfection
(calcium phosphate precipitation [Stratagene]) of TELCeB6 cells with
the FBCsalf vector (FeLV-C Sarma envelope gene [41]
cloned into the FBsalf retroviral expression vector [6]). Transfectants were selected with phleomycin (50 µg/ml), and resistant colonies were pooled 2 weeks after the start of selection. Infection of target cells with lacZ pseudotype virus was
carried out as previously described (47).
Replication-defective FeLV-C pseudotype virus carrying the puromycin
resistance gene, puro(FeLV-C), was generated by transducing TECeB15
cells with replication-defective puro(RD114) pseudotype virus to
introduce the puromycin resistance gene [the puro(RD114) pseudotype
virus was produced from FLYRD18 cells (6) transfected with
pBabe-puro retroviral expression vector (31)]. Transduced
cells were selected with puromycin (1 µg/ml), and pooled resistant
colonies were then transfected with FBCsalf envelope expression vector.
Transfected cells were selected with phleomycin, and puro(FeLV-C)
pseudotype virus was harvested from a pooled population of resistant
cells and used for infection studies.
Receptor cloning.
A human T-lymphocyte cDNA library, cloned
into the retroviral vector pBabe-X (25), was generously
provided by R. Sutton (Baylor College of Medicine, Houston, Tex.).
Approximately 10 µg of retroviral plasmid library DNA was transfected
into Phoenix-Eco packaging cells (2 × 106 cells in a
100-mm-diameter tissue culture plate) with SuperFect transfection
reagent (Qiagen, Valencia, Calif.). Two days after transfection, the
viral supernatant was filtered and added with Polybrene (8 µg/ml) to
10 100-mm tissue culture dishes, each containing 5 × 105 NIH 3T3 cells. After 16 h of incubation, the viral
supernatant was replaced with fresh medium. The following day, the
transduced NIH 3T3 cells were transferred to 150-mm tissue culture
plates and incubated with 10 ml of puro(FeLV-C) supernatant for 16 h. The cells were then incubated with another 10 ml of puro(FeLV-C) supernatant for a further 4 h, after which the medium was
replaced. The next day, the cells in each plate were transferred to two 150-mm plates and puromycin was added at 5 µg/ml. Selection medium was replaced every 2 days until resistant colonies had appeared. Resistant colonies were then tested for susceptibility to the lacZ(FeLV-C) pseudotype.
Isolation of receptor cDNA and expression in mammalian
cells.
The transduced cDNA was recovered by subjecting 250 ng of
genomic DNA, isolated from lacZ(FeLV-C)-sensitive NIH 3T3 clones, to
PCR amplification with the Expand PCR kit from Boehringer Mannheim (Indianapolis, Ind.). The 2.0-kb FLVCR cDNA C10 was amplified with
primers complementary to pBabe-X vector sequences flanking the cDNA
insert (upstream primer, 5'-GATCCCAGTGTGCTGGAAAG-3'; downstream primer, 5'-GGTGGGGTCTTTCATTCC-3'). The PCR
was run for 30 cycles with annealing at 54°C for 1.5 min and
extension at 68°C for 7 min. The amplified DNA was cloned into the
pCDNA3.1V5HisTOPO vector (Invitrogen, Carlsbad, Calif.) and was named
pCD3.1VHC10. The DNA sequence was determined at the Microbiology and
Molecular Immunology Core Facility on the PE/ABD 377 sequencer by using dye terminator cycle-sequencing chemistry (Applied Biosystems, Foster
City, Calif.).
CHO cells expressing human FLVCR were generated by transfection of the
pCD3.1VHC10 expression vector. Transfectants were selected
with G418 (1 mg/ml), and resistant cells were analyzed for sensitivity
to
lacZ(FeLV-C).
Northern blots.
Multiple-tissue Northern blots containing
approximately 2 µg of poly(A)+ RNA from various human
tissues were obtained from Clontech (Palo Alto, Calif.). The blots were
probed with full-length FLVCR cDNA that was labeled with
32P by using the random-primer extension system from NEN
Life Science Products (Boston, Mass.).
Nucleotide sequence accession number.
The human FeLV-C
receptor DNA and protein sequences has been assigned Genbank accession
no. AF118637.
| |
RESULTS |
Isolation of a putative human FLVCR cDNA.
To isolate the FLVCR
cDNA, we used a human T-lymphocyte cDNA library subcloned into the
pBabe-X retroviral vector (see Materials and Methods). NIH 3T3
murine fibroblasts, which are naturally resistant to FeLV-C infections,
were transduced with the retroviral library. Transduced cells were
challenged with a replication-defective FeLV-C pseudotype virus that
encodes the dominant selectable marker for puromycin resistance.
Puromycin selection yielded 16 resistant clones, 1 of which (termed
C10) was reproducibly susceptible to infection by lacZ(FeLV-C)
pseudotype virus. The putative receptor cDNA was isolated by subjecting
genomic DNA isolated from C10 cells to PCR amplification with primers
specific for the pBabe-X vector. A DNA product of 2.0 kbp was amplified
and cloned into a mammalian expression vector (see Materials and
Methods) to generate the expression plasmid pCD3.1VHC10. NIH 3T3
fibroblasts and CHO cells transiently transfected with pCD3.1VHC10 were
highly susceptible to lacZ(FeLV-C) infection, whereas cells transfected
with vector alone were resistant (data not shown). We then isolated a
CHO cell clone, CC1011, that stably expressed the FLVCR. CC1011 cells were highly susceptible to lacZ(FeLV-C), whereas no infections were
observed in control CHO cells (Table 1).
In contrast, the CC1011 cells were completely resistant to FeLV-B,
amphotropic murine leukemia virus, and RD114 feline endogenous
retrovirus (Table 1). These results strongly suggest that the 2.0-kbp
C10 DNA encodes a protein that specifically mediates FeLV-C infections.
The FLVCR protein.
Figure 1
shows the predicted amino acid sequence and major structural features
of the human FLVCR. The open reading frame encodes a protein of 555 amino acids that has three NX(S/T) sites for potential N-linked
glycosylation. However, the first of these sites (NDT) may be
inefficiently glycosylated because it contains an acidic amino acid in
the X position (reviewed in reference 18). The
Kyte-Doolittle (26) plot of FLVCR suggests the presence of
12 hydrophobic potential transmembrane (TM) sequences, consistent with
its possible presence in cellular membranes (see Fig. 2).
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FIG. 1.
Amino acid sequence and major structural features of the
human FLVCR protein. The FLVCR protein contains 555 amino acids with 12 hydrophobic potential TM sequences. TM sequences were identified by the
Kyte-Doolittle algorithm (26) and are indicated by bold
lines over the amino acid sequence. Potential N-linked glycosylation
sites are shown by asterisks. The protein contains several dileucines,
which may be required for endocytosis (15).
|
|
BLAST (
2) comparisons with sequences in the databases
showed 99% identity to an expressed sequence tag
(EST176269) derived
from human colon carcinoma Caco-2 cells.
FLVCR also showed strong
homology (45% identity) to a
Caenorhabditis elegans protein (
P = 1 × 10
113) of unknown function and weaker homologies
(21% identity) to
a bacterial glycerol-3-phosphate transporter
(
P = 1 × 10
5) and to a bacterial
glucarate transporter (
P = 2 × 10
5). Interestingly, the bacterial transporters are
both members
of the ancient major-facilitator superfamily (MFS) of
transporters,
which also generally contain 12 hydrophobic TM sequences
(
38).
The MFS transporters have been classified into 17 evolutionary
branches or subgroups that transport distinct categories
of solutes
(
38). FLVCR is most closely related to the
organic-phosphate
antiporter (OPA) and anion/cation symporter (ACS)
subfamilies,
which both transport organic anions. As shown in Fig.
2, FLVCR
has a size and hydrophobicity
profile that is closely similar
to those of various MFS transporters,
including the type 2 glucose
transporter from human liver and the
bacterial glucarate and glycerol-3-phosphate
transporters. Like the MFS
transporters, FLVCR has a relatively
large hydrophilic sequence between
TM6 and TM7.
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FIG. 2.
Hydrophobicity plot of human FLVCR and comparison to
plots of several members of the MFS transporters. Hydrophobicity plots
were generated by the Kyte-Doolittle algorithm (26). The
human FLVCR has a similar hydrophobicity plot to that of bacterial
glucarate and glycerol-3-phosphate transporters (GlucarateT and G3PT,
respectively) and to that of the type 2 glucose transporter (GlucT)
from human liver (12). The results suggest that FLVCR
contains 12 TM domains, which are numbered above the hydrophobicity
plot. This is a general characteristic of MFS transporters, which
typically contain 12 TM region with a large hydrophilic region between
TM6 and TM7.
|
|
Further evidence that FLVCR is a member of the MFS superfamily is shown
in Fig.
3. Specifically, it has been
found that the
most highly conserved sequence in the MFS transporters
occurs
in the hydrophilic sequence of defined length that separates TM2
and TM3 (
38). The FLVCR sequence in this region is compared
in Fig.
3 with the corresponding sequences of representative
transporters
in the MFS superfamily and with the consensus sequence for
all
17 evolutionarily distinct subfamilies of the MFS transporters
(
38). The subscripts at the consensus amino acid positions
indicate
the numbers of the 17 MFS subfamilies that adhere to the
consensus
at that site. Thus, the most highly conserved amino acids are
glycine at position 1 (in 11 of the 17 subfamilies), leucine at
position 3 (in 11 of the 17 subfamilies), aspartic acid at position
5 (in 12 of the 17 subfamilies), and glycine at position 8 (in
15 of the
17 subfamilies). The similarity scores indicated in
Fig.
3 are the sums
of the subscripts for all positions of identity
of the query sequence
and the consensus sequence. In contrast
to the scores indicated for
these MFS transporters, random peptides
of this length would have an
average similarity score of only
4.0. Thus, FLVCR closely matches this
consensus in both its length
and its sequence.
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FIG. 3.
Signature sequence for MFS transporters compared with
the corresponding sequence of the human FLVCR. The MFS signature
sequence, which occurs in the hydrophilic loop that separates TM2 and
TM3, is the most highly conserved sequence in this transporter
superfamily. The sequence for FLVCR is shown at the top, and residues
identical to the consensus are boxed. Other MFS transporter sequences
are also shown for C. elegans (accession no. AF002196) and
for the bacterial glucarate transporter (GlucarateT) (accession no.
P42237) and glycerol-3-phosphate transporter (G3PT) (accession no.
AJ235270). In addition, the consensus sequences are shown for the ACS,
OPA, and monocarboxylate porter (MCP) subfamilies of the MFS
transporters (38). The bottom line shows the overall
consensus sequence for all MFS transporters as compiled by Pao et al.
(38). In this line, each amino acid is indicated by a
subscript, which indicates its frequency of occurrence in the consensus
sequences of the 17 distinct lineages of MFS transporters. For example,
glycine (G) at position 1 occurs in 11 of the 17 subfamilies whereas G
at position 8 occurs in 15 of the 17 subfamilies. The similarity score
for each sequence is the sum of these subscript score for each
corresponding position. Random sequences would have very low average
similarity scores.
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|
FLVCR expression in human tissues.
We analyzed the tissue
distribution of FLVCR expression by Northern blot analysis with the
2.0-kb C10 cDNA as a probe (Fig. 4). A
2.0-kb RNA transcript was present in all hematopoietic tissues including peripheral blood lymphocytes, and was most abundant in the
fetal liver. However, relatively little of this RNA transcript was
present in nonhematopoietic tissues except the pancreas and kidneys. In
addition, a 7.5-kb RNA was detected in most tissues except the liver
and skeletal muscle.
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FIG. 4.
FLVCR RNA expression in human tissues. The multiple
tissue Northern blots containing poly(A)+ RNA from various
human tissues were probed with 32P-labeled C10 cDNA. FLVCR
RNA transcript was present in all hematopoietic tissues and also in the
pancreas and kidney. Relatively little RNA expression was present in
other tissues. An additional transcript of 7.5 kb was present in most
tissues. P, pancreas; K, kidney; SM, skeletal muscle; L, liver; Lg,
lung; Pl, placenta; B, brain; H, heart; FL, fetal liver; BM, bone
marrow; PBL, peripheral blood lymphocyte; T, thymus; LN, lymph node; S,
spleen.
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|
| |
DISCUSSION |
Functional cloning of human FLVCR.
We report the functional
cloning and characterization of a 2.0-kbp human cDNA from human T
lymphocytes that encodes a putative cell surface receptor (FLVCR) that
mediates FeLV-C infections. This cloning was done with a representative
cDNA library in a retroviral vector, by procedures that have recently
been successfully used to clone coreceptors for simian immunodeficiency
viruses (8) and receptors for xenotropic and polytropic
murine leukemia viruses (3, 48, 55), and for type D primate
retroviruses (39, 49). Expression of FLVCR cDNA in NIH 3T3
fibroblasts (data not shown) and in CHO cells (Table 1), which are both
resistant to FeLV-C, resulted in substantial susceptibility of the
cells to this virus. In contrast, FLVCR did not confer susceptibility to infections of FeLV-A (data not shown), FeLV-B, amphotropic murine
leukemia viruses, or RD114 feline endogenous retrovirus (Table 1),
suggesting that FLVCR is highly specific for FeLV-C. Northern blot
analyses indicated expression of a 2.0-kb FLVCR mRNA in diverse
hematopoietic tissues including peripheral blood lymphocytes, with
relatively little expression in other tissues. These results are in
accordance with a report that FeLV-C can infect diverse hematopoietic
cells in vivo (7) and with our isolation of the FLVCR cDNA
from a T-lymphocyte library.
Our evidence does not exclude the possibility that additional or
alternative FeLV-C receptors occur in certain cells. The
FLVCR that we
have identified is a member of a large and ancient
MFS transporter
superfamily that has many related members. Moreover,
FeLV-C is closely
related to FeLV-A and apparently is formed from
FeLV-A within infected
cats by a small number of mutations that
modify the vr1 domain of the
envelope glycoprotein (
32,
42).
These mutations evidently
change the receptor-binding specificity
of the virus (
4,
42)
and are critical for the onset of aplastic
anemia (
1,
10,
20,
32,
35,
40-42). The close precursor-progeny
and sequence
similarities of FeLV-A and FeLV-C imply that FeLV-A
might use a
receptor that is related to FLVCR. In accordance with
this hypothesis,
a putative receptor for FeLV-A that was identified
by biochemical
methods has an apparent
Mr 70,000 (
13), consistent
with the expected size of a glycosylated
form of FLVCR (Fig.
1)
and with the sizes of MFS transporters
(
38). Furthermore, it
is conceivable that some FeLV-C
isolates might use FLVCR or closely
related MFS transporters with
differing efficiencies. These relationships
appear strikingly similar
to those for human immunodeficiency
virus type 1, which is transmitted
in a form that employs the
CCR5 coreceptor but progresses by in vivo
mutations that modify
the small variable region V3 in the surface
envelope glycoprotein
to form derivatives that use related coreceptors
such as CXCR4
(
5,
8,
11,
14,
29). All of the
immunodeficiency virus
coreceptors are members of a common protein
superfamily, and the
different forms of human immunodeficiency virus
also have different
cellular tropisms and pathogenic effects (
11,
29).
The FLVCR protein.
The FLVCR protein contains 555 amino acids
with three NX(S/T) sites for potential N-linked glycosylation and with
12 hydrophobic potential TM sequences. The FLVCR protein is closely
related (45% identity) to a C. elegans protein of unknown
function and is less closely related (21% identity) to bacterial
glycerol-3-phosphate and glucarate transporters, which are both members
of the MFS superfamily of transporters (38). This large
superfamily, which is ancient, contains at least 17 evolutionarily
distinct branches, most of which separated from the trunk shortly after
the origin of the superfamily approximately 2 billion years ago
(38). As a consequence of their ancient divergences, most
MFS transporters are only distantly related in their sequences
(38). Moreover, each branch or subfamily has specialized in
the transport of a particular category of solutes. The
glycerol-3-phosphate and glucarate transporters occur in the OPA and
ACS subfamilies of MFS transporters, which have both specialized for
transport of different types of organic anions. These relationships
suggest that FLVCR is also a transporter and that it may transport an
organic anion.
The identification of FLVCR as a member of the MFS transporter
superfamily is strongly supported by additional considerations.
Like
FLVCR, MFS transporters typically contain 12 TM sequences
with a
relatively large hydrophilic loop between TM6 and TM7.
These
similarities are illustrated in Fig.
2, which compares the
sizes and
hydrophobicity profiles of FLVCR with those of several
members of the
MFS family including the widely studied mammalian
facilitated glucose
transporters. In addition, FLVCR has a conserved
signature sequence in
the hydrophilic region between TM2 and TM3
that is highly conserved
throughout the MFS superfamily (Fig.
3). These features strongly
suggest that FLVCR is a member of
the MFS transporter superfamily and
imply that its amino and carboxyl
termini are in the cytosol and that
its TM domains each traverse
the membrane. Accordingly, the regions
most likely to interact
with FeLV-C occur in the hydrophilic
extracellular loops between
TM1 and TM2, TM3 and TM4, TM5 and TM6, TM7
and TM8, TM9 and TM10,
and TM11 and TM12. As expected, this implies
that the potential
sites of N-linked glycosylation between TM5 and TM6
are in the
lumen of vesicles or facing the extracellular
milieu.
General implications.
Our results strongly suggest that FLVCR
is expressed in diverse hematopoietic cells including peripheral blood
lymphocytes and T cells but is only weakly or negligibly expressed in
many other tissues (Fig. 4). This is consistent with evidence that FeLV-C preferentially infects different lineages of hematopoietic cells
in vivo (7). Thus, although the major determinant of FeLV-C-induced aplastic anemia has been mapped to the vr1 region, which
controls the receptor-binding specificity of the envelope glycoprotein
(42), it is clear that FLVCR is not restricted to
erythroblasts. In addition, our results suggest that FLVCR is likely to
be a facilitative transporter for organic anion(s) (see above), and we
propose that it transports an important nutrient or regulator that is
critical for erythroblast survival in vivo and in stroma-dependent bone
marrow cultures. By analogy to other retroviral receptors (22, 24,
34, 53), it is likely that FLVCR transport function would be
inhibited or down-modulated following infection with FeLV-C, and it is
conceivable that this transport function is more critical for
erythroblasts than for other hematopoietic cells. Alternatively, this
transport activity might be necessary for function or signaling by
stromal or nurse cells in the erythropoietic microenvironment (1,
27, 28). Our present results should facilitate further
investigations and elucidation of the normal FLVCR function. In
addition, these results raise the possibility that FeLV-A, which is a
closely related progenitor of FeLV-C, may also use an MFS transporter
for infection. Such a result would confirm a strong similarity to human
immunodeficiency virus type 1, which also evolves by env
gene mutations in vivo to form divergent progeny that utilize distinct
members of a receptor superfamily and that have different cellular
tropisms and pathogenic effects (5, 11, 14, 29).
| |
ACKNOWLEDGMENTS |
We are extremely grateful to Richard Sutton for providing the
human retroviral cDNA library, to Yasuhiro Takeuchi and Garry Nolan for
providing the packaging cells, and to Susan Kozak for helping with the
Northern blot analysis. We are also grateful to our coworkers Emily
Platt, Mariana Marin, Navid Madani, Shawn Kuhmann, and Ali Nouri for
helpful suggestions.
This work was supported by NIH grant CA25810.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Department of
Biochemistry and Molecular Biology, Oregon Health Sciences University, 3181 SW Sam Jackson Park Rd., Mail Code L224, Portland, OR 97201-3098. Phone: (503) 494-2548. Fax: (503) 494-8393. E-mail:
tailorc{at}ohsu.edu and kabat{at}ohsu.edu.
| |
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Abstract
Introduction
