A molecular genetic linkage map of mouse chromosome 18 reveals extensive linkage conservation with human chromosomes 5 and 18

GENOMICS

13, l%l-1288

(19%)

A Molecular Genetic Linkage Map of Mouse Chromosome 18 Reveals Extensive Linkage Conservation with Human Chromosomes 5 and 18 MONICA J. JUSTICE,* DEBRA J. GILBERT,* KENNETH W. KINzLER,t BERT VOGELSTEIN, t ARTHUR M. BUCHBERG, *,’ JEFFREYD. CECI,* YOICHI MATSUDA,* VERNE M. CHAPMAN,* CHRISTOS PATRIOTIS, !j ANTONIOS MAKRIS, 5 PHILIP N. TSICHLIS, § NANCY A. JENKINS,* AND NEAL G. COPELAND” *Mammalian Genetics Laboratory, ABL-Basic Research Program, NC/-Frederick Cancer Research and Development Center, P.O. Box B, Frederick, Mary/and 21702; tMo/ecular Genetics Laboratory, The Johns Hopkins Oncology Center, 424 North Bond Street, Baltimore, Maryland 21237; *Department of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo, New York 14263; and § Department of Molecular Oncology, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19 17 7 Received

March

27, 1992;

An interspecific backcross between C57BL/6J and Mus spretus was used to generate a molecular genetic linkage map of mouse chromosome 18 that includes 23 molecular markers and spans approximately 86% of the estimated length of the chromosome. The Ape, Camk2a, Dl8FcrZ, DlSFcr2, DlSLehl, Dl8Leh2, Dee, Emb-rs3, Fgfa, Fim-S/Csfmr, Gnal, Grl-1, Grp, Hk-lrsl, Ii, Kns, Lmnb, Mbp, Mcc, Mtv-38, Palb, Pdgfrb, and Tpl-2 genes were mapped relative to each other in one interspecific backcross. A second interspecific backcross and a centromere-specific DNA satellite probe were used to determine the distance of the most proximal chromosome 18 marker to the centromere. The interspecific map extends the known regions of linkage homology between mouse chromosome 18 and human chromosomes 5 and 18 and identifies a new homology segment with human chromosome lop. It also provides molecular access to many regions of mouse chromosome 18 for the first time. o 1992 Academic PEW, IDC.

INTRODUCTION

Molecular genetic linkage maps of the mouse genome have proven invaluable for establishing comparative maps of mouse and human (or other) genomes, studying genome organization and chromosome evolution, identifying genes encoding known mutations, and identifying mouse models of human disease. Interspecific backcrosses (IBs) between distantly related species of mice provide a powerful method for establishing multilocus The U.S. Government’s right to retain a nonexclusive royalty-free license in and to the copyright covering this paper, for governmental purposes, is acknowledged. ’ Current address: Department of Microbiology and Immunology, Jefferson Medical College, 233 S. 10th Street, Philadelphia, PA 1910’7.

revised

May

15, 1992

maps of the mouse genome (reviewed by Guenet, 1986; Avner et al., 1988; Copeland and Jenkins, 1991). The evolutionary distance between mouse species involved in an IB has allowed for the accumulation of DNA sequence differences (Bonhomme et al., 1984) that facilitate the detection of restriction fragment length polymorphisms (RFLPs). These RFLPs can be used to establish the map location of virtually any molecular marker (Ferris et al., 1982; Robert et al., 1985). We have used an IB involving the inbred laboratory mouse strain C57BL/6J and the wild mouse species Mus spretus to create a multilocus molecular genetic linkage map of mouse chromosome (Chr) 18. Nine of the 23 loci included in this study have been mapped previously to mouse Chr 18 by IB or intraspecific backcross analysis. These loci include Dl8Lehl and D18Leh2 (Cox et al., 1991), Fgfu (Cox et al., 1991), Fim-B/Csfmr (Sakai et al., 1991; Siracusa et al., 1991), G&I (Cox et al., 1991; Oakey et al., 1991; Sakai et al., 1991), Ii (Cox et al., 1991), Mbp (Cox et al., 1991; Oakey et al., 1991; Sikela et al., 1990; Sakai et al., 1991; Siracusa et al., 1991), Mtv-38 (Siracusa et al., 1991), and Pdgfrb (Oakey et al., 1991). The map locations of the remaining 14 loci (Ape, Camk2u, DlBFcrl, D18Fcr2, Dee, Emb-rs3, Gnal, Grp, Hk-lrsl, Kns, Lmnb, Mcc, Palb, and Tpl-2) are reported here for the first time. The position of the most proximal marker relative to the centromere was also determined with a second IB (Brannan et al., 1992), which was analyzed for the centromere using a DNA satellite probe that specifically detects C57BL/6Ros centromeres (Matsuda and Chapman, 1991). These mapping studies provide an unambiguous orientation of 23 molecular markers that span approximately 86% of the estimated genetic length of mouse Chr 18, identify a new homology segment of human chromosome lop, extend the regions of known link-

1281 All

0888-7543/92 $5.00 Copyright 0 1992 by Academic Press, Inc. rights of reproduction in any form reserved.

JUSTICE

age homology with human chromosomes 5 and 18, and provide molecular access to many regions of Chr 18, which may eventually provide molecular access to mutations on the chromosome. MATERIALS

AND

METHODS

Mice. The interspecific backcross (C57BL/6J x M. spreta.s)F, X C57BL/6J was performed at the NCI-Frederick Cancer Research and Development Center as described by Copeland and Jenkins (1991). Subsets of a total of 205 mice were used in the mapping studies. The (C57BL/6Ros X M. spretus)F, X C57BL/6Ros and (C57BL/6Ros X M. spretus)F, X M. spretus interspecific backcrosses were performed at the Roswell Park Cancer Institute and analyzed for centromeric satellite sequences as described by Brannan et al. (1992). Probes. The probes for the adenomatosis polyposis coli gene (Apt) were: (1) a mouse 1.1.kb EcoRV cDNA fragment cloned into pBluescript SK (mAPC 73.lA), and (2) a 327-bp human DNA fragment amplified by the polymerase chain reaction (PCR) and purified in agarose (L.-K. Su and K. W. Kinzler, unpublished data). The probe for the calmodulin-dependent protein kinase type IIa gene (Camk2a) was a rat 4.5-kb cDNA fragment cloned into pBR322 (pk2a-3; Tobimatsu and Fujisawa, 1989), a gift from H. Fujisawa (Asahikawa Medical College, Asahikawa, Japan). The probe for the DNA fragment, Frederick Cancer Research-l (D18Fcrl) was a mouse 2.8kb EcoRI cDNA fragment cloned into pBluescript KS (pAB7). The probe for the DNA fragment, Frederick Cancer Research-2 (D18Fcr2), was a 320.bp DNA fragment amplified by PCR and purified in agarose (748 7), a gift from T. Wilkie (California Institute of Technology, Pasadena, CA). The probe for the deleted in colorectal carcinoma gene (Dee) was a human 1.65-kb EcoRI fragment cloned into pBluescript SK (pDCC 1.65; Fearon et al., 1990). The probe for the embigin-related sequence-3 (Et&rs3) was a mouse agarose-purified 250-bp PstI cDNA fragment (p17P; Ozawa et al., 1988), a gift from T. Muramatsu (Kagoshima University Faculty of Medicine, Kagoshima, Japan). The probe for the G protein olfactory subunit (Gnal) was a rat full-length cDNA cloned into pBluescript (rga; Jones and Reed, 1989) that was a gift from R. Reed (The Johns Hopkins University, Baltimore, MD). The probe for the gastrin-releasing peptide gene (Grp) was a human 650-bp BamHI/HindIII fragment cloned into pSP64 (pB12; DeteraWadleigh et al., 1987), a gift from E. R. Spindel (Oregon Regional Primate Center, Beaverton, OR). The probe for the hexokinase-l-related sequence 1 (Hk-Irsl) was a rat 3.6-kb EcoRI hexokinase cDNA fragment cloned into pUC19 (rHK-1; Schwab and Wilson, 1989), a gift from J. Wilson (Michigan State University, East Lansing, MI) and P. Overbeek (Baylor College of Medicine, Houston, TX). The probes for the heavy chain kinesin gene (Ens) were two agarose-purified fragments from the full-length human cDNA cloned into pSP72 (pXPE; Navone et al., 1992), a gift from R. Vale (University of California School of Medicine, San Francisco, CA). These fragments were (1) head, a 1.4-kb BamHI (linker)/SacI fragment; and (2) stalk, a 1.3-kb SacI/FspI fragment. A third probe for Ens representing the tail (an FspI/linker fragment) was also used to check for intragenic recombinants; however, no crossovers were detected among the three probes. The probe for the lamin B gene (Lmnb) was a mouse 1.36-kb EcoRI cDNA fragment cloned into pUC19 (aa77-533; Hoger et al., 1988), a gift from W. Reeves (University of North Carolina School of Medicine, Chapel Hill, NC) and N. Chaudhary (Rockefeller University, New York, NY). The probe for the mutated in colorectal cancer gene (Mcc) was a mouse l.l-kb EcoRI fragment cloned into pBluescript SK (MCC MB56-1) (L.-K. Su and K. W. Kinzler, unpublished data). The probe for prealbumin (Palb) was a human 0.3.kb EcoRI/HindIII cDNA fragment cloned into pSP64 (pHH64; Yoshoioka et al., 1986). a gift from Y. Sakaki (Kyushu University, Japan). The probe for the platelet-derived growth factor receptor-0 gene (Pdgjrb) was a mouse P.l-kb PstI cDNA fragment cloned into pGEM-2 (Yarden et al., 1986), a gift from U. Drager (Harvard Medical School, Boston, MA). The probe for the tumor progression locus-2 (Tpl-2) was a rat 2.5.kb PstI fragment cloned into pUC18 (pTP2).

ET AL,. The DZBLehl and D18Leh2 loci are interrepeat polymerase chain reaction products that have been described previously (Cox et al., 1991). The probes for the fibroblast growth factor-acidic (Fgfa), Friend murine leukemia virus integration site-2/macrophage colonystimulating factor receptor (Fim-Z/C’sfmr), glucocorticoid receptor (Grl-I), Ia antigen-associated invariant chain (Ii), myelin basic protein (Mbp), and mouse mammary tumor virus envelope (Mtu-38) genes have been described previously (Cox et al., 1991; Siracusa et al., 1991). DNA isolation and Southern blot analysis. High-molecular-weight genomic DNAs were prepared from frozen mouse tissues as described (Jenkins et al., 1982). Restriction endonuclease digestions, agarose gel electrophoresis, Southern transfers, and hybridizations were also performed as described (Jenkins et al., 1982), except that Zetabind membrane (CUNO, Inc.) was used for Southern transfers. Probes were labeled with [a-32P]dCTP (Amersham), using a nicktranslation kit (Boehringer Mannheim) for plasmid subclones and a random prime kit (Boehringer Mannheim) for gel-purified fragments. After hybridization with s*P-labeled probes, the filters were usually washed three times at 65°C under conditions that ranged from 0.1X to 1X SSCP, 0.1% SDS, depending on the probe. Filters hybridized with human probes were routinely washed three times at 65°C in 1X SSCP, 0.1% SDS. Probes were removed from the filters before rehybridization as described by Justice et al. (1990). Statistical analysis. Standard errors of the recombination frequencies from the results of the IB study were determined as described by Green (1981), using the computer program SPRETUS MADNESS. Gene order was determined by minimizing the number of multiple crossovers along the length of the chromosome (pedigree analysis; reviewed by Avner et al., 1988).

RESULTS

IB Analysis Twenty-three loci were mappedusing Southern analysis of DNAs from the N, progeny of a (C57BL/6J X M. spretus)F, X C57BL/6J backcross. C57BL/6J and M. spretus DNAs were digested with several restriction enzymes and analyzed by Southern hybridization with each of the probes listed in Table 1 and cited under Materials and Methods. At least one informative RFLP was identified for each probe. The segregation of the M. spretus allele(s) detected by each probe was followed in the N, progeny by Southern analysis. Only M. spretusspecific RFLPs were followed in backcross mice, since all backcrosses were to C57BL/6J, and C57BL/GJ-specific RFLPs could only be followed by hybridization intensity. The order of 16 of the 23 loci was determined by the analysis of 68 N, progeny (Fig. 1). Although only 68 animals were analyzed for all 16 loci, up to 193 animals were analyzed in pairwise combinations for some loci. The most likely gene order and the ratio of the total number of mice carrying recombinant chromosomes to the total number analyzed for each pair of loci are: centromereKns-7/164-Palb-14/164-Apc-8/l9l-Fgfa-2/189-Grl-l -4/179-Mcc-14/169-Lmnb-4/173-Pdgfrb-6/192D18Fcr2-5/182-Emb-rs3-2/147-Grp-2/154-Gnal-2/ 193-Mtu-38-3/176-Dcc-1/110-D18Fcr2-10/12. Six loci did not recombine with one of the 16 loci. NO crossovers were detected between (1) Kns and Tpl-2 (O/ 189, suggesting that the two loci are within 1.6 CM of

LINKAGE

MAP

OF

MOUSE

CHROMOSOME

TABLE Loci Mapped

by RFLP Analysis

1283

18

1

in Interspecific

Backcross

Mice

Restriction Locus

Gene

name

Probe

APC

Adenomatosis

Camk2a

Calmodulin-dependent protein kinase 11-a DNA fragment, Frederick Cancer Research-l DNA fragment, Frederick Cancer Research-Z Deleted in colorectal carcinoma

DIBFcrl D18Fcr2 DCC

polyposis

coli

Embigin-related

Gnal GYP Hk-lrsl

G protein, olfactory subunit Gastrin-releasing peptide Hexokinase-l-related sequence1 Kinesin heavy chain

Lmnb Mcc Palb Pdg frb Tpl-2

sequence-3

Lamin B Mutated in colorectal cancer Prealbumin Platelet-derived growth factor receptor-@ Tumor progression locus-2

73-1A

size in kb

C57BL/6J

Mus

spretus”

EcoRI HincII TaqI

13.0, 2.8 8.6, 0.9 4.4, 3.4, 0.9

g, 2.8 8.6, J.J 7.2, 3.4, 2.4, 1.4

pAB7

Hind111

748-7

XbaI

10.5, 6.4, 5.1, 4.6, 1.8, 0.8 10.0, 5.2

>23, 6.4 , 4.0 1.)3 6 1.8, 0.8 8.6, g >23, 15.0, 6.4, @, 2.4, 1.8, 1.4

pDCC

Emb-rs3

Kns

mAPC APC5’ pk2a3

Enzyme

fragment

1.65

PstI

p17P

EcoRI

>23, 17.0, 9.0, 6.8, 4.4, 3.1, 2.4, 1.8, 1.4 14.0, 6.2, 6.0, 3.5, 3.1

rga pB12 rHK-1

SphI SphI PUUII

1.8 9.3 5.3. 1.9

11.0, 8.4,b 8.2,’ 5.0,’ 3.5 4.9 8.8 Z,d34 -L

pXPE

Head:

77-533 MCC MB56-1 pHH64

Stalk: KpnI KpnI BamHI PVUII

16.0 18.0 7.5 8.8, 4.4 8.6, 8.2, 6.2, 1.2 r23, 7.6 7.5, 3.6, 2.7, 2.3

12.0 214 8.6 ii& 10.0, 11.0, 13.0,

pTP2

Hind111

3.4

4.6 -

Sac1 PUUII TaqI

n The RFLP that was mapped in the IB is underlined. When more than one RFLP are underlined, cosegregated. b This polymorphism detects an embigin-related sequence on Chr 6 (data not shown). ’ This polymorphism detects an embigin-related sequence on Chr 17 (data not shown). ’ This RFLP identifies the Hk-1 structural locus on mouse Chr 10 (data not shown). e This RFLP was used to map Kns in the C57BL6/Ros X M. spretus IBs.

each other, upper 95% confidence limit), (2) Ape and Dl8Leh2 (O/25, within 11.3 CM), (3) Pdgfrb and Ii (O/ 192, within 1.5 CM), Pdgfrb and Fim-2/Csfmr (O/175, within 1.7 CM), and (4) D18Fcr2 and Camk2a (O/94,

both

were

followed

6.2,

4.4 6.8, 6.2, a, 1.2 7.6 3.6, 2.7, 2.3, -1.2

in the IB, but

they

within 3.1 CM), and D18FcrZ and D18Leh2 (O/49, within 5.9 CM). Hk-lrsl was analyzed for a subset of the 68 N, animals, and the results placed Hk-lrsl between Palb and Apt (Fig. 1). The total number of animals analyzed

Ku.5 Palb Apt F& G&I

:8

ET”” 300 DfSFcrl MM

10 DO

18 12 98 FIG. 1. the left. Each parent. The bottom, the animals that

2

2

2 5

2

4

1 2

Segregation of alleles in 68 (C57BL/6J column represents the chromosome, black boxes represent the presence of first line indicates the total number of were analyzed for Hk-lrsl.

0

1

1 1 1

2

1

1 1

1

2 0 1

I

1

11 1

01

01

111014 1

1

3

X Mus spretus)F, X C57BL/6J IB progeny. Genes that were mapped in all 68 animals are at identified in the N, progeny, that was inherited from the (C57BL/6J x M. spretus)F, female a C57BL/6J allele, and the white boxes represent the presence of a M. spretus allele. At the offspring inheriting each type of chromosome, and the second line indicates the number of

1284

JUSTICE

Kns,

roprr

by the Poisson distribution (data not shown). This interference of multiple crossovers has been noted previously in both interspecific and intraspecific multilocus mouse crosses (Kingsley et al., 1989; Siracusa et al., 1989; Seldin et al., 1989; King et al., 1989).

T&Z

palb

18ql l-972

ET AL.

Hk-lrsl A&c,

5q21-q22

Map Distance to the Centromere

DlBLehl

1.1

4.2 5q31-q33 5q31

2.2

5q21-qz

8.3

Lmnb 5q33-q34,

5933-934,

5q32

2.3

&@+I,

Fim-2lCsfmr,

!a!

3.1

DIBFcr2, 18q21

rsq21

2.7 1.4 1.3 1.0 1.7

Camk2a,

g DlBLeh2

Emb-rs3 Gnal Mtv38 D i%FcrJ

0.9 1.9

lSq22-qler

&&

(shi)

t FIG. 2. A molecular genetic linkage map of mouse chromosome 18. The loci are aligned with the centromere at Km, and the map distances determined in our IB are to the left of the chromosome. Loci that have been mapped in humans are underlined. The human map locations are at the far left, and citations to the human locations can be found in the text.

for Palb, Hk-1 rsl, and Apt and the number of crossovers between them is Palb-2/32-Hk-lrsl-l/32-Ape. The recombination distances between each cluster of genes and flanking loci were determined using the locus from each cluster that was typed for the most N, animals. For the Palb/Hk-lrsl/Apc segment, the recombination distance from Palb to Apt is given, and Hk-1 r-s1 is positioned between the two loci. The recombination frequencies (expressed as genetic distance in centimorgans k standard error) between the loci are centromere-[Kns, Tpl-21-4.3 + 1.6-Palb-8.5 t 2.2-[Apt, D18Lehl]-4.2 -t 1.5-Fgfa-1.1 f 0.7-Grl-1-2.2 -t l.l-Mcc-8.3 + 2.1Lmnb-2.3 + l.l-[Pdgfrb, Fim-B/Csfmr, Ii]-3.1 * 1.3[D18Fcr2, CamkBa, D18Leh2]-2.7 + 1.2-Emb-rs3-1.4 & l.O-Grp-1.3 + 0.9-Gnal-1.0 rt 0.7-Mtu-38-1.7 -+ l.ODee-0.9 +- 0.9-D18Fcrl-7.9 + 2.4-Mbp. Although Dl8Leh1, D18Leh2, Fgfa, Fim-B/Csfmr, Grl-1, Ii, Mbp, and Mtv-38 have been reported previously by our laboratory (Cox et al., 1991; Siracusa et al., 1991), these studies give the order of these loci relative to 16 other loci mapped in a single cross (Fig. 2). No multiple crossover events were detected among the 68 N, animals; thus, fewer animals carrying multiple recombinants were observed than expected as calculated

The markers were oriented relative to the centromere by the analysis of a second IB between C57BL/6Ros and M. spretus (Y. Matsuda and V. Chapman, unpublished results) that was typed for the centromere by using a major mouse satellite DNA probe that preferentially detects C57BL/6Ros centromeres (Matsuda and Chapman, 1991; Brannan et al., 1992). No crossovers were detected between the centromere and Kns, the most proximal marker on Chr 18, in 142 N, animals. This suggests that Kns lies within 2.1 CM of the centromere (upper 95% confidence limit). Probes That Detect Multiple

Loci

A number of the probes detected more than one polymorphism that did not cosegregate in the mouse. First, the probe for hexokinase (rHK-1) detected RFLPs that identified the Hk-1 structural locus on mouse Chr 10 (data not shown), as well as the Hk-lrsl locus on Chr 18. Second, the probe for embigin (p17P) detected RFLPs that mapped to mouse chromosomes 18 (Emb-rs3), 17, and 6 (data not shown). Finally, the probe for the mouse mammary tumor virus envelope detects multiple loci in addition to Mtv-38 on Chr 18 (Siracusa et al., 1991). DISCUSSION

Using an IB between C57BL/6J and M. spretus mice, we have developed a molecular genetic linkage map of Chr 18 that includes 23 molecular markers (Fig. 2). From a second IB that was typed for the centromere, we were also able to position our mouse Chr 18 map with respect to the centromere. The Ape, Camk22a, Dee, D18Fcr1, D18Fcr2, Emb-rs3, Grp, Gnal, Hk-lrsl, Kns, Lmnb, Mcc, Palb, and Tpl-2 loci were mapped to mouse Chr 18 for the first time. The molecular markers mapped in this study span 50.9 CM, and thus extend along much of the estimated 59 CM of Chr 18 (Davisson and Roderick, 1989). Comparison

with Previous Mapping

Results

Few molecular markers have been previously placed on mouse Chr 18 (Davisson et al., 1991). Of the loci examined in our IB, the map positions of Fgfa, Fim-B/Csfmr, Grl-I, Ii, Mbp, and Pdgfrb have been reported previously in backcrosses performed by other laboratories. The map distances (in CM t SE), the most likely gene order, and the type of cross are Grl-l-9.8 + 2.8-Ii-17.0 + 3.6Mbp (intraspecific backcross, Byrd et al., 1991); Grl-l14.0 -t 3.3-Pdgfrb-3.5 * 1.7-Adrb-2-19.3 -+ 3.7-Mbp (interspecific backcross; Oakey et al., 1991); Grl-l-B.2

LINKAGE

MAP

OF

MOUSE

Insertion

lop

18q

Translocation

l@ 18q

with inversion

5q

18q

5q?

FIG. 3. Two possible mechanisms for the evolution of conserved linkage groups on mouse Chr 18 and human chromosomes 5q and 18q. The disruption of the human 18q linkage group by the human chromosome 5q linkage group could be explained by (A) a break with an accompanying insertion or (B) a translocation followed by an inversion.

f 3.0-Fim-2/Csfmr-42.4 -t 5.4-Mbp (interspecific backcross; Sakai et al., 1991). Our results are consistentwith the map distances and gene order reported in these crosses with the exception of the Csfmr to Mbp map distance reported by Sakai et al. (1991). Of the markers mapped in our cross, Fim-B/Csfmr and Mbp have been mapped by in situ hybridization to bands D and E3-4, respectively (Sola et al., 1988; Koizumi et al., 1991). These locations place Mbp on the distal end of Chr 18, and Fim-B/Csfmr proximal to Mbp, which is consistent with our map. Previously, the most proximal marker on Chr 18, Twirler (Tw), was mapped 9 CM from the centromere using two different centromere markers: (1) a duplication of centromeric heterochromatin and (2) a Robertsonian translocation (Lane et al., 1981). It is difficult to orient our map relative to a composite map of the chromosome (Davisson et al., 1991), since at this time, only Mbp (shi) has been mapped relative to both molecular markers and phenotypic markers. Linkage Homologies with Human

16

1285

kinase type IV [CAMK4 (Camk4)], monocyte differentiation antigen CD14 (Cd14), and cGMP phosphodiesterase (Y [PDEA (Pdea)], which have been assigned to

18q

B.

CHROMOSOME

Chromosomes

Many markers mapping to the central region of mouse Chr 18 are conserved on human chromosome 5q (Figs. 2 and 3). Of the loci mapped in our IB, those that have been localized previously on human chromosome 5q include Ape, Fgfa, Fim-B/Csfmr, Grl-1, Ii, Mcc, and Pdgfrb. APC (mouse homolog Ape) has been assigned to human 5q21-q22 (Ashton-Rickardt et al., 1989); MCC (Mcc) to 5q21-q22 (Kinzler et al., 1991b); GRL (G&I) to 5q31 (Huebner et al., 1990); FGFA (Fgfa) to 5q31-q33 (Jaye et al., 1986); DHLAG/HL (Ii) to 5q32 (Genuardi and Saunders, 1988); PDGFRB (Pdgfrb) to 5q33-q34 (Roberts et al., 1988); and CSFlR (Fim-2/Csfmr) to 5q33-q34 (Le Beau et al., 1986; Roberts et al., 1988). A number of other loci that have been previously assigned to Chr 18 also map to human chromosome 5q. These include adreneraic 8-2 receotor TADRB2R (A&b-2)1.~,~, calmodulin -- ~---- -.----L

5q31-q32,5q21-q23,5q23-q31, and 5, respectively (Kobilka et al., 1987; Sikela et al., 1989; Goyert et al., 1988; Pittler et al., 1990). On human chromosome 5q21-q22, MCC and APC are separated by less than 200 kb; however, in mouse they are interrupted by Fgfa and Grl-1, which map to human chromosomes 5q31-q33 and 5q31, respectively. The order of the 5q genes in human is centromere-MCCAPC-(FGFA, GRLl )-(PDGFRB, CSFl R), as determined by pulsed-field gel analysis of yeast artificial chromosomes (Kinzler et al., 1991b) and analysis of somatic cell hybrids with interstitial deletions (Huebner et al., 1990). Thus, although linkage is highly conserved between mouse Chr 18 and human chromosome 5q215q34, gene order is not conserved. Two other mouse chromosomes exhibit extensive linkage homology with human chromosome 5q. Mouse Chr 13 has extensive linkage homology with human chromosome 5qll-q14 (Justice and Stephenson, 1991), whereas mouse Chr 11 has extensive linkage homology with 5q31-q34 (Buchberg et al., 1991). Several of the loci mapping to mouse Chr 18 and Chr 11 define a cluster of growth factors, cytokines, and growth factor receptors on human chromosome 5q31-q34. Thus, a break in linkage homology occurred on the distal end of human chromosome 5q during the divergence of mouse and human lineages. Loci on both the proximal and distal ends of mouse Chr 18 are conserved on human chromosome 18q. PALB (Palb) maps to human chromosome 18qll-q12 (Sparkes et al., 1987). We have mapped another locus, N-cadherin [NCAD (N-cad)], between Kns and Palb on mouse Chr 18, which also maps to human chromosome 18 (Walsh et al., 1990; Miyatini et al., submitted for publication). DCC, GRP, and MBP have been assigned to human chromosomes 18q21, 18q21, and 18q22-qter, respectively (Fearon et al., 1990; Huebner et al., 1990; Kamholz et al., 1987). The relative order of these loci appears to be conserved between mouse and human, although they are disrupted by a region of human chromosome 5q linkage homology. One explanation for the disruption is a simple insertion of a region homologous to human chromosome 5q into a region homologous to human chromosome 18q (Fig. 3A). A second explanation for this disruption is that a translocation linking 5q and 18q markers was followed by a large inversion, creating the rearrangement shown in Fig. 3B. If this hypothesis is true, we would expect additional 5q markers to map distal to Mbp on mouse Chr 18. Further mapping studies will test this hypothesis. The assignment of Tpl-2 to the proximal end of mouse Chr 18 identifies a new homology segment. TPL2 maps to human chromosome lop11 (J. Testa and P. N. Tsichlis, unpublished results) and is the first human chromosome 10 gene to be mapped to mouse Chr 18.

1286

JUSTICE

Genes Encoding Known Mouse Mutations Among the eight phenotypic mutations that have been mapped to mouse Chr 18 (Green, 1989; Davisson et al., 1991; Byrd et al., 1991), the molecular basis of only one mutation, shiverer (shi), has been identified. The shi mutation represents a partial deletion of the myelin basic protein gene (Kimura et al., 1985; Katsuki et al., 1988). While none of the genes mapped in this study represent obvious candidate genes for other Chr 18 mutations, they will serve as useful anchors for additional physical mapping studies of Chr 18, which will aid in determining the gene products encoded by these mutant loci. Mouse Models of Human

Neoplasia

At least two loci on mouse Chr 18 represent candidates for mouse models of human hematologic disease. CSFlR is often deleted in the human 5q- myeloid syndrome (reviewed by Bunn, 1986), is a transforming protooncogene (v-fms), and is also activated by retroviral integration in mouse myeloid leukemias (Fim-2; Sola et al., 1988; Gisselbrecht et al., 1987). 1531-2 is a common site of retroviral integration in rat T-cell leukemia cell lines (C. Patriotis, A. Makris, S. E. Bear, and P. N. Tsichlis, unpublished results). The human homolog of TPL2 maps to chromosome lop11 within a region that could be involved in hereditary multiple endocrine neoplasia type II (Tokino et al., 1992). The same region is also frequently affected by chromosomal aberrations in human adult T-cell leukemia (ATL) (Shiraishi et al., 1985; Fujita et al., 1986) and acute nonlymphocytic leukemia (ANLL) (Bernstein et al., 1984). Three genes (APC, DCC, and MCC) that have been implicated in the progression of colorectal cancer (Vogelstein et al., 1988) map to mouse Chr 18. DCC is deleted in many colorectal carcinomas and may encode a tumor suppressor gene (Fearon et al., 1990). MCC is mutated in some colorectal carcinomas (Kinzler et al., 1991b), and mutations in APC predispose patients to adenomatous polyps that often develop into colon carcinoma (Kinzler et aL., 1991a; Groden et al., 1991). Mouse models of these loci would be very useful in examining the development and treatment of colon carcinoma. Recently, a dominant mouse mutation called multiple intestinal neoplasia (Min) was induced by ethylnitrosourea mutagenesis. Affected mice develop multiple adenomas throughout the intestinal tract and are predisposed to colorectal cancer (Moser et al., 1990). Mapping studies placed Min near Apt on Chr 18 (Luongo et al., submitted for publication), and subsequent studies showed that Min represents a mutant allele of the mouse homolog of Apt. Min. mice thus represent a valuable model for the study of colorectal cancer (Su et al., 1992). ACKNOWLEDGMENTS We thank H. Fujisawa, T. Wilkie, T. Muramatsu, del, .J. Wilson. P. Overbeek, R. Vale, W. Reeves,

R. Reed, E. SpinN. Chaudhary, Y.

ET AL. Sakaki, and U. Drager for kindly providing their resources. We thank Dr. J. Testa, Dr. W. F. Dove, Dr. A. K. Moser, and C. Luongo for providing information prior to its publication. D. S. Swing, B. C. Cho, and M. E. Barnstead are thanked for providing excellent technical assistance. We thank Dr. L. F. Lock for critically reviewing the manuscript. This research was supported, in part, by the National Cancer Institute, DHHS, under Contract NOl-CO-74101 with ABL. M. J. J. is the recipient of a National Research Service Award, Postdoctoral Fellowship 5F32CA088.53.03.

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