GENOMICS 9, :3$6-s’% (1991)

SHORT COMMUNICATION

Localization of the Gene for Plasma Retinol Binding Protein to the Distal Half of Mouse Chromosome 19

MEENA CHAINANI,* BONNIE kwsu..,t AND ROSEMARY W. ELLIOTT*

*Department of Moiecuiar and Celiuiar Biology, Rosweii Park Cancer institute, New York State Department of Heaith, 666 Elm Street, Buffalo, New York 14263; and tDepartment of Biology, State University College at Buffalo,

1300 Elmwood Avenue, Buffalo, New York 14222

Received June 18. 1990. revised October 8. 1990

A DNA polymorphism for the mouse retinol binding pro- tein has been identified using the enzyme BamHI and a rat partial cDNA probe. Analysis of the polymorphism in DNA from 64 inbred mouse strains demonstrated the presence of a single gene with two alleles, Rbp-4 b and Rbp-4d. Compari- son of the segregation patterns of these alleles in three sets of recombinant inbred strains with allele segregation pat- terns of previously characterized loci shows that the Rbp-4 locus is closely linked to the locus for phenobarbital-induc- ible cytochrome P450-2c (Cyp-2~) that has been shown by in situ hybridization to lie on chromosome 19, bands Dl- D2. The Rbp-4 locus is just proximal to Cyp-2c at the distal end of chromosome 19. k. IWI Academic PRSS. he.

Retinoids are a class of molecules including retinol, retinaldelhyde, retinoic acid, and their synthetic ana- logs. The retinoids are fundamental mediators of cell differentiation and cell proliferation (Goodman, 1984). Vitamin A (retinol) is an essential component of the mammalian diet because it is necessary for growth, differentiation of epithelial tissue, mainte- nance of reproductive capacity, glycoprotein produc- tion, and prevention of night blindness. Like other fat-soluble vitamins, vitamin A is poorly soluble in plasma and other biological fluids and must complex with specialized carrier molecules to reach its site of action in the body. There are four retinol binding pro- teins (RBPs), each encoded by its own gene and per- forming a unique function. The genes for RBPl and RBPS, the two intracellular RBPs (CRBPs), are closely linked and located on mouse chromosome (Chr) 9 (Demmer et al., 1987). They are also syntenic on human Chr 3 (Demmer et al., 1987), and RBPl has been localized to 3q21-q22 (Rocchi et al., 1989). It is responsible for transport inside the target cell, while RBP2 is responsible for transport inside intestinal

cells. The locus for RBP3, the interstitial (or inter- photo) receptor, responsible for transport in the ret- ina, has been mapped to Chr 11 in the mouse (Kozak et al., 1989) and to Chr lOq11.2 in man (White et al., 1990). The locus for plasma RBP, also called RBP4, has been localized in man to lOq23-q24.

The gene encoding plasma RBP (Rbp-4) is a member of a large gene family whose products are secreted into various body fluids. The genes retain similarity in sequence and intron/exon organization. The proteins also show significant amino acid se- quence similarities and many of them function in binding small hydrophobic molecules. Proteins in this family include plasma retinol binding protein, fi-lac- toglobulin (Papiz et al., 1986), a,-acid glycoprotein AGP (Pervaiz and Brew, 1987), apolipoprotein D (Drayna et al., 1987), insect bilin binding protein (Holden et al., 1987; Huber et al., 1987), bovine pyra- zine binding protein (Cavaggione et al., 1987), frog olfactory-specific Bowman’s gland protein (Lee et al., 1987), rat olfactory odorant binding protein (Pevsner et al., 1988), a rat salivary gland protein (Schmale et al., 1990), and the mouse major urinary proteins (MUP) as well as the homologous rat urine cu,-mi- croglobulin.

In the mouse, genes for the MUP multigene family and the two AGP proteins have been mapped to Chr 4 (Bennett et al., 1982; Bishop et al., 1982; Krauter et al., 1982; Baumann and Berger, 1985). Since gene fam- ilies are thought to arise through gene duplication and subsequent sequence divergence, we thought it of in- terest to see if another member of the gene family, Rbp-4, is located on this chromosome. Our data indi- cate that the mouse Rbp-4 locus is unlinked to the other mapped genes in this family; instead it is lo- cated on mouse chromosome 19.

To search for a polymorphism for Rbp-4, genomic

088%754:1/91 $3.00 Copyright ril 1991 by Academic Press, Inc. All rights of reproduction in any form resrrvrd.

376

SHORT COMMUNICATION 377

TABLE 1

Distribution of Rbp-4 Alleles among Inbred Mice

BUmHI Allele fragment Mouse strains

b :%.6 kh AEJ/GnRk, AU/&J, BNT/Ha, C57BL/J6,* C57BL,‘lOJ,” C57BR,‘cd.J,* (:57BL/KsJ, C57/e/Ha, (‘571,/J. FS/EiJ, IS/CamEid, LT/ReSv,J, MA/My-J, RIIIs/.J, SF/ CamEi.J, SHR/Gn, STX/Le, YEBT/Ha

d ‘i.3 kl, A/.J, ABP/I,eJ, AKR/J,$ BALB/cByJ,f BART/Ha. BDP/J, BLJB/BnJ, CBA/ Cad,t CBA/.J. CE/J,$ C3H/He.J, C3Heb/ Fe,J. DBA/lJ. DBA/2J,t DBA/2Ha, DBA/De.J. DW/J. GRS/A,$ HRS/J, I/ LnJ. I/St-J, IC/LeJ. ICR/Ha. JE/Le.J, LG/,J, LP/,J MOLD/Rk, NZB/BlNJ, OBO/A,t P/J, PL/J, RF/.J, RSV/LeJ, SB/ Le,J. SEA/GnJ, SEC/lRe,J. S.JL/J, SM/J. ST/b,J,$ STS/A. SWR/.J. WB/Re.J. YBR/Ki. 101. 129/J:

* Strain carries the (‘*VP-zc” allele. t Strain carries the Cq-2cb allele. 1 Strain carries the C>p-Zc’ allele.

DNA from 64 inbred strains was digested with BamHI and analyzed by the method of Southern (1975) using t.he 256-bp BamHI-PstI fragment of the rat retinol binding protein cDNA probe. Only two al- leles were observed among the 64 inbred strains (Ta- ble 1). The 3.6-kb DNA fragment is associated with the Rbp-4’ allele, while the 7.3-kb fragment is asso- ciated with the Rbp-4d allele (Fig. 1). In heterozygous animals both fragments were observed.

To determine the map position of the Rbp-4 gene, DNA from animals of the BXD, AKXL, and BXH RI st.rain sets were analyzed for the segregation of the BamHI polymorphic fragments, since the parental strains of each RI set differed in their Rbp-4 alleles. Only parental fragments were observed, indicating that the cDNA probe detected a gene encoded by a single locus. Strain distribution patterns for alleles of Rbp-4 (Table 2) showed complete concordance with the alleles of C-vp-2c (cytochrome P450, subfamily IIc, phenobarbital-inducible) (Meehan et al., 1988) in the 18 AKXL and the 12 BXH RI strains, while there were three recombinants in the 26 BXD RI strains. The probability of linkage between the two loci is greater than 99.9% (Silver and Buckler, 1986) and the distance estimate between the two loci is 1.5 ? 0.7 map units. As the locus for cytochrome Cyp-2c was assigned to bands Dl/D2 of Chr 19 in the mouse by Meehan et al. (1988) by in situ hybridization, Rbp-3 must also be also on Chr 19, probably in the same chromosomal band. Furthermore, comparison of BXD RI strains SDPs with those of Ly-20 (Hogarth et al., 1988), located proximally on Chr 19, suggests that

Rbp-4 lies proximal to Cyp-Bc, predicting the order Ly- 1 O-Rbp-4-Cyp-2c. A further confirmation of close linkage between Rbp-4 and Cyp-SC is shown in Table 1. Among the inbred strains typed for both loci, all strains carrying the Cyp-2~” allele also carry the Rbp- 4’ allele. Furthermore, all strains carrying either the Cyp-2cb or the Cyp-2c” allele carry the Rbp-4d allele. Thus only three of the six allele combinations for the three loci are found. This linkage disequilibrium sug- gests that the two loci are tightly linked (Murray et al., 1987).

The human gene CYP2c has been localized to Chr lOq24.1-q24.3, linked to the human gene for plasma ret,inol binding protein, RBP4, which has been local- ized to Chr 1Oq23-q24 (Rocchi et al., 1989). As our data show, this linkage has been preserved in the mouse. Conservation of synteny and linkage of genes between man and mouse has been established for sev- eral regions of the genome. A conserved syntenic seg- ment has already been established for the distal end of human Chr 1Oq and the distal end of mouse Chr 19 (Nadeau and Reiner, 1989) and includes the locus for glutamate oxalase transaminase-1, which has been regionally localized in both genomes, as well as loci for lysosomal acid lyase, phosphoglyceromutase-1, and terminal deoxynucleotidyltransferase, which have not yet been localized in either genome. The data on Cyp-2c and Rbp-4 serve to extend this conserved re- gion from Got-l toward the distal end of mouse Chr 19, and from GOT1 toward the centromere on human Chr 1Oq.

7 a Kb

9.4 7.3 Irl *1*rrr

6.7

4.4

3.6

2.3

FIG. 1. RamHI fragment length polymorphism detected with the 266.bp RamHI-p,s’stI fragment of the cloned plasma retinol binding protein, pRbp-2. Mouse DNA was isolated from frozen livers as described in Mann et al. (15). Ten to fifteen micrograms of DNA was digested with BornHI under conditions described by the manufacturer (Bethesda Research Laboratories). DNA fragments were separated in 0.8% agarose (Seakem. FMC Carp) gels blotted to Zetabind (AMF-Curio). Probe DNA was labeled using the method of Feinberg and Vogelstein (7) and prehybridization and hybridization conditions were as described in Mann et al. (15). The clone, pRbp-2, was the kind gift of Dr. Brehon C. Laurent of Wal- lenberg Laboratories, Ilppsala, Sweden (27). It consists of 740.bp rat RBP cDNA cloned into pUC9.

378 SHORT COMMLJNICATION

TABLE 2

Segregation of Alleles at Rbp-4 and Flanking Loci in Three Sets of RI Strains

BXD

1 2 5 6 8 9 11 1% 13 14 15 16 18 19 20 21 ‘” 23 24 25 27 28 29 :w :11 32

LF-IO DBBDDBBBDBBBBBDBB D L) B LJ B H B D B

Rbp-4 DBBDDBBB ii B B B B h D B H I1 iBBHBR ii B

Cyp-2c D B B D D B ;5 H B B B B B D D ;r; R I) B R B B H B B k

AKXI,

5 6 ‘7 8 9 12 13 14 16 17 19 21 24 “5 ‘8 29 37 38

L.%-10 A A A L A U L A L L L I, A A A A A L x

Rbp-l A I, : ; A A L I, L L L A x x

i I, I, A A I, Cypp-2c A 1, L L A A L I, L L L A A L I, A A 1,

BSH

2 3 4 6 7 8 9 IO 11 12 14 19

L\,- 1 HHHBBHHBB B H H

Rbp-=I HB;;H;1B tI ifI ii B ;r C’ypp-2c H B B B H H B H H B B H

Interval R N % Recombination (*SD) 95% Conf. limits 99% Conf. limits

Ly-lO.l-Rbp-3 17 5 4 14.9 f 5.7 6.9-86 5.5-49.1 Rbp-l-Cyp-:! 3 56 1.5 f 0.9 O.L4 0.2-6.3

Note. A, B. D, H, and L are generic symbols for alleles inherited from strains AKR/J, C57BL/6J, DBA/2J, C3H/HeJ, and C57L/J, respectively. A crossover is indicated hy an x. A U indicates that the allele is not known. R represents the number of recombinants in N strains for a given interval. Strain distribution patterns for Rbp-4 were determined in this study. Strain distribution patterns for other loci have been published (9, 16, 28). Analysis of the segregation of alleles in RI strains was performed by methods described previously (24,25).

ACKNOWLEDGMENT

This work was supported by Grant GM28464 from the National Institutes of Health, which is not responsible for its contents nor do they necessarily represent the official views of that agency.

5.

REFERENCES

BAUMANN, H., AND BERGER, F. G. (1985). Genetics and evo- lution of the acute phase proteins in mice. Mol. &n. &net. 210:505%512.

6.

BENNETT, K. L., LALLEY, P. A.. BARTH, R. K., AND HASTIE, N. D. (1982). Mapping the structural genes coding for the major urinary proteins in the mouse: Combined use of recom- binant inbred strains and somatic hybrids. Proc. N&l. Acad. Sci. USA 79: 1220-1224.

7.

BISHOP, J. O., CLARK, A. J., CLISSOLD, P. M., HAINEY, S., AND FRANKE, U. (1982). Two main groups of mouse major urinary protein genes, both largely located on chromosome 4. EMBO J. 1: 615-620.

8.

CAVAGGIONE, A., SORBI, R. T., KEEN, ,J. N., PAPPIN, D. .J. C., AND FINDLEY, J. B. C. (1987). Homology between the pyra-

9.

10.

zinc-binding protein from the nasal mucosa and the major urinary proteins. &X&S Lett. 212: 225-228.

DEMMER, L. A., BIRKENMEIER, E. H., SWEETSER, L). A., LE- VIN, M. S., ZOLLMAN, S.. SPARKES, R. S., MOHANDAS. T., LUSIS, A. cJ.. AND GORDON, J. I. (1987). The cellular retinol hinding protein II gene: Sequence analysis of the rat gene, chromosomal localization in mice and humans, and documen- tation of its close linkage to the cellular retinol binding pro- tein gene. J. Bid. C’hem. 262: 2458-2467.

DRAYNA, D. T., MCLEAN, J. W., WION, K. L., TRENT, .J. M., DRABKIN, H. A.. AND LAWN. R. M. (1987). Human apolipo- protein D gene: Gene sequence, chromosome localization, and homology to the a2u-globin superfamily. DNA 6: 199-204.

FEINBERG, A. P.. AND VOGELSTEIN, 13. (1983). A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity. Anal. Biochem. 132: 6-13.

GOODMAN, D. S. (1984). Biosynthesis, absorption and hepatic metabolism of retinal. In “The Retinoids” (M. B. Sporn, A. B. Roberts, and D. S. Goodman, Eds.), Vol. 2, pp. 64-85. Academic Press, New York.

Deleted in proof.

HOGARTH, I’. M.. HOULDEN, B. A., LATHAM, S. E., CHERRY,

SHORT COMMCJNICATION 379

11.

1’.

13.

14.

15.

16.

17.

18.

19.

20.

M., TAYLOR, B. A., AND MCKENZIE, 1. F. C. (1988). The mouse L-v-12.1 specificity: Genetic and biochemical relation- ship to Ly- 1. Immunogenetics 27: 383-387.

HOLDEN, H., RYPNIEWSKI, W., LAW, J., AND RAYMENT, R. (1987). The molecular structure of insectcyanin from the to- bacco hormworm Manduca sexta I. at 2.6 A resolution. EMHO J. 6: 1565- 1570.

HUBER, R., SCHNEIDER, M., EPP, O., MAYR, I., MESSER- SCHMIDT, A., AND PFLUGRATH, J. (1987). Crystallization, crystal structure analysis and preliminary molecular model of the bilin binding protein from the insect Pieris brassicae. J. Mol. Biol. 94: 414-417.

KOZAK, C., DANCIGER, M., AND FARBER, D. (1989). Chromo- some assignment of the genes for several proteins involved in visual transduction. Cytogenet. Cell Genet. 51: 1025-1026.

KRAUTER, K., LEINWAND, L., D’EUSTACHIO, P., RUDDLE. F., AND DARNELL. J. E. (1982). Structural genes of the mouse major urinary protein are on chromosome 4. J. Cell Biol. 94: 414-417.

LEE, K.-H.. WELLS, R. G., AND REED, R. R. (1987). Isolation of an olfactory cDNA: Similarity to retinol-binding protein suggests a role for olfaction. Science 235: 1053-1056.

MANN, E. A.. SILVER, L. M., AND ELLIOTT, R. W. (1986). Ge- netic analysis of a mouse t complex locus that is homologous to a kidney cDNA clone. Genetics 114: 993-1006.

MEEHAN, R,. R.. SPEED, R. M.. GOSDEN, J. R., ROUT, D., HUT- TON, J. .J.. TAYLOR, B. A., HILKENS, J., KROEZEN, V., HILGERS, J., ADESNIK, M., FREIDBERG, T., HASTIE, N. D., AND WOLF, C. R. (1988). Chromosomal organization of the cytochrome P450-2C gene family in the mouse: A locus asso- ciated with constitutive aryl hydrocarbon hydroxylase. Pmt. Natl. Acad. Sci. IJSA 85: 2622-2666.

MURRAY, J. C., BUETOW, K. H., DONOVAN, M., HORNUNG, S., MOTULSKY, A. G., DISTECHE, C., DYER, K., SWISSHELM, K., ANDERSON, ,J.. GIBLETT, E., SADLER, E., EDDY, R., AND SHOWS, T. 13. (1987). Linkage disequilibrium of plasminogen polymorphisms and assignment of the gene to human chro- mosome 6q26-6q27. Amer. J. Hum. Genet. 40: 338-350.

NADEAU, -7. H., AND REINER, A. H. (1989). Linkage and syn- teny homologies in mouse and man. In ‘*Genetic Variants and Strains of the Lahoratory Mouse” CM. F. Lyon and A. J. Searle, Eds.), 2nd ed., pp. 506-536, Oxford Univ. Press, Ox- ford.

PAPIZ, M. Z.. SAWYER, L., ELIOPOULOS. E. E.. NORTH, A. C. T.. FINDLAV, J. B. C., SIVAPRASADARIO, R., *JONES, T. A., NEWCOMER. M. E.. AND KRAULIS. (1986). The structure of

21.

22.

23

24.

25.

26

27.

28.

29

30.

31

beta-lactoglobulin and its similarity to plasma retinol binding protein. Nature (London) 324: 383-385.

PERVAIS, S., AND BREW, K. (1987). Homology and structure- function correlations between alpha l-acid glycoprotein and serum retinol-hinding protein and its relatives. FASEB J. 1:

209-214.

PEVSNER, J., REED, R. R., FEINSTEIN, P. G., AND SNYDER, S. H. (1988). Molecular cloning of odorant-binding protein: Member of a ligand carrier family. Science 241: 336-339.

ROCCHI, M., COVONE, A., ROMEO, G., FARAONIO, R., AND Co-

LANTUONI, V. (1989). Regional mapping of RBP4 to lOq23- q24 and RBPl to 3q21--q22 in man. Somat. I’ell Mol. Grnet. 15: 185-190.

SCHMAI,E, H., HOLTGREVE-GREZ, H.. AND CHRISTIANSEN, H. (1990). Possible role for salivary gland protein in taste recep- tion indicated by homology to lipophilic-ligand carrier pro- teins. Nature (London) 343: 366-369.

SILVER, J. (1985). Confidence limits for estimates ofgene link- age based on analysis of recombinant inbred strains. J. Hered. 76: 4X6-440.

SILVER, J., AND BUCKI,ER, C. (1986). Statistical consider- ations for linkage analysis using recombinant inbred strains and backcrosses. Proc. Natl. Acad. Sci. ITSA 83: 1423-1427.

SOUTHERN, E. M. (1975). Detection of specific sequences among DNA fragments separated by elrctrophoresis. J. Mol. iBid. 98: 503-517.

SJNDELIN, .J., LAIJRENT, B. C., ANUNDIT, H.. TRAGARDH, L., LARHAMMAR, D., B.JORCK, I,., ERIKSSON, U.. AKERSTROM, B., CJ~~~~, A., NEWCOMER, M., PETERSON, P. A., AND RASK, I,.

(1985). Amino acid sequence homologies between rabhit, rat, and human serum retinol-binding proteins. J. Riol. C’hem. 260: 6472-6480.

TADA, N.. KIMURA, S.. AND HAMMERLINC.. U. (1982). Immu- nogenetics of mouse B-cellalloantigen systems defined hy

monoclonal antibodies and gene cluster formation of these loci. Immunol. Ret>. 69: 99-125.

TAYLOR, B. A. (1989). Recombinant inbred strains. In “Ge- netic Variants and Strains of the Laboratory Mouse” (M. F. I,yon and A. .J. Searle, Eds.), 2nd. ed., pp. 773-796, Oxford Univ. Press, Oxford.

WHITE, R. I,.. LALOUEL,. J.-M., NAKAMURA, Y., DONIS- KELI~ER. H.. GREEN, I’.. BOWDEN, D. W., MATHEW, C. G. P.,

EASTON. D. F., ROBSON, E. B.. MORTON, N. E.. GUSELLA, .J. F., HAINES, .J. L., RETIEF, A. E.. KIDD, K. K., MURRAY, d. C.. LATHROP. G. M., AND CANN, H. M. (1990). The CEPH consortium primary linkage map of human chromosome 10. Gcnomics 6: 39:1-412.