1 Division of Developmental and Clinical Immunology, 2 Division of Hematology/Oncology, 3 Department of Medicine, 4 Department of Pediatrics, 5 Department of Pathology and 6 Department of Microbiology, University of Alabama at Birmingham, Birmingham, AL 35294-3300, USA
7 Howard Hughes Medical Institute, Birmingham, AL 35294-3300, USA
8 Laboratory of Immunopathogenesis and Bioinformatics, Science Applications International Corporation-Frederick, Building 550, Room 204 FCRDC PO Box B, Boyles Street, Frederick, MD 217021201, USA
9 Present address: Department of Pathology, Stanford University, 269 Campus Drive, Room CCSR 3250, Stanford, CA 943055176, USA
Correspondence to: M. D. Cooper; E-mail: max.cooper{at}ccc.uab.edu
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Abstract |
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Keywords: B cell differentiation, Fc receptors, immunoglobulin superfamily, immunoreceptor tyrosine-based motifs, phylogeny
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Introduction |
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Members of the human FcRH15 subfamily belong to the network of receptors that possess immunoreceptor tyrosine-based activating motifs (ITAM), inhibition motifs (ITIM) or both (12). ITAMs are characterized by two repeats of the consensus sequence Y-X-X-L/I separated by 68 amino acids (E/D)-X-X-Y-X-X-(L/I)-X68-Y-X-X-(L/I), while ITIMs characteristically have a six amino acid consensus sequence (I/V/L/S)-X-Y-X-X-(L/V) (1315). The FcRH ITIM sequence motifs closely match the defined consensus, whereas most FcRH ITAM candidate motifs correspond less precisely to the consensus sequence, and some of the sequences ambiguously resemble either part of an ITAM or an ITIM. Like the FcRII receptors, the presence of these potential immunoregulatory motifs in the FcRH cytoplasmic tails suggests autonomous signaling potential. In contrast, the other activating classical FcRs require co-association with the FcR common
-chain (FcR
c) or other adaptor subunits for signaling competence (1619).
The classical Fc receptors in mice have been shown to positively or negatively modulate antibody mediated responses of lymphocytes and inflammatory cells (20). The locus for the mouse Fc receptor genes is split between mouse chromosome 3, where the moFcRI resides, and mouse chromosome 1, where moFc
R
, moFc
RII and moFc
RIII are located (2124). There are a limited number of mouse FcR genes relative to their more highly diversified human counterparts. This feature has facilitated their targeted disruption to enable clarification of their biological roles as activating and inhibitory receptors. Mice deficient in the Fc receptors have profoundly altered humoral immune responses, immediate hypersensitivity, cytotoxic inflammatory responses and immune complex mediated inflammation (2530). These findings emphasize the important pathophysiologic roles of Fc receptors in autoimmunity, allergy and inflammation.
The immunoregulatory potential of the huFcRH15 family members and their preferential B cell expression raise interesting issues regarding their biological importance. Their identification also reveals that this 1q21-23 region contains an unexpectedly large number of genes with potential Ig binding function and signaling capacity. As a first step towards gaining greater insight into the diversity of this gene family and the functional potential of the individual family members, we sought to identify and characterize the FcRH relatives in mice. Using amino acid sequences of Ig-like domains specific to the huFcRH15 subfamily to search the available databases, three potential mouse orthologs (moFcRH13) were identified. In this report we describe their chromosomal location, genomic configuration, sequence and cellular expression patterns. In addition, these mouse Fc receptor homologs are compared with their human counterparts.
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Methods |
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Generation of full-length moFcRH13 cDNAs
Primers used in end-to-end amplification to generate full-length cDNAs were as follows: moFcRH1, forward 5'-GTGAGAGGCACCTTCAAGTTACCAT-3' and reverse 5'-GGTGAAGGACTCATCTAATGAACCG-3'; moFcRH2, forward 5'-CCACAGTGTTCTATCCCAGATCCGT-3' and reverse 5'-GAGGCCCAGTGCAGAAAGTAGGAG-3'; and moFcRH3, forward 5'-GTGAGTGACTACCATTGCGAGCAAG-3' and reverse 5'-CAGGCCCAGTAGAAGCATCGG-3'. Each amplification reaction underwent initial denaturation at 94°C for 30 s followed by 30 cycles of denaturation at 94°C for 5 s and annealing at 68°C for 4 min, and final extension at 72°C for 6 min.
DNA sequence analysis
PCR products were ligated into the pCR2.1 TOPO T/A vector (Invitrogen, Carlsbad, CA). Inserts were sequenced on both strands by the dideoxy chain termination method using SequiTherm EXCEL II (Epicentre Technologies, Madison, WI) and an automated sequencer (LiCor, Lincoln, NE). Nucleotide and amino acid sequence alignment was analyzed with a DNASTAR (Madison, WI) software package and homology searches were performed using the Basic Local Alignment Search Tool (BLAST) (31).
RNA blot analysis
A mouse tissue northern blot (Ambion, Austin, TX) was hybridized with the following PCR amplified and [-32P]dCTP labeled gene specific probes from respective moFcRH cDNAs: a 549 bp fragment (5691117) corresponding to D23'UTR of moFcRH1, a 198 bp fragment (48245) corresponding to the S2D1 region of moFcRH2 and a 440 bp fragment (289728) corresponding to D1D3 of moFcRH3. According to the manufacturer's instructions, membranes were hybridized overnight at 65°C, washed and exposed to X-ray film.
For northern blots of mouse cell lines, RNA was extracted from freshly grown cells with the RNeasy kit (Qiagen, Valencia, CA). Ten micrograms of total RNA were electrophoresed through a 1% formaldehyde/agarose gel and blotted onto Nytran membrane following the manufacturer's instructions (Schleicher & Schuell, Keene, NH). The membrane was hybridized with [-32P]dCTP labeled gene specific probes (as described above) overnight at 65°C, in 0.25 M NaH2PO4 (pH 7.2), 1 mM EDTA, 5% SDS, 0.1% sodium pyrophosphate (Na4P2O7.10H2O) and 100 µg/ml of total yeast RNA. Washes were performed at 65°C: twice in 0.125 M NaH2PO4 (pH 7.2), 0.05 mM EDTA and 2.5% SDS and twice in 0.025 M NaH2PO4 (pH 7.2), 0.01 mM EDTA and 0.5% SDS. Membranes were exposed to film for 17 days at 80°C.
Immunofluorescence and cell sorting
Bone marrow cell suspensions prepared from BALB/cJ mice (Jackson Laboratory, Bar Harbor, ME) were stained with APC-conjugated anti-CD19, PE-conjugated anti-CD43 (both from BD Biosciences, San Diego, CA) and FITC-conjugated goat anti-µ heavy chain (Southern Biotechnology Associates, Birmingham, AL) and cells representative of the different stages in B cell differentiation (32) were separated using a MoFlo flow cytometer (Cytomation, Fort Collins, CO). Spleen cells from BALB/cJ mice were stained with FITC-conjugated anti-CD19, PE-conjugated CD23 (BD Biosciences) and Cy5-conjugated anti-CD21 (a kind gift of Dr John Kearney), APC-conjugated anti-CD4 and PE-conjugated anti-CD8 (BD Biosciences), or PE-conjugated DX5 and FITC-conjugated anti-CD11c (BD Biosciences) and the cells were differentially sorted using a MoFlo flow cytometer. Newly-formed (NF) B cells were isolated as CD19+CD21CD23, follicular (FO) B cells were isolated as CD19+CD21+CD23+, marginal zone (MZ) B cells were isolated as CD19+CD21hiCD23lo (33), NK cells were isolated as DX5+CD11c, and dendritic cells were isolated as DX5CD11c+. Peritoneal lavage cells (PLC) from BALB/cJ mice were obtained by saline lavage of the peritoneal cavity. PLC cells were stained with APC-conjugated anti-CD19, PE-conjugated anti-CD5 and FITC-conjugated anti-CD11b (all from BD Biosciences), and separated with a MoFlow flow cytometer. Sorted cells were >98% pure and real-time PCR was performed at least three times from two independent sorts.
Reverse transcription (RT)PCR
Total cellular RNA extracted from mouse cell lines with the RNeasy kit (Qiagen) was primed with random hexamers and oligo dT primers and reverse transcribed with SuperScript III (Life Technologies, Carlsbad, CA) into single-stranded cDNA. Gene specific primers for RTPCR were moFcRH1 forward 5'-GCATCATCCTGGGGAACAGTTCAGCAC-3' and reverse 5'-CTCATCTAATGAACCGCAGTG-3'; moFcRH2 5'-GGCAACGACCCAGCTACGCTA-3' and reverse 5'-CGCCACATCTCCGATGAAG-3'; and moFcRH3 forward 5'-AGGTGAACATCAGTGACGC-3' and reverse 5'-TGGTTGAGTTCTCCGTACTTCT-3'. Each amplification reaction underwent initial denaturation at 94°C for 5 min followed by 35 cycles of denaturation at 94°C for 30 s, annealing at 60°C for 30 s, extension at 72°C for 1 min and final extension at 72°C for 7 min. Amplified products were visualized in 1% agarose gels containing ethidium bromide and documented with the Bio-Rad Fluor-S Imager (Hercules, CA).
Real-time PCR
RNA from sorted cells, mouse tissues and cell lines was isolated using the RNeasy Mini kit with on column DNase digestion as recommended by the manufacturer (Qiagen). Total cellular RNA was primed with random hexamers and oligo dT primers and reverse transcribed with SuperScript III (Life Technologies) into single-stranded cDNA. Gene specific primers for real-time PCR were as follows: moFcRH1 forward 5'-GAACCTGCTGGAATCTCTGATGT-3' and reverse 5'-TCCCTCCATCACCCATCCT-3'; moFcRH2 forward 5'-TGACTGCCTCTCGCAGTGTCT-3' and reverse 5'-TGACTTGAGATACAGGGATCCTCTCTA-3'; and moFcRH3 forward 5'-GCCAAGCCGACAGCTTACTTC-3' and reverse 5'-ACAGCAGGTGGAGCTTGCA-3'; ß-actin forward 5'-GCTCTGGCTCCTAGCACCAT-3' and reverse 5'-GCCACCGATCCACACAGAGT-3'. The real-time PCR reactions were set up per manufacturer's instructions with 2x Syber Green PCR Master Mix (Applied Biosystems, Warrington, UK) and run on a 7900HT Sequence Detection System (Applied Biosystems).
Cell lines
Mouse cell lines included RAW8.1 and SCID7 pro-B cell lines, the 70Z/3 pre-B cell line, WEHI-231 and WEHI-279 immature-B cell lines, the CH-12, A20 and X16C8.5 B cell lines, the EL4 T cell line and the myeloid cell line WEHI-3.
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Results |
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The two moFcRH1 isoforms differ by alternative splicing of the transmembrane exon. The short isoform, moFcRH1S, lacks the sequence encoded by this exon and is presumably a secreted protein. The longer isoform, moFcRH1L, is a type I transmembrane protein with an uncharged transmembrane region and a cytoplasmic tail that contains a potential ITAM.
Both moFcRH2 isoforms, moFcRH2Ig and moFcRH2sc, have four extracellular Ig-like domains and lack transmembrane regions. The carboxy terminus of the moFcRH2sc isoform contains in addition a single type B scavenger receptor cysteine-rich (SRCR) domain with eight cysteines (37). This unusual chimeric protein, not found among other presently characterized human and mouse proteins, shares 56% amino acid identity with the amino terminal domain of the product of its closest neighboring gene, moSp/CD5L/Api6 (38).
The two moFcRH3 isoforms identified in this analysis are type I transmembrane proteins that differ by alternative splicing of the first of five Ig-like domains. The short isoform, moFcRH3S, lacks the sequence encoding the first Ig domain, but the rest of the sequence is identical to moFcRH3L which contains all five Ig domains. Their cytoplasmic tails contain a consensus ITIM and an ITAM-like sequence with a glutamic acid at the +3 position relative to the second tandem tyrosine. This analysis suggests the mouse has only two FcRH members that closely resemble their human FcRH15 counterparts. Despite the conserved extracellular regions for these FcRH family members, only moFcRH1 and moFcRH3 may possess immunoregulatory tyrosine-based signaling potential.
MoFcRH1, moFcRH2 and moFcRH3 genomic organization
Genomic sequences of the moFcRH locus were identified by BLASTN searches of the Ensembl, Celera and NCBI databases using the moFcRH cDNA sequences. Exon/intron boundaries were determined by comparing moFcRH cDNAs with genomic DNA sequences using the AG/GT rule. A C57BL/6 derived BAC clone, RP23 135C10, corresponding to the 5' end of the locus was found to contain moFcRH2 and moFcRH1, but not moFcRH3 (Fig. 1A). The position of moFcRH3 was confirmed by PCR verification of its position on a partially overlapping BAC clone, RP23 228P24, which lies just telomeric of RP23 135C10 (data not shown). The moFcRH locus thus spans 215 kb and is organizationally more complex than the human syntenic region. The 5' end of the locus is demarcated by the 3' end of the moFcRH2 gene, which lies in the reverse transcriptional orientation relative to moFcRH1 and moFcRH3.
MoFcRH1 consists of 10 exons that span 15.2 kb. All exon/intron boundaries follow the phase 1 splicing pattern, in that splicing occurs after the first nucleotide of the triplet codon except for the CY1/CY2 boundary, which follows a phase 2 pattern, and the CY4/CY5-3'UTR boundary, which follows the phase 0 pattern (Fig. 2). Like other members of the extended FcR family, the first exon 5'UT/S1 encodes the 5' untranslated region, the ATG translation initiation codon, and the beginning of a split signal peptide. The second exon, S2, encodes the second half of the split signal peptide and is only 21 bp in length, a characteristic feature of all FcR and FcRH relatives except for FcRX (1,2,10,22,24,3942). The Ig-like extracellular domain is encoded by two exons, D1 and D2, separated by an intron of only 167 bp. Exon 5 encodes the membrane-proximal, hydrophobic/uncharged transmembrane domain and the beginning of the cytoplasmic tail. The presumably secreted moFcRH1S isoform splices out this transmembrane encoding exon and maintains the same ORF with the remaining exons that encode the cytoplasmic region of the moFcRH1L isoform. The cytoplasmic tail is encoded by five exons, CY1CY5, the last of which also encodes the translation termination codon and the 3'UT. MoFcRH1 and moFcRH3 are separated by
44 kb and, unlike moFcRH2, both are transcriptionally oriented towards the telomere.
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MoFcRH3 contains 13 exons that span 32 kb. All of its exon/intron boundaries follow the phase 1 splicing pattern, except for the CY1/CY2 boundary which is phase 2. Like other family members, moFcRH3 has a split signal peptide encoded by two exons, the second of which is also 21 bp. The extracellular Ig domains, D1D5, are encoded by five exons clustered within a
6.4 kb region. The first of these, D1, is spliced out in the moFcRH3S isoform while maintaining the ORF with the remaining 3' encoding exons. These are followed by an
6.1 kb intron and exon 8 that encodes the membrane-proximal, hydrophobic/uncharged transmembrane region and proximal portion of the cytoplasmic domain. The CY1CY4 regions of the cytoplasmic tail are encoded by four exons, the last of which also encodes the translation termination codon and the beginning of the 3'UT. Unlike moFcRH1 and moFcRH2, moFcRH3 has a second 3'UT transcribed exon that is
9 kb from CY4.
Relationship of mouse FcRH13 with other FcR/FcRH family members
The Ig domains present in members of the extended human FcR/FcRH family can be divided into five subtypes on the basis of their respective amino acid identities in a CLUSTAL driven analysis (1,7,43). Comparison of the moFcRH13 domain relationships with the mouse FcRs and FcRHs in an unrooted pairwise analysis indicates that the extracellular regions of the mouse FcR/FcRH family members consist of different combinations of the five Ig-like domain subtypes (Fig. 3A and B). The ordering of the membrane-proximal to membrane-distal Ig domains for the mouse FcRH and FcR family members resembles that observed for their human relatives. Greater amino acid sequence relatedness is usually seen for membrane-proximal domains, while more membrane-distal Ig domains characteristically exhibit greater diversity.
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An interspecies comparison of mouse and human FcRH orthologs indicates that moFcRH1 has greatest extracellular identity (61%) with huFcRH1 (Fig. 3C). The Ig domains of moFcRH2 have 46% identity with both huFcRH1 and huFcRH2. Although moFcRH3 exhibits 45% extracellular identity with huFcRH2 and 40% with huFcRH1, huFcRH3 and huFcRH5, the domain composition of moFcRH3 is most similar to huFcRH3. Comparisons of the cytoplasmic domains indicate 43% identity between huFcRH1 and moFcRH1 and 48% identity between moFcRH3 and huFcRH5 (data not shown).
Pairwise Ig domain comparisons of FcR/FcRH family members indicate that moFcRH3 possesses the D1 and D2 Ig binding domain subtypes employed by the FcRs. To extend this analysis, we threaded the amino acid sequence of moFcRH3 D1 and D2 into the Position Specific Scoring Matrix (PSSM) database to compare this sequence with currently solved protein folds in the library (44). This analysis indicates that huFcRIIB is the closest recognizable structure to the linear and predicted secondary structure of moFcRH3 D1 and D2. A significance value of 2.26e-05 indicates that moFcRH3 has a high probability of forming a similar structural fold to that of huFc
RIIB.
Tissue distribution of moFcRH1, moFcRH2 and moFcRH3 expression
MoFcRH1 transcripts of 1.6 kb were easily identified by RNA blot analysis in spleen and thymus and at lower abundance in kidney and lung (Fig. 4A). MoFcRH2 was more widely expressed, with transcripts of
2.4 kb being detected in whole embryo (14 day), lung, ovary, brain, testes, thymus, heart and kidney, albeit in trace levels in the latter four tissues. Notably, moFcRH2 transcripts were not seen in spleen. In contrast, moFcRH3 transcripts were found only in the spleen, where a faint band of
2.5 kb was sometimes detected with gene specific probes (data not shown). The inconsistency of this finding suggested that northern blot analysis is insufficiently sensitive for accurate detection of moFcRH3 expression in tissues, perhaps because moFcRH3 is expressed at relatively low levels or by a minor subpopulation of cells.
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MoFcRH expression in primary B lineage cells
The patterns of moFcRH13 expression were examined during normal B lineage differentiation by isolating subpopulations of B lineage cells via their defining cell surface antigens from bone marrow, spleen and peritoneal lavage samples. Gene specific primers were then employed in a real-time PCR analysis of moFcRH13 transcripts. Whereas moFcRH1 transcripts were barely detectable in early B lineage cells of bone marrow origin, they were present in higher levels in the newly-formed, follicular and marginal zone subpopulations of splenic B cells (Fig. 5). Minimal moFcRH1 expression was evident in sorted populations of splenic T cells, dendritic cells and NK cells. In peritoneal lavage samples, moFcRH1 expression was detectable in each of the B cell subsets, B1a, B1b and B2, but not in macrophages (data not shown). MoFcRH2 transcripts were not detectable in bone marrow and splenic B lineage cells, or in other types of splenocytes, but were surprisingly abundant in peritoneal macrophages (Fig. 5; and data not shown). In contrast, moFcRH3 expression was most prominent in marginal zone B cells, much less evident in the newly-formed and follicular splenic B cell subpopulations, and not seen in macrophages, DC, NK and T cells from either spleen or the peritoneal cavity. These results indicate a relatively broad pattern of expression for moFcRH1 during B cell differentiation, while moFcRH3 is preferentially expressed by marginal zone B cells.
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Discussion |
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Our identification of huFcRH15 derived from an informatics based search using a consensus sequence generated from the Fc binding regions of FcRI, Fc
RII and Fc
RIII (1). The related extracellular composition, gene structure and signaling potential of these receptors suggests a common phylogenetic origin of the FcR and FcRH families (17). The validity of this interpretation is strengthened by the characterization of the mouse FcRH13 gene family here and in a previous report describing ESTs for two of these family members, moFcRH1/mIFGP1 and moFcRH2/mIFGP2 (6). As for the other FcR/FcRH family members, with the exception of moFcRX and huFcRX, the moFcRH13 genes share structural similarity that includes a trademark 21 bp S2 exon encoding the second portion of a split signal peptide. MoFcRH proximity to the most closely related FcR family member, moFc
RI, on chromosome 3 also attests to the evolutionary relationship between the FcR and FcRH gene families.
The FcR and FcRH genes comprise a larger, more diverse family in humans and mice than formerly appreciated. The FcR gene family members, like the FcRH family members, are composed of multiple genetic units that reflect significant duplication and diversification over the 90 million years since rodents and primates shared a common ancestor (39,47). While the location of moFcRH13 near moFc
RI, CD1 and Sp
is not surprising, given the relative positions of these genes in humans, it is noteworthy that the FcR family cluster is divided between chromosomes 1 and 3 in mice. A high recombinatorial rate for this chromosomal region may account for the inverted orientation of moFcRH2, the inclusion of an SRCR domain encoding exon which may have its origin in the Sp
gene, and the apparent loss of moFcRH2 lymphoid specific promoter elements. Involvement of the IRTA1/FcRH4 gene in a chromosomal translocation with the human Ig locus accords with other evidence indicating that the FcR/FcRH region is a recombinatorial hotspot (3).
The mammalian FcRH and FcR genes appear to represent an ancient gene family with members of the FcRH gene family being conserved in chickens, frogs and bony fish (4850). Remarkably, at least 20 FcRH family members have recently been identified in Xenopus (50). Additional phylogenetic analysis of the FcR and FcRH families is needed to clarify the evolution of the FcR/FcRH locus, and this endeavor may also yield informative clues to the functions of these Ig-like receptors. In this regard, the closest phylogenetic relative of the huFcRIII (CD16) gene has only recently been identified in the mouse, FcRL3/CD16-2 (11).
The tandem order of the different subtypes of Ig domains in the extracellular regions of FcRH and FcR family members (Fig. 3) is highly conserved. The Ig Fc binding sites are located in the FcR D1 (red in Fig. 3B) linker regions and the neighboring D2 (dark blue) domains. These Ig domain subtypes are seen less frequently in the moFcRHs than in huFcRH family members, although moFcRH3 has D1 and D2 domains that may have Ig-binding potential. This possibility is supported by a three dimensional position specific scoring matrix (3D-PSSM) analysis indicating a high degree of structural similarity between these two moFcRH3 domains and the related huFcRIIB domains. In contrast, the more membrane-proximal Ig domains of moFcRH3 and moFcRH1 (light blue and green in Fig. 3B) are FcRH family specific. The absence of FcR-like Ig domains in moFcRH1 suggests that, like the huFcRH1, it may not have Fc binding potential, but rather a different ligand(s).
The tyrosine-based motifs in the cytoplasmic regions of the transmembrane moFcRH1 and moFcRH3 molecules resemble those of the huFcRHs. Like its nearest human relative, huFcRH1, moFcRH1 has a canonical ITAM in its cytoplasmic tail, suggesting the potential for cellular activation. A different function would be predicted for the moFcRH1S isoform that lacks a transmembrane region; such alternatively spliced isoforms have similarly been observed for moFcRII (51). MoFcRH3 differs in that it possesses an ITIM and an ITAM-like sequence, while its closest human relative, huFcRH5, has two consensus ITIMs and an ITAM-like sequence. The presence of ITIM and ITAM-like motifs in moFcRH3 and several members of the human FcRH family suggests they may have either inhibitory or activating potential depending upon the signaling context. A precedent for this type of dual functionality has been demonstrated for CD22, another transmembrane molecule on B cells (52,53).
MoFcRH2 differs from all of the human FcRHs in its preferential expression by non-lymphoid lineage cells. Neither of the two predicted moFcRH2 isoforms has a transmembrane region, and both are therefore likely to be secreted. The Ig/SRCR domain-containing moFcRH2sc isoform is an unusual chimeric protein. The exon encoding the SRCR domain could have been derived from a recombination event involving moSp, its chromosome 3 neighbor, given the high degree of identity (56%) with the Sp
N-terminal SRCR domain (38,54). Moreover, in contrast with the lymphoid tissue-specific expression of mouse and human Sp
, moFcRH2 is primarily expressed in non-lymphoid tissues.
Like their human counterparts, the moFcRH1 and moFcRH3 genes are selectively expressed by B lineage cells at different stages in differentiation. MoFcRH1 is expressed by many of the mature B cell subpopulations, including newly formed, follicular and marginal zone B cells. MoFcRH1 may thus serve an activating role on mature B cells, as appears to be the case for its human counterpart, huFcRH1 (46). The most remarkable feature of moFcRH3 is that it is preferentially expressed by B cells in the marginal zone subpopulation. Marginal zone B cells are typified by their topographic location, pre-activated state and distinct receptor repertoire, characteristics that enable them to participate in early responses to T independent blood-borne antigens (55,56). However, it may be noteworthy that memory B cells are also present in the marginal zone (57), especially since huFcRH4/IRTA1 has been shown to be preferentially expressed by memory B cells, where it may serve an important regulatory role in antibody responses (45,58). It will therefore be important to determine which type(s) of marginal zone B cells account for the moFcRH3 expression that we have observed.
The true functions of moFcRH1 and moFcRH3 on B lineage cells will only become clear with further investigation, but differences in their structure and expression patterns suggest potential immunoregulatory roles. MoFcRH1 possesses an ITAM-like sequence in its cytoplasmic tail, whereas moFcRH3 has both an ITIM and ITAM-like motif. Given the presence of domain subtypes (D1 and D2) that resemble the Fc binding domains of classical FcRs, it is possible that moFcRH3 may bind Ig or antigen-antibody complexes on marginal zone B cells. Since these cells also express FcRIIB, a competitive or cooperative relationship could exist between these receptors. The resulting impact on activation or inhibition is difficult to predict, especially given it may depend upon the Ig isotype and other microenvironmental factors. MoFcRH1 is more likely to have an activating function on the B cells that express it. Our studies of human FcRH1 indicate broad expression among mature B cell populations, where it can serve a role as an activation co-receptor (C.-M. Leu, R. S. Davis, L. A. Gartland, W. D. Fine, and M. D. Cooper, submitted for publication). In conclusion, the differential B lineage expression and signaling potential of moFcRH1 and moFcRH3 suggest that these receptors will differentially modulate B cell activation and differentiation.
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Acknowledgements |
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Notes |
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Transmitting editor: T. Watanabe
Received 6 May 2004, accepted 30 June 2004.
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References |
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