GENOMICS

13,1082-1087

(19%)

Molecular Cloning of Mouse &-Glycoprotein I and Mapping of the Gene to Chromosome 11 MAYUMI NONAKA, * YOICHI MATsuDA, t TOSHIHIKO SHIROISHI, + KAZUO MORIWAKI, $ MASARU NONAKA, * AND SHUNNOSUKE NATSUUME-SAKAI*,’ *Department of Immunobiology, Cancer Research Institute, Kanazawa University, 13-l Takaramachi, Kanazawa 920, Japan; SDivision of Genetics, National Institute of Radiological Science, Anagawa, Chiba 260, Japan; and *Department of Cytogenetics, National Institute of Genetics, Mishima 411, Japan Received

December

27, 1991;

&-Glycoprotein I (&GPI), a plasma protein that binds to anionic phospholipids, is composed of five repeating units called a short consensus repeat (SCR), which is found mostly in the regulatory proteins of the complement system. Recently the human &GPI gene has been assigned to chromosome 17, not to chromosome 1 where most of the genes of the SCR-containing proteins are clustered. In this report, we have isolated a full-length cDNA clone of mouse &GPI and determined the chromosomal localization of the gene. The amino acid sequence deduced from the nucleotide sequence of mouse &GPI revealed 76.1% identity with that of human &GPI. A genetic mapping by in situ hybridization and linkage analysis using 50 backcross mice has shown that the mouse &GPI gene (designated B2gpl) is located on the terminal portion of the D region of chromosome 11, closely linked to Gfup, and is 18 CM distal to Acrb, extending a conserved linkage group between mouse chromosome 11 and human chromosome 17. On the basis of these results, the evolutionary relationships among the SCR-containing proteins are discussed. LQ1992 Academic Press, Inc.

INTRODUCTION &Glycoprotein I (P,GPI), a human plasma protein also called apolipoprotein H, is known to bind to negatively charged molecules such as platelets, heparin, DNA, and anionic phospholipids, but its physiological function has not been established. However, several findings have suggested a regulatory role for @,GPI in the pathway of blood coagulation (Schousboe, 1985; Nimpf et al., 1986). Recently, McNeil et al. (1990) have reported that anti-cardiolipin antibodies are directed against an Sequence data from this article have been EMBL/GenBank Data Libraries under Accession ’ To whom correspondence should be addressed.

088%7543/92 $5.00 Copyright 0 1992 by Academic Press, Inc. All rights of reproduction in any form reserved.

deposited with No. D10056.

the

1082

revised

April

24, 1992

antigen that includes P2GPI, suggesting that @,GPI is involved in the production of the autoantibodies. The complete amino acid sequence of human P,GPI determined by Lozier et al. (1984) showed that the protein is composed of five contiguous internal repeats, each of about 60 amino acids containing four conserved half-cystine residues. This repetitive unit is termed short consensus repeat (SCR) and is widely distributed in the proteins of the complement system and in some noncomplement proteins (reviewed in Reid and Day, 1989). That is, the regulators of the complement activation (RCA) family, including complement receptor 1 (CRl), complement receptor 2 (CR2), (Yand @chains of C4-binding protein (C4BP), decay acceleration factor (DAF), membrane cofactor protein (MCP), and factor H, as well as the b subunit of coagulation factor XIII (FXIII-b), are constructed with SCRs in most parts of the protein structure. Some other complement proteins, such as Clr, Cls, C2, factor B, C6, and C7, and several noncomplement proteins, such as interleukin-2 receptor, contain one to three SCRs as a part of their domain structure. In addition, the selectin family including leukocyte adhesion molecule 1 (LAM-l), granule-membrane protein 140 (GMP-140), and endothelial leukocyte adhesion molecule 1 (ELAM-1) is known to contain two to nine SCRs in addition to the lectin domain and an epidermal growth factor-like domain. In particular, the genes of the RCA family, FXIII-b, and the selectin family are clustered on human and mouse chromosome 1 (Pardo-Manuel et al., 1990; Rodrigez de Cordoba et al., 1988; Watson et al., 1990; Kingsmore et al., 1989), and evolutionary and/or functional relationships between them are strongly suggested. Recently, the human &GPI gene has been assigned to chromosome 17, not linked to a set of related genes on chromosome 1 (Haagerup et al., 1991). To further explore the evolutionary relationships among the SCRcontaining proteins, we isolated a full-length cDNA clone of mouse /3,GPI and determined the chromosomal localization of the gene by both in situ hybridization and

CLONING

AND

MAPPING

linkage analysis using [(C3H/HeJ X Mus spretus)F, X C3H/HeJ] backcross progeny. Here we demonstrate extensive conservation of the &GPI sequence and chromosomal localization during mammalian evolution.

MATERIALS

AND

METHODS

Isolation of the mouse cDNA clones. To obtain a cDNA probe for screening, we first isolated a full-length cDNA clone encoding human &GPI from a human liver XgtlO library purchased from Clontech (Palo Alto, CA) using two synthesized oligonucleotide probes based on the amino acid sequence of Lazier et al. (1984). One was a positive strand codon-preference 36-mer, 5’ GCTGACTGTGCCAAGTGCACCGAGGAGGGCAAGTGG 3’, corresponding to amino acids 100 to 111; and the other was a mixture of 23-mer, 5’ TT(T/C)TC(T/ C)TT(A/G)TT(C/T)TT(A/G)CA(A/G)AA(A/G)AA 3’, synthesized complementary to the mRNA sequence corresponding to amino acids 279 to 286 (marked by brackets in Fig. 1). A mouse cDNA library constructed using the h ZAP II vector with size-selected poly(A)+ RNA from the livers of BlO.DS/oSn was kindly made available to us by Dr. R. A. Wetsel (Department of Pediatrics, School of Medicine, Washington University, MO) (Wetsel et al., 1990). A plasmid containing human &GPI cDNA was linearized at the 5’ end of the insert, labeled with [32P)UTP using a Riboprobe system (Promega, Madison, WI), and used as a probe for screening the mouse cDNA library. DNA sequencing. The nucleotide sequence was determined by the chain termination method (Sanger et al., 1977) using Sequenase Version 2.0 (United States Biochemical, Cleveland, OH) and [cu-?SdATP with the synthesized oligonucleotide primers as described (Natsuume-Sakai et al., 1990). All nucleotide positions were sequenced at least twice on both strands. In situ hybridization. Preparation of R-banded chromosomes and fluorescence in situ hybridization were performed as described by Matsuda et al. (1992). The mouse &GPI cDNA clone inserted in the pBluescript vector was labeled by nick-translation with biotin-16. dUTP (Boehringer-Mannheim). Excitation wavelengths 450-490 nm (Nikon filter set B-2A and B-2E), 510-560 nm (G-2A), and near 365 nm (UV-PA) were used for observation. Kodak Ektachrome ASA 100 films were used for photomicrographs. Genome mapping. The [(C3H/HeJ X M. spretu.s)F, x C3H/HeJ] backcross progeny were obtained at National Institute of Genetics, Mishima, Japan. The genomic DNAs from livers of 50 backcross mice were prepared by standard techniques. Genome mapping was performed using polymerase chain reaction (PCR) amplification. For @,GPI, the restriction fragment length variant was utilized because preliminary study showed that the C at nucleotide position 414 of the nucleotide sequence of BlO.D2/oSn is the T in the nucleotide sequence of @,GPI of M. spretus, forming a Hi&I site, GAGTC (shown by an arrow and the dotted underline in Fig. l), whereas it is not replaced in the nucleotide sequence of C3H/HeJ. Primer sequences for PCR were 5’ ACCAGCTCATCTAAGTG 3’ (the sense strand) and 5’ CGAGCACAAGCAGGAAT 3’ (the anti-sense strand), corresponding to nucleotides 376-392 and nucleotides 421-437, respectively (underlined in Fig. 1). For markers Acrb (acetylcholine receptor o-chain) and Gfup (glial fibrillary acidic protein), a microsatellite polymorphism was utilized (Love et al., 1990). The primers for Acrb were synthesized on the basis of the reported genomic sequences of mouse acetylcholine receptor &chain for (GT),, repeat (Buonanno et al., 1989), independently of the primers reported by Hearne et al. (1991). Primer sequences were 5’ TGATGTGGTGCTGCTGAACAA 3’ and 5 CAACGTCGAAATTTCCGTCAT 3’, producing a 207-bp fragment with BALB/c. The PCR primers for Gfup were prepared as described by Love et al. (1990, Table 1, No. 39). All PCRs were carried out in 10.~1 volumes containing 50 ng of genomic DNA and 15 pmol of each oligonucleotide primer. Amplification conditions were 95°C for 5 min; 27 to 32 cycles of 95°C for 0.5 min, 50°C or 55°C for 0.5 min, 72°C for 1

OF

1083

&GLYCOPROTEIN

min; followed by 72”C, 5 min. Annealing temperature was 50°C for Gfup and 55°C for P,GPI and Acrb. PCR products for &GPI were analyzed by 10% polyacrylamide gel electrophoresis after incubation with Hid at 37°C for 1 h, and those for Acrb and Gfap were analyzed by 2% agarose gel electrophoresis. The sizes of the amplified PCR products were in agreement with the predicted sizes. Maximum likelihood estimates of recombination probabilities and their standard errors among backcross progeny were calculated according to Green (1981).

RESULTS

The sequence of mouse &GPI. The complete cDNA sequence and the derived amino acid sequence of mouse &GPI are shown in Fig. 1, aligned with those of human &GPI obtained from a cDNA clone that we have isolated. The mouse cDNA clone contained a 21-bp 5’ untranslated region, 57 bp for the leader peptide region (resulting in 19 amino acids), 978 bp for the protein coding region (resulting in 326 amino acids), and a 102-bp 3’ untranslated region followed by a poly(A) tail. The putative polyadenylation recognition signal, AATAAA, was located 20 bp upstream from the poly(A) tail. The predicted amino acid sequence has the same number of amino acids as that of human and is also arranged into five repeating units of approximately 60 amino acids containing four conserved half-cystine residues. The calculated MW is 36,646. Five potential sites for asparagine-linked carbohydrate chain attachment based on the Asn-Xaa-Ser/Thr sequence were found (amino acid positions 86, 98, 143, 164, and 174) (marked by diamonds in Fig. 1). The amino acid identity between human and mouse &GPI was 76.1 and 87.4% when the equivalent amino acid replacements (A = S = T = P = G, N = D = E z Q, H = R = K, M = L = 1 = V, F = Y = W) were taken into account. The nucleotide homology in the protein coding region was 78.1%. This strong homology confirmed that the clone isolated from the mouse library encodes the mouse counterpart of human &GPI. In situ hybridization. To localize the gene encoding mouse &GPI, we first performed in situ hybridization. As shown in Fig. 2, the hybridization signal was present on the terminal portion of the D region of chromosome 11. No signals were seen on other chromosomes, including chromosome 1. Linkage analysis. To confirm this result and define the position of the &GPI gene more precisely, the linkage with two markers on chromosome 11 was investigated using 50 [ (C3H/HeJ X M. spretus)F, X C3H/HeJ] backcross mice. Examples of gene mapping analysis using the PCR amplification method are shown in Fig. 3. A 62-bp PCR fragment for P,GPI produced from M. spretus DNA using the two synthesized primers described under Materials and Methods was split into 36- and 26bp fragments by HinfI, while that obtained from C3H/ HeJ DNA was not digested. The size difference could be clearly resolved by 10% polyacrylamide gel electrophore-

1084

NONAKA

ET

AL.

-19

-1 +1 MVSPVLALFSAFLCHVAIAGRICPKPDDLPFAT MO CGCTGGTGGGACGCATCCGCAATGGTTTCCCCGGTGCTCGCCTTGTTCTCCGCCTTCCTCTGCCATGTTGCTATTGCAG~CG~TCTGTCC~GCCGGAT~CCTACCATTTGCTACG HU .T.....A.....A....T..A......AT.........GAGT..T............................C......C.....A.....TT........T.C..A M I I S T

1:: S

VVPLKTSYDPGEQIVYSCKPGYVSRGGMRRFTCPLTGMWP MO GTTGTCCCCTTAAAGACATCCTACGACCCTGGGGAGCAGATTGTCTACTCCTGC~GCCAGGCTACGTGTCCAGGG~GG~T~~CGGTTTACCTGTCCTCTCACAG~TGTGGCCC Hu ..G.....G.....A....T...T..G..A..A..AG.....ACG..T...........G.....T......C.A............AA.....T...C............C........ F E E T K I

54 240 L

INTLRCVPRVCPFAGILENGIVRYTSFEYPKNISFACNPG MO ATCAACACCCTGAGATGTGTCCCCAGAGTATGTCCTTTCGCTGG~TCTTAG~TGG~TTGTACGCTACACGAGTTTTG~TATCCC~~CATCAGTTTTGCTTGT~CCCTGGG HU . . . . . . ..T....A....ACA.................T.....................GCC........T....C...............C.CG.........T........A..... K T A T N T

+

134 ii0

ACCACCAGTTCCAAAGTiTkACkCiT P

+ KDYRPSAGNNSLYQDTVVFKCLPHFAMIGND MO AAGGATTATAGGCCTTCAGCTGGGAACAACTCTTTGTATCAG~CACAGTGGTCTTT~TGCTTGCCACACTTTGCCAT~TCG~T~C Hu CGT.T . . . ..A...A........A.....T..CC.C....G........CA..T...G....T........ACA...G...TiT....N...T...AIT.~C.....GA~..I.T.....T RV K N R A E 0 H

I

SI

T

T A

6 TVMCTEQGi :ACAGTCATGTGCACAFAArAAccAAAr l",".",""."",~

D

K

A

T

F

,. I, r I ” u I DGPEEAECTKTGAWSFLPTCRESCKLPVKKATV ,,,,,,,,,,,,,,,,,,CJ\CGGCCCAGAI\GAAGCGW\ATGTACC/U\GACGGGAACTTGGTCCTTCTTGCCGACCTGTA~~GTCTTGC~CTCCCCGTT~~GCCAC~GTG TtY TTPT . . . . . . ..I”“...IIbl... . . T . . . . . G . . . . ..ATA...........ACT.....AC.....TGC.A....A.~....~..~.....T...GI.A..T..A..A........ DG S I L N AM

HCRDGTIEIPSCFKEHSSLAFWKTDASELTPC* MO CATTGCAGAGATGGCACTATCGAGATTCCCTCGTGCTTC~G~GCACAGTTCTCTGGCTTTCTGG~CGGATGCATCAG~CTGACACCGTGCTG~GTCGTTTCCAG~TC~T Hu ..G..T.T...............AG.C...AAA...........A.................T........T........C.~TG~A.~G..A....~.G..G....T....T..C.C.C V K 0 I TTACTGGATTGCCT-CAGTAAAAAAAAAAAA C . . . ..A...TAT.G.C.CA

FIG. 1. Alignment of the nucleotide and derived amino acid sequences of mouse and human P,GPI cDNA clones. The shown below the mouse sequences only when differences were observed between mouse and human. To obtain a maximal in the 3’ untranslated region of each nucleotide sequence. The amino acids numbered -19 to -1 represent a portion sequence, while those numbered +1 to 326 represent the amino acids of mature protein regions. Stop codons are indicated the polyadenylation signals are boxed. The amino acid sequences used to construct the probes for isolation of human brackets. The potential N-glycosylation sites are indicated by diamonds. The nucleotide positions of the primers for the linkage analysis of the mouse P,GPI gene are underlined. The nucleotide C indicated by an arrow is substituted by the sequence, forming a Hid site, GAGTC, shown with the dotted underline.

sis. Microsatellite variations in Gfap and Acrb between C3H/HeJ and M. spretus were detected by 2% agarose gel electrophoresis. The segregation pattern is presented in Table 1. The locus of the &GPI gene (designated B2gpl) mapped 18.0 -+ 5.4 CM distal to Acrb and no recombinants were identified between B2gpl and Gfap (95% upper confidence limit r = 5.8). The distance between Acrb and Gfap is in good agreement with the current location (Buchberg et al., 1991). There were no significant relationships be-

G

214 720

254 840 ...

LYQGNRVKIQEQFKNGMMHGDKIHFYCKNKEKKCSYTVEA ~GTACCAAGGGATGAGGGTWV\tATCCAGGAACAGTTTGTGGGAT~TGCATGGCGAC~TTCACTTCTACTGC~~~~~~GTGCAGCTACACTGTGGAGGCT . . . . . . . . ..AcA...A..A.....T......A.A...........A...C.A.....T..T...G~.T~T....~......... . . . . . . . . . . . . T . . . ..T..A.A...T... E V E K L I I

MO TGAATGTCGTATTTGTATTTCTGCTTGTCTTAGGAATCCATCTG~TAGCA Hu AA......AC.C....-..CT..T.CA..CA.....C.T.AT......TTA.

174 mfl 1"" N

W T R L P E CL E V K C P F P P R P E N G Y V N Y P A K P V L L Y K MO TGGACCCGATTGCCAGAATGCCTGGAAGTAAAATGTCCCTTCCCTCCGAGGCCAG~TGGGTATGT~TTATCCTGC~GCCGGTGCTTCTATAT~GGAT~GCCACATTTGGT Hu . . . ..TAA...A.........AG............C..A.....AT.A..A.....C.....A.T......C...........A..~~...T~..C....................C K R S D F

;l

T

S

7

FFLNGTSSSKCTEEGKWSPDIPACARITCPPPPVPKFALL MO TTTTTTCTGAATGGGACCAGCTCATCTAAGTGCACGGAGGCCTGCCCGCC Hu . . . . A . . . . . . . ..CG. TGAT..TG.C........T....................G..GC......TC,......C.....T...,..T......T.CA.A..T.C.......ACA... ,AD A Y EL V t

I-8” U.. nu

94 360

294 960 D 326 1080

1170

human sequences are match, a gap is made of the leader or signal by the asterisks and /3,GPI are shown with PCR amplification in T in the Mus spretus

tween the &GPI gene and the genes on chromosome 1, including Cfh whose gene product, factor H, is a protein constructed with SCRs only, similar to &GPI (data not shown).

DISCUSSION

The complete been determined

cDNA sequence of mouse &GPI has and compared with that of humans.

CLONING

FIG. 2. photographed

Fluorescence with UV-2A

in situ hybridization (a), B-2E (b), B-2A

AND

MAPPING

1085

&-GLYCOPROTEIN

using the cDNA clone of mouse &GPI as a biotinylated (c, e, f, g, h), and G-2A (d) filters. The hybridization

Recently, several cDNA sequences for human &GPI have been published, and our results were in complete agreement with the nucleotide sequences reported by Mehdi et al. (1991) and Matsuura et al. (1991). One nucleotide difference was found in comparison with the b

t32GPI

OF

--62

probe. The metaphase spreads were signals are indicated by the arrows.

nucleotide sequences reported by Kristensen et al. (1991) and Steinkasserer et al. (1991); the G at position 817 is replaced by the T, resulting in an amino acid replacement from Val to Leu. The difference is likely to be a polymorphism, as suggested by Steinkasserer et al. (1991). The clone we isolated from mouse liver has the same number of nucleotides in the leader peptide region, the

bp

-36bp

TABLE

-26bp

Segregation

Locus

Gfap

of t.he &GPI Gene and of Markers in 50 Backcross Mice Number

of recombination

events”

Acrb

H/S

H/H

S/H

H/H

S/H

H/H

B%W

H/S

H/H

H:H

S;H

S/H

H/H

Gfap

H/S

H/H

H/H

S/H

H;H

S;H

19

22

5

4

0

0

Acrb Locus FIG. 3. Demonstration of the segregation patterns of the Hinfldigested PCR products of the @zGPI gene and the PCR products of Gfap and Acrb. Lanes S and H correspond to the parental DNA samples of M. spretus and C3H/HeJ, respectively. Lanes 1 to 10 correspond to the first 10 backcross mice analyzed. The first lane on the left contains the molecular weight markers.

1

pair

Acrb-B2gpI BZgpl-Gfap

Recombinants/total 9/50 o/50

’ Mice were scored as homozygous or heterozygous (S/H) with the Mus * This number represents the 95%

Distance

in CM * SE 18 & 5.4 0 =G 5.P

(H/H) with the C3H/HeJ allele spretus and C3H/HeJ alleles. upper confidence limit.

1086

NONAKA

protein coding region, and the 3’ untranslated region as that of humans. The predicted amino acid sequence showed a high level of identity with human &GPI, indicating that this cDNA clone encodes the full-length mouse counterpart of the &GPI protein. To assess the size of the mRNA of mouse &GPI and investigate the site of synthesis, Northern blotting analysis was performed using the mouse cDNA clone as a probe. A hybridizing band of about 1.2-1.3 kb was detected in poly(A)+ RNA from liver, while no bands were detected in poly(A)+ RNA from kidney, heart, spleen, lung, and brain (data not shown). This result confirms that the clone we isolated is an almost full-length clone and supports the conclusion by Steinkasserer et al. (1991) that hepatocytes are a major site of human &GPI synthesis. A cDNA sequence of rat &GPI (Aoyama et al., 1989) and the amino acid sequence of bovine &GPI (Kato and Enjyoji, 1991) have been reported, although the rat cDNA lacks 145 bp at the N-terminal end of the human and mouse protein coding region and a polyadenylation signal, AATAAA, in the 3’ untranslated region. Matsuura et al. (1991) aligned the amino acid sequences of human, bovine, and rat &GPI, showing the extensive conservation among the three species. Mouse &GPI also showed extensive homology with them. In particular, the nucleotide sequences of mouse and rat &GPI in the corresponding protein coding region (nucleotide positions 232-1056) shared 88.1% homology. However, the amino acid identity in the same region, 82.9%, is rather lower than the nucleotide homology, compared with the identities between the human and rat sequences in the same region, 80.5% in nucleotide and 81.9% in amino acid. Approximately half of the nucleotide substitutions result in the amino acid replacements. However, the biological significance of this finding is not clear at present since the function of @,GPI has not been established. The mouse &GPI gene has been localized to distal chromosome 11 by both in situ hybridization and linkage analysis in this study. Recently, the human &GPI gene has been assigned to chromosome 17 (Haagerup et al., 1991). Comparative mapping of the human and mouse genomes has revealed that mouse distal chromosome 11 has extensive homology with human chromosome 17 (Buchberg et al., 1989). The homolog of every gene that has been mapped to human chromosome 17 has been located on mouse chromosome 11, including the human homolog of Gfap, which has been recently assigned to chromosome 17 (Brownell et al., 1991). Therefore, our data extend the degree of linkage conservation. Moreover, the human P,GPI gene has been located 5 CM from HOX2. Since the mouse &GPI gene is tightly linked to Gfap, which is located 3 CM distal to Hox-2 (Buchberg et al., 1991), the distance between Hox-2 and the ,&GPI gene is in good agreement with the linkage data for their human homologs. The short consensus repeat, the constituent domain of &GPI, is widely distributed in the complement proteins and some other proteins. In particular, &GPI, the RCA family, and FXIII-b are composed of SCRs in most parts of the structure. Furthermore, the genes of the hu-

ET AL.

man RCA family and FXIII-b are all located on chromosome lq32 closely linked with each other, suggesting a strong evolutionary relationship among them. Likewise, the genes of their mouse homologs are also located on the homologous regions of chromosome 1, except for Crry (mouse homolog of human CRl) and Cr2, which are located on the most telomeric portion of the chromosome. However, it has been found that the &GPI gene is not linked to the RCA gene family, including FXIII-b in both man and mouse. The significant degree of conservation of the &GPI gene between man and mouse in the chromosomal localization suggests that there was no close linkage between the &GPI gene and the RCA gene family even before the divergence of man and mouse. ACKNOWLEDGMENTS We thank Dr. R. A. Wetsel for providing us with the cDNA library constructed from the livers of BlO.DP/oSn mice. We also thank M. Maemori for her technical assistance with in situ hybridization. This work was supported in part by a Grant-in-Aid from the Ministry of Education, Science and Culture of Japan.

REFERENCES Aoyama, Y., ture of rat Brownell, E., and Bayney, tic-specific 1087-1089.

Chan, Y.-L., and Wool, I. G. (1989). The primary struc&-glycoprotein I. Nucleic Acids Res. 17: 6401. Lee, A. S., Pekar, S. K., Pravtcheva, D., Ruddle, F. H., R. M. (1991). Glial fibrillary acid protein, an astrocymarker, maps to human chromosome 17. Genomics 10:

Buchberg, A. M., Brownell, E., Nagata, S., Jenkins, N. A., and Copeland, N. G. (1989). A comprehensive genetic map of murine chromosome 11 reveals extensive linkage conservation between mouse and human. Genetics 122: 153-161. Buchberg, A. M., Moskow, J. J., Buckwalter, M. S., and Camper, S. A. (1991). Mouse chromosome 11. Mammalian Genome 1: S158-S191. Buonanno, A., Mudd, J., and Merlie, J. P. (1989). Isolation and characterization of the 0 and t subunit genes of mouse muscle acetylcholine receptor. J. Biol. Chem. 264: 7611-7616. Green, E. L. (1981). Linkage, recombination ics and Probability in Animal Breeding Ed.), pp. 77-113, MacMillan, New York. Haagerup, A., Kristensen, T., and Kruse, T. and genetic mapping of the gene encoding to chromosome 17. Cytogenet. Cell Genet.

and mapping. Experiments”

In “Genet(E. Green,

A. (1991). Polymorphism human &glycoprotein 58: 2004-2012.

I

Hearne, C. M., McAleer, M. A., Love, J. M., Aitman, T. J., Cornall, R. J., Ghosh, S., Knight, A. M., Prins, J.-B., and Todd, J. A. (1991). Additional microsatellite markers for mouse genome mapping. Mammalian Genome 1: 273-282. Kato, H., and Enjyoji, K. (1991). Role of & glycoprotein I (&GPI) in surface-mediated activation of factor XII and prekallikrein. Thromb. Haemostasis 65: 1246. Kingsmore, S. F., Vik, D. P., Curtz, C. B., Leroy, P., Tack, B. F., Weis, J. H., and Seldin, M. F. (1989). Genetic organization of complement receptor-related genes in the mouse. J. Exp. Med. 169: 1479-1484. Kristensen, T., Schousboe, I., Boel, E., Mulvihill, E. M., Hansen, R., Meller, K. B., Moller, N. P. H., and Sottrup-Jensen, L. (1991). Molecular cloning and mammalian expression of human &glycoprotein I cDNA. FEBS L&t. 289: 183-186. Love, J. M., Knight, A. M., McAleer, M. A., and Todd, J. A. (1990). Towards construction of a high resolution map of the mouse genome using PCR-analysed microsatellites. Nucleic Acids Res. 18: 4123-4130.

CLONING

AND

MAPPING

Lozier, J., Takahashi, N., and Putnam, F. W. (1984). Complete amino acid sequence of human plasma /3x-glycoprotein I. Proc. Natl. Acad. Sci. USA 81:3640-3644. Matsuda, Y., Harada, Y.-N., Natsuume-Sakai, S., Lee, K.-H., and Chapman, V. M. (1992). The use of replication R- and G-banding of mouse chromosomes in diploid lymphocytes in the fluorescence in situ hybridization (FISH) analysis of the mouse complement factor H gene. Cytogenet. Cell Genet., in press. Matsuura, E., Igarashi, M., Igarashi, Y., Nagae, H., Ichikawa, K., Yasuda, T., and Koike, T. (1991). Molecular definition of human & glycoprotein I (&-GPI) by cDNA cloning and inter-species differences of &-GPI in alternation of anticardiolipin binding. Znt. Zmmunol. 3: 1217-1221. McNeil, H. P., Simpson, R. J., Chesterman, C. N., and Krilis, S. A. (1990). Anti-phospholipid antibodies are directed against a complex antigen that includes a lipid-binding inhibitor of coagulation: f12glycoprotein I (apolipoprotein H). Proc. Nut/.. Acad. Sci. USA 87: 4120-4124. Mehdi, H., Nunn, M., Steel, D. M., Whitehead, A. S., Perez, M., Walker, L., and Peeples, M. E. (1991). Nucleotide sequence and expression of the human gene encoding apolipoprotein H (fix-glycoprotein I). Gene 108: 293-298. Natsuume-Sakai, S., Nonaka, M., Nonaka, M., Harada, Y., Shreffler, D. C., and Moriwaki, K. (1990). Demonstration of an unusual allelic variation of mouse factor H by the complete cDNA sequence of the H.2 allotype. J. Zmmunol. 114: 358-362. Nimpf, J., Bevers, E. M., Bomans, P. H. H., Till, U., Wurm, H., Kostner, G. M., and Zwaal, F. A. (1986). Prothrombinase activity of human platelets is inhibited by &glycoprotein-I. Biochem. Biophys. Acta 884: 142-149.

OF

1087

&GLYCOPROTEIN

Pardo-Manuel, F., Rey-Campos, J., Hillarp, A., Dahlblick, B., and Rodriguez de Cordoba, S. (1990). Human genes for the (Y and fi chains of complement C4b-binding protein are closely linked in a head-totail arrangement. Proc. Natl. Acad. Sci. USA 87: 4529-4532. Reid, K. B. M., and Day, ships of the complement 180.

A. J. (1989). components.

Structure-function Zmmunol. Today

relation-

10: 177-

Rodriguez de Cordoba, S., Rey-Campos, J., Dykes, D. D., McAlpine, P. J., Wong, P., and Rubinstein, P. (1988). Coagulation factor XIII B subunit is encoded by a gene linked to the regulator of complement activation (RCA) gene cluster in man. Immunogenetics 28: 452-454. Sanger, F., Nicklen, S., and Coulson, with chain terminating inhibitors. 5463-5467. Schousboe, I. (1985). &Glycoprotein tact activation of the intrinsic 66: 1086-1091.

A. R. (1977). DNA sequencing Proc. Natl. Acad. Sci. USA 74: I: A plasma inhibitor of the conblood coagulation pathway. Blood

Steinkasserer, A., Estaller, C., Weiss, E. H., Sim, R. B., and Day, A. J. (1991). Complete nucleotide and deduced amino acid sequence of human &-glycoprotein I. Biochem. J. 277: 387-391. Watson, M. L., Kingsmore, S. F., Johnston, G. I., Siegelman, M. H., Le Beau, M. M., Lemons, R. S., Bora, N. S., Howard, T. A., Weissman, I. L., McEver, R. P., and Seldin, M. F. (1990). Genomic organization of the selectin family of leukocyte adhesion molecules on human and mouse chromosome 1. J. Exp. Med. 1'72: 263-272. Wetsel, R. A., Fleischer, D. T., and Haviland, D. L. (1990). Deficiency of the murine fifth complement component (C5). A 2-base pair gene deletion in 5’-exon. J. Biol. Chem. 266: 2435-2440.