GENOMICS
8,57%582
(1990)
SHORT COMMUNICATION Genetic Mapping of the Gene for Ca2+/Calmodulin-Dependent Protein Kinase IV (Camk-4) to Mouse Chromosome 18 J. M. SIKELA,* *Department
M. C. ADAMSON,t
D. WILSON~HAW,*
AND C. A. KOZAKt
of Pharmacology, University of Colorado Health Sciences Center, Denver, Colorado 80262; Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892 Received
April
18, 1990;
revised
and tLaboratory
of
July 6, 1990
brain expression library using calmodulin as a probe (Sikela and Hahn, 1987). Additional cDNAs have subsequently been isolated and characterized from both mouse and rat and indicate that the gene is differentially processed to produce multiple and distinct mRNAs in brain and testis (Sikela et al., 1989; Ohmstede et al., 1989; Ono et al., 1989; Sikela, unpublished results). Also, the deduced amino acid sequences show that CaM kinase IV exhibits the same domain organization as other known CaM kinases and appears to be most closely related to brain CaM kinase II (Sikela et al., 1989; Schulman, 1988). In this paper, we report the chromosomal location of the gene for CaM kinase IV in the mouse. DNAs extracted from Chinese hamster E36 cells and NFS/N mouse liver were digested with various restriction enzymes, electrophoresed on 0.8% agarose gels, and hybridized with a probe which represents an 850-bp EcoRI fragment of X8, a mouse genomic clone containing part of the CaM kinase IV gene (Sikela et al., 1989). For all enzymes tested, digestion produced one or two major cross-reactive bands. PstI digestion produced an 8.0-kb band in hamster DNA and a 3.6kb band in mouse DNA (Fig. 1). DNAs from 14 Chinese hamster X mouse somatic cell hybrids (Hoggan et al., 1988) were analyzed by Southern blotting (Fig. 1). These hybrids were selected from a larger panel of 40 independently derived lines and 46 secondary clones. Eight of the 14 hybrids contained the mouse fragment, and comparison of the presence or absence of this 3.6-kb P&I band with the mouse chromosome content of the hybrids showed a positive correlation with mouse chromosome 18 (Table l), suggesting that this chromosome carries the gene for mouse CaM kinase IV, here termed Card-4. The single discordancy for chromosome 18 represents
Southern blot analysis of Chinese hamster X mouse somatic cell hybrids was used to map the gene for a serine/ threonine protein kinase expressed in brain and testis. This locus, termed Camk-4, encodes Caz+/calmodulin-dependent protein kinase IV. Progeny of an interspecific backcross were analyzed to position Camk-4 in the centromeric region of chromosome 18 near two mutations known to affect neurological function and fertility. This raises the possibility that a defect in Camk-4 may be responsible for one of these mutant phenotypes. o Iwo Academic press, IDC.
The protein kinases represent a large and diverse group of enzymes, many of which are known to mediate cellular responses to external stimuli (Hanks et al., 1988). These enzymes are subclassed by substrate specificity into serine/threonine-specific and tyrosine-specific kinases and include numerous cellular homologs of transforming proteins. Regulation of these enzymes varies, but one related group of serine/ threonine kinases is characterized by modulation by Ca2+/calmodulin (CaM kinases) (Stull et al., 1986; Kennedy et al., 1987). Recently, a new CaM kinase, i.e., CaM kinase IV, has been identified and appears to be found primarily, if not exclusively, in brain and testis (Sikela and Hahn, 1987; Sikela et al., 1989; Sikela, unpublished results). While its function is at present unknown, its expression in brain is postnatal and parallels a developmental period of active synaptogenesis (Ohmstede et al., 1989). That CaM kinase IV may be involved in synaptic development and/or activity is also suggested by its ability to phosphorylate synapsin, a brain phosphoprotein that has been implicated in mediating neurotransmitter release (Ohmstede et al., 1989). A partial-length mouse cDNA for CaM kinase IV was isolated by screening a 579
All
Copyright 0 1990 rights of reproduction
o&B-7543/90 $3.00 by Academic Press, Inc. in any form reserved.
580
SHORT
a
b
c
COMMUNICATION
de
-
8.0
-
3.6
FIG. 1. Southern blot analysis of P&I-digested mouse and Chinese hamster X mouse hybrid DNAs using the CaM kinase IV cDNA as probe. Lanes a-d, independent somatic cell hybrids; lane e, NFS/N mouse. Fragment sizes in kilobase pairs are indicated to the right.
a hybrid in which this chromosome is present at low frequency. To position this gene on chromosome 18, DNAs from progeny of an interspecies backcross were typed by Southern blot analysis for inheritance of restriction enzyme polymorphisms (RFLPs) of Camk-4 and other markers on chromosome 18. Mice of the laboratory strain parent used for this cross, NFS/N, were obtained from the Division of Natural Resources, NIH (Bethesda, MD). NFS/N females were mated with males of a laboratory colony of Mus mus muscuZU..Soriginally trapped in Skive, Denmark, and maintained by Dr. M. Potter (NC1 Contract NOl-CB71085 at Hazelton Laboratories, Rockville, MD); F, females were mated with musculus males to produce first backcross progeny. Southern blot analysis of NFS/N and musculus DNAs indicated that NFS/N DNA produces 6.9- and 2.9-kb BglII fragments cross-reactive with the CaM kinase IV cDNA, whereas M. m. musculus DNA produces a 9.4-kb fragment. DNAs from 45 of 74 backcross mice produced the NFS/N fragment (Fig. 2). The same DNAs were typed for RFLPs of Mbp (myelin basic protein) following digestion with ApaI and P&a (cGMP phosphodiesterase, LYsubunit) following digestion with EcoRI as described previously (Dantiger et al., in press). The data confirm that Cumk-4 maps to mouse chromosome 18 (Table 2) and further indicate that the most likely gene order is Cumk-4Pdea-Mbp. Since Mbp has been mapped near the telomeric end of chromosome 18 (Sidman et al., 1985), these data indicate that Camk-4 is the most centromerit of the three markers. The number of identified protein kinases in vertebrates now totals over 100 (T. Hunter, personal correspondence) and many have been cloned by virtue of
their sequence similarity to the catalytic domains of known protein kinases (Hanks et al., 1988). Genes for many members of the protein kinase family have been mapped in man and mouse, and the great majority of these mapped protein kinases represent those with known transforming potential (Kozak, 1990; Harper et al., 1989). Among the serine/threonine kinases, mapping data are available for members of the RafMos proto-oncogene subfamily and for Pim-1. Cloned probes for protein kinase C of the Ca’+/phospholipiddependent subfamily have been used to map PRKCA, PRKCB, and PRKCC in man (Coussens et aZ., 1986) and Pkca and Pkcc in mouse (Buchberg et al., 1989; Saunders and Seldin, 1990). Several members of the CaM kinase subfamily have now been identified and can be distinguished by one or more of a number of criteria (e.g., sequence, molecular weight, tissue distribution, substrate specificity). CaM kinases II and IV have been cloned along with myosin light chain kinase (both skeletal and smooth muscle forms), phosphorylase kinase (catalytic subunit), and putative serine kinase-HI (Hanks et al., 1988; Kennedy et aZ., TABLE
1
Analysis of Concordance between Specific Mouse Chromosomes and the Presence or Absence of the X Mouse SoCamk-4 Gene in 14 Chinese Hamster matic Cell Hybrids Number of hybrids: DNA hybridization/ chromosome” Mouse chromosome 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 X
+/+ 4 6 0 1 2 7 6 2 3 0 0 3 4 1 3 5 3 8 2 2
-/-
+/3 3 4 5 5 4 2 4 4 6 6 1 1 5 0 4 1 5 3 3
4 2 3 5 6 1 2 6 5 7 6 0 4 7 0 3 3 0 6 6
-/+ 3 3 2 1 0 2 4 1 2 0 0 3 5 1 3 1 5 1 2 3
% Discordancy 50 36 56 50 46 21 43 54 50 54 50 43 64 57 50 31 67 7 62 64
“Symbols indicate the presence (t-/) or absence (-/) of the mouse Camk-4 restriction fragment as related to the presence (/+) or absence (/-) of a particular mouse chromosome. Seven hybrids were karyotyped; the remainder were typed for specific marker loci including Fms and Mbp on chromosome 18.
SHORT
1987). CaM kinase IV is the only member of this subfamily for which information regarding chromosome map position in both mouse and man is known. Previous studies show that the human CAMK-4 locus maps to chromosome 5q (Sikela et al., 1989), and it has been shown that homologs of a number of genes on this human chromosome map to mouse chromosome 18 (Nadeau, 1989). Thus, our data extend information on this linkage homology and suggest that this segment of linkage homology is in the centromeric region of chromosome 18. The mapping of Gunk-4 provides a useful molecular marker for this region of chromosome 18 and should aid in positioning other genes predicted to map in this same region, such as Csfmr and Pdgfr. While no physiological role has been assigned to CaM kinase IV, its map location is of interest for several reasons. First, the human CaM kinase IV gene maps to 5q21-23 in a region that is rich in genes related to cell growth (Chandrasekharappa et al., 1990) and that has been implicated in various human disorders (Nakamura et al., 1988; LeBeau et al., 1989). Second, Camk-4 maps near two previously described mutations on mouse chromosome 18. Tw affects brain and ear development and may contribute to sterility. ax is a recessive mutation that causes paralysis and degeneration in various brain regions, as well as sterility (Green, 1989). Since CaM kinase IV is expressed uniquely in brain and testis, it is possible that mutations in this gene could produce the type of abnormal neurological or sexual development seen in mice carrying either ax or Tw. Thus, further studies on
a b
581
COMMUNICATION
TABLE
2
Segregation of the Camk-4 Hybridizing Restriction Fragment with Alleles of Mbp and Pdea in 74 Progeny of an IntersDecies Backcross Inheritance NFS/N Mice
Single
Pdea
Mb
Parentals recombinants
+ -
+ -
+ -
+ -
+ Double
recombinants
of the allele
+ -
+ -
+
Camk-4
Number of mice
+ + + + -
29 18 3 8 8 6 2 0
Recombination” Locus
pair
Mbp, Pdea Pdea, Camk-4 Mbp, Camk-4
r/n 16/74 13174 29/74
% Recombination k 1 SE 21.6 f 4.8 17.6 f 4.4 39.2 (NS)
a Percentage recombination between restriction fragments standard error were calculated according to Green (5) from number of recombinant5 (r.) in a sample size of n. NS, significant.
and the not
these kinases should describe the extent of diversity among members of this subgroup, shed some light on their normal physiological roles, and may assist in determining the molecular mechanisms that underlie a known mutation.
cd ACKNOWLEDGMENTS -
9.4
We thank Brenda Rae Marshall for preparing the manuscript and M. Peyser for technical assistance. This work was supported in part by NIH Grant NS27322.
6.9 REFERENCES
-
2.9
FIG. 2. Southern blot analysis of BglII-digested mouse liver DNAs using the CaM kinase IV cDNA as probe. Lanes a-c, liver DNAs from individual progeny of the interspecific backcross; lane d, NFS/N mouse DNA. Fragment sizes in kilobase pairs are indicated to the right.
BUCKBERG, 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. C~NDRASEKHARAPPA, S. C., REBELSKY, M. S., FIRAK, T. A., LJZBEAU, M. M., AND WESTBROOK, C. A. (1990). A long-range restriction map of the interleukin-4 and interleukin-5 linkage group on chromosome 5. Gerwmics 6: 94-99. COUSSENS, L., PARKER, P. J., RHEE, L., YANG-FENG, T. L., &EN, E., WATERFIELD, M. D., FRANCKE, U., AND ULWCH, A. (1986). Multiple, distinct forms of bovine and human protein kinase C suggest diversity in cellular signaling pathways. Science 233: 859-866. DANCIGER, M., KOZAK, C. A., LI, T., APPLEBURY, AND FARBER,
582
SHORT
COMMUNICATION
D. B. (1990). Genetic mapping demonstrates that the cumsuhunit of retinal cGMP-phosphodiesterase is not the site of the rd mutation. Genomics, in press. 5.
GREEN, Breeding
E. L. (1981). Experiments,”
6.
GREEN, M. C. (1989). phic loci. In “Genetic Mouse” (M. F. Lyon Press, Oxford.
“Genetics and Probability MacMillan, New York.
14.
NAKAMURA, Y., LATHROP, M., LEPPERT, M., DOBBS, M., WASMUTH, J., WOLFF, E., CARLSON, M., FUJIMOTO, E., KRAPCHO, K., SEARS, T., WOODWARD, S., HUGHES, J., BURT, R., GARDNER, E., LALOUEL, J.-M., AND WHITE, R. (1988). Localization of the genetic defect in familial adenomatous polyposis within a small region of chromosome 5. Amer. J. Hum. Genet. 43: 638-644.
15.
OHMSTEDE, C.-A., JENSEN, K. F., AND SAHYOUN, N. E. (1989). Caa+/calmodulin-dependent protein kinase enriched in cerebellar granule cells: Identification of a novel neuronal calmodulin-dependent protein kinase. J. Biol. Chem. 264: 58665875.
16.
ONO, T., SLAUGHTER, G. R., COOK, R. F., AND MEANS, A. R. (1989). Molecular cloning sequence and distribution of rat calspermin, a high affinity calmodulin-binding protein. J. Biol. Chem. 264: 2081-2087. SAUNDERS, A. M., AND SELDIN, M. F. (1990). Comparative mapping of genes localized to human chromosome 19 using an interspecific MLLS cross. Genomics 6: 324-332. SCHULMAN, H. (1988). The multifunctional Cae+/calmodulindependent protein kinase. Zn “Advances in Second Messenger and Phosphoprotein Research” (P. Greengard and G. A. Robinson, Eds.), Vol. 22, pp. 59-112, Raven Press, New York.
in Animal
Catalog of mutant genes and polymorVariants and Strains of the Laboratory and A. G. Searle, Eds.), Oxford Univ.
7.
HANKS, S. K., QUINN, protein kinase family: logeny of the catalytic
8.
HARPER, P. S., F&ZAL, J., FERGUSON-SMITH, SCHINZEL, A. (1989). Report of the Committee Disorders and Chromosomal Deletion Syndromes. Cell Genet. 51: 563-611.
9.
HOGGAN, M. D., HALDEN, N. F., BUCKLER, C. E., AND KODAK, C. A. (1988). Genetic mapping of the mouse c-fins proto-oncogene to chromosome 18. J. Virol. 62: 1055-1056.
18.
10.
KENNEDY, M. B., BEN=, M. K., ERONDU, N. E., AND MILLER, S. G. (1987). Calcium/calmodulin-dependent protein kinases. In “Calcium and Cell Function” (E. W.-Y. Cheung, Ed.), Vol. 7, pp. 62-106, Academic Press, New York.
19.
11.
12.
13.
A. M., AND HUNTER, T. (1988). Conserved features and deduced domains. Science 241: 42-52.
The phy-
M. A., AND on Clinical Cytogenet.
KOZAK, C. A. (1990). Retroviral and cancer related genes of the mouse. In “Genetic Maps, Locus Maps of Complex Genomes” (S. J. O’Brien, Ed.), pp. 4.69-4.79, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. LEBEAU, M. M., ALBAIN, K. S., LARSON, R. A., VARDIMAN, J. W., DAVIS, E. M., BLOUGH, R. R., GOLOMB, H. M., AND SHERR, C. J. (1986). Clinical and cytogenetic correlations in 63 patients with therapy-related myelodysplastic syndromes and acute nonlymphatic leukemia: Further evidence for characteristic abnormalities of chromosomes No. 5 and 7. J. Clin. Oncol. 4: 325-345. NADEAU, J. H. (1989). Maps of linkage and synteny homologies between mouse and man. Trends Genet. 5: 82-86.
17.
20.
21.
22.
SIDMAN, R. L., CONOVER, C. S., AND CARSON, J. H. (1985). Shiverer gene maps near the distal end of chromosome 18 in the mouse. Cytogenet. Cell Genet. 39: 241-245. SIKELA, J. M., AND HAHN, W. E. (1987). Screening an expression library with a ligand probe: Isolation and sequence of a cDNA corresponding to a brain calmodulin-binding protein. Proc. Natl. Acad. Sci. USA 84: 3038-3042. SIKELA, J. M., LAW, M. L., KAO, F. T., HARTZ, J., WEI, Q., AND HAHN, W. E. (1989). Chromosomal localization of the human gene for brain Ca2+/calmodulin-dependent protein kinase type IV. Genomics 4: 21-27. STULL, J. T., NUNNALLY, M. H., AND MICHNOFF, C. H. (1986). Calmodulin-dependent protein kinases. In “The Enzymes” (P. D. Boyer and E. G. Krebs, Eds.), Vol. 17, pp. 113-166, Academic Press, New York.
8,57%582
(1990)
SHORT COMMUNICATION Genetic Mapping of the Gene for Ca2+/Calmodulin-Dependent Protein Kinase IV (Camk-4) to Mouse Chromosome 18 J. M. SIKELA,* *Department
M. C. ADAMSON,t
D. WILSON~HAW,*
AND C. A. KOZAKt
of Pharmacology, University of Colorado Health Sciences Center, Denver, Colorado 80262; Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892 Received
April
18, 1990;
revised
and tLaboratory
of
July 6, 1990
brain expression library using calmodulin as a probe (Sikela and Hahn, 1987). Additional cDNAs have subsequently been isolated and characterized from both mouse and rat and indicate that the gene is differentially processed to produce multiple and distinct mRNAs in brain and testis (Sikela et al., 1989; Ohmstede et al., 1989; Ono et al., 1989; Sikela, unpublished results). Also, the deduced amino acid sequences show that CaM kinase IV exhibits the same domain organization as other known CaM kinases and appears to be most closely related to brain CaM kinase II (Sikela et al., 1989; Schulman, 1988). In this paper, we report the chromosomal location of the gene for CaM kinase IV in the mouse. DNAs extracted from Chinese hamster E36 cells and NFS/N mouse liver were digested with various restriction enzymes, electrophoresed on 0.8% agarose gels, and hybridized with a probe which represents an 850-bp EcoRI fragment of X8, a mouse genomic clone containing part of the CaM kinase IV gene (Sikela et al., 1989). For all enzymes tested, digestion produced one or two major cross-reactive bands. PstI digestion produced an 8.0-kb band in hamster DNA and a 3.6kb band in mouse DNA (Fig. 1). DNAs from 14 Chinese hamster X mouse somatic cell hybrids (Hoggan et al., 1988) were analyzed by Southern blotting (Fig. 1). These hybrids were selected from a larger panel of 40 independently derived lines and 46 secondary clones. Eight of the 14 hybrids contained the mouse fragment, and comparison of the presence or absence of this 3.6-kb P&I band with the mouse chromosome content of the hybrids showed a positive correlation with mouse chromosome 18 (Table l), suggesting that this chromosome carries the gene for mouse CaM kinase IV, here termed Card-4. The single discordancy for chromosome 18 represents
Southern blot analysis of Chinese hamster X mouse somatic cell hybrids was used to map the gene for a serine/ threonine protein kinase expressed in brain and testis. This locus, termed Camk-4, encodes Caz+/calmodulin-dependent protein kinase IV. Progeny of an interspecific backcross were analyzed to position Camk-4 in the centromeric region of chromosome 18 near two mutations known to affect neurological function and fertility. This raises the possibility that a defect in Camk-4 may be responsible for one of these mutant phenotypes. o Iwo Academic press, IDC.
The protein kinases represent a large and diverse group of enzymes, many of which are known to mediate cellular responses to external stimuli (Hanks et al., 1988). These enzymes are subclassed by substrate specificity into serine/threonine-specific and tyrosine-specific kinases and include numerous cellular homologs of transforming proteins. Regulation of these enzymes varies, but one related group of serine/ threonine kinases is characterized by modulation by Ca2+/calmodulin (CaM kinases) (Stull et al., 1986; Kennedy et al., 1987). Recently, a new CaM kinase, i.e., CaM kinase IV, has been identified and appears to be found primarily, if not exclusively, in brain and testis (Sikela and Hahn, 1987; Sikela et al., 1989; Sikela, unpublished results). While its function is at present unknown, its expression in brain is postnatal and parallels a developmental period of active synaptogenesis (Ohmstede et al., 1989). That CaM kinase IV may be involved in synaptic development and/or activity is also suggested by its ability to phosphorylate synapsin, a brain phosphoprotein that has been implicated in mediating neurotransmitter release (Ohmstede et al., 1989). A partial-length mouse cDNA for CaM kinase IV was isolated by screening a 579
All
Copyright 0 1990 rights of reproduction
o&B-7543/90 $3.00 by Academic Press, Inc. in any form reserved.
580
SHORT
a
b
c
COMMUNICATION
de
-
8.0
-
3.6
FIG. 1. Southern blot analysis of P&I-digested mouse and Chinese hamster X mouse hybrid DNAs using the CaM kinase IV cDNA as probe. Lanes a-d, independent somatic cell hybrids; lane e, NFS/N mouse. Fragment sizes in kilobase pairs are indicated to the right.
a hybrid in which this chromosome is present at low frequency. To position this gene on chromosome 18, DNAs from progeny of an interspecies backcross were typed by Southern blot analysis for inheritance of restriction enzyme polymorphisms (RFLPs) of Camk-4 and other markers on chromosome 18. Mice of the laboratory strain parent used for this cross, NFS/N, were obtained from the Division of Natural Resources, NIH (Bethesda, MD). NFS/N females were mated with males of a laboratory colony of Mus mus muscuZU..Soriginally trapped in Skive, Denmark, and maintained by Dr. M. Potter (NC1 Contract NOl-CB71085 at Hazelton Laboratories, Rockville, MD); F, females were mated with musculus males to produce first backcross progeny. Southern blot analysis of NFS/N and musculus DNAs indicated that NFS/N DNA produces 6.9- and 2.9-kb BglII fragments cross-reactive with the CaM kinase IV cDNA, whereas M. m. musculus DNA produces a 9.4-kb fragment. DNAs from 45 of 74 backcross mice produced the NFS/N fragment (Fig. 2). The same DNAs were typed for RFLPs of Mbp (myelin basic protein) following digestion with ApaI and P&a (cGMP phosphodiesterase, LYsubunit) following digestion with EcoRI as described previously (Dantiger et al., in press). The data confirm that Cumk-4 maps to mouse chromosome 18 (Table 2) and further indicate that the most likely gene order is Cumk-4Pdea-Mbp. Since Mbp has been mapped near the telomeric end of chromosome 18 (Sidman et al., 1985), these data indicate that Camk-4 is the most centromerit of the three markers. The number of identified protein kinases in vertebrates now totals over 100 (T. Hunter, personal correspondence) and many have been cloned by virtue of
their sequence similarity to the catalytic domains of known protein kinases (Hanks et al., 1988). Genes for many members of the protein kinase family have been mapped in man and mouse, and the great majority of these mapped protein kinases represent those with known transforming potential (Kozak, 1990; Harper et al., 1989). Among the serine/threonine kinases, mapping data are available for members of the RafMos proto-oncogene subfamily and for Pim-1. Cloned probes for protein kinase C of the Ca’+/phospholipiddependent subfamily have been used to map PRKCA, PRKCB, and PRKCC in man (Coussens et aZ., 1986) and Pkca and Pkcc in mouse (Buchberg et al., 1989; Saunders and Seldin, 1990). Several members of the CaM kinase subfamily have now been identified and can be distinguished by one or more of a number of criteria (e.g., sequence, molecular weight, tissue distribution, substrate specificity). CaM kinases II and IV have been cloned along with myosin light chain kinase (both skeletal and smooth muscle forms), phosphorylase kinase (catalytic subunit), and putative serine kinase-HI (Hanks et al., 1988; Kennedy et aZ., TABLE
1
Analysis of Concordance between Specific Mouse Chromosomes and the Presence or Absence of the X Mouse SoCamk-4 Gene in 14 Chinese Hamster matic Cell Hybrids Number of hybrids: DNA hybridization/ chromosome” Mouse chromosome 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 X
+/+ 4 6 0 1 2 7 6 2 3 0 0 3 4 1 3 5 3 8 2 2
-/-
+/3 3 4 5 5 4 2 4 4 6 6 1 1 5 0 4 1 5 3 3
4 2 3 5 6 1 2 6 5 7 6 0 4 7 0 3 3 0 6 6
-/+ 3 3 2 1 0 2 4 1 2 0 0 3 5 1 3 1 5 1 2 3
% Discordancy 50 36 56 50 46 21 43 54 50 54 50 43 64 57 50 31 67 7 62 64
“Symbols indicate the presence (t-/) or absence (-/) of the mouse Camk-4 restriction fragment as related to the presence (/+) or absence (/-) of a particular mouse chromosome. Seven hybrids were karyotyped; the remainder were typed for specific marker loci including Fms and Mbp on chromosome 18.
SHORT
1987). CaM kinase IV is the only member of this subfamily for which information regarding chromosome map position in both mouse and man is known. Previous studies show that the human CAMK-4 locus maps to chromosome 5q (Sikela et al., 1989), and it has been shown that homologs of a number of genes on this human chromosome map to mouse chromosome 18 (Nadeau, 1989). Thus, our data extend information on this linkage homology and suggest that this segment of linkage homology is in the centromeric region of chromosome 18. The mapping of Gunk-4 provides a useful molecular marker for this region of chromosome 18 and should aid in positioning other genes predicted to map in this same region, such as Csfmr and Pdgfr. While no physiological role has been assigned to CaM kinase IV, its map location is of interest for several reasons. First, the human CaM kinase IV gene maps to 5q21-23 in a region that is rich in genes related to cell growth (Chandrasekharappa et al., 1990) and that has been implicated in various human disorders (Nakamura et al., 1988; LeBeau et al., 1989). Second, Camk-4 maps near two previously described mutations on mouse chromosome 18. Tw affects brain and ear development and may contribute to sterility. ax is a recessive mutation that causes paralysis and degeneration in various brain regions, as well as sterility (Green, 1989). Since CaM kinase IV is expressed uniquely in brain and testis, it is possible that mutations in this gene could produce the type of abnormal neurological or sexual development seen in mice carrying either ax or Tw. Thus, further studies on
a b
581
COMMUNICATION
TABLE
2
Segregation of the Camk-4 Hybridizing Restriction Fragment with Alleles of Mbp and Pdea in 74 Progeny of an IntersDecies Backcross Inheritance NFS/N Mice
Single
Pdea
Mb
Parentals recombinants
+ -
+ -
+ -
+ -
+ Double
recombinants
of the allele
+ -
+ -
+
Camk-4
Number of mice
+ + + + -
29 18 3 8 8 6 2 0
Recombination” Locus
pair
Mbp, Pdea Pdea, Camk-4 Mbp, Camk-4
r/n 16/74 13174 29/74
% Recombination k 1 SE 21.6 f 4.8 17.6 f 4.4 39.2 (NS)
a Percentage recombination between restriction fragments standard error were calculated according to Green (5) from number of recombinant5 (r.) in a sample size of n. NS, significant.
and the not
these kinases should describe the extent of diversity among members of this subgroup, shed some light on their normal physiological roles, and may assist in determining the molecular mechanisms that underlie a known mutation.
cd ACKNOWLEDGMENTS -
9.4
We thank Brenda Rae Marshall for preparing the manuscript and M. Peyser for technical assistance. This work was supported in part by NIH Grant NS27322.
6.9 REFERENCES
-
2.9
FIG. 2. Southern blot analysis of BglII-digested mouse liver DNAs using the CaM kinase IV cDNA as probe. Lanes a-c, liver DNAs from individual progeny of the interspecific backcross; lane d, NFS/N mouse DNA. Fragment sizes in kilobase pairs are indicated to the right.
BUCKBERG, 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. C~NDRASEKHARAPPA, S. C., REBELSKY, M. S., FIRAK, T. A., LJZBEAU, M. M., AND WESTBROOK, C. A. (1990). A long-range restriction map of the interleukin-4 and interleukin-5 linkage group on chromosome 5. Gerwmics 6: 94-99. COUSSENS, L., PARKER, P. J., RHEE, L., YANG-FENG, T. L., &EN, E., WATERFIELD, M. D., FRANCKE, U., AND ULWCH, A. (1986). Multiple, distinct forms of bovine and human protein kinase C suggest diversity in cellular signaling pathways. Science 233: 859-866. DANCIGER, M., KOZAK, C. A., LI, T., APPLEBURY, AND FARBER,
582
SHORT
COMMUNICATION
D. B. (1990). Genetic mapping demonstrates that the cumsuhunit of retinal cGMP-phosphodiesterase is not the site of the rd mutation. Genomics, in press. 5.
GREEN, Breeding
E. L. (1981). Experiments,”
6.
GREEN, M. C. (1989). phic loci. In “Genetic Mouse” (M. F. Lyon Press, Oxford.
“Genetics and Probability MacMillan, New York.
14.
NAKAMURA, Y., LATHROP, M., LEPPERT, M., DOBBS, M., WASMUTH, J., WOLFF, E., CARLSON, M., FUJIMOTO, E., KRAPCHO, K., SEARS, T., WOODWARD, S., HUGHES, J., BURT, R., GARDNER, E., LALOUEL, J.-M., AND WHITE, R. (1988). Localization of the genetic defect in familial adenomatous polyposis within a small region of chromosome 5. Amer. J. Hum. Genet. 43: 638-644.
15.
OHMSTEDE, C.-A., JENSEN, K. F., AND SAHYOUN, N. E. (1989). Caa+/calmodulin-dependent protein kinase enriched in cerebellar granule cells: Identification of a novel neuronal calmodulin-dependent protein kinase. J. Biol. Chem. 264: 58665875.
16.
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