Silver fox gene mapping: conserved chromosome regions in the order Carnivora

Twenty-three silver fox x hamster somatic cell hybrid clones were used to assign 15 fox genes: GPI to chromosome 1; PGD to chromosome 2; MDH2 to chromosome 3; ESD to chromosome 6; LDHB to chromosome 8; NP to chromosome 10; LDHA to chromosome 11; APRT, ENO1, and PGM1 to chromosome 12; IDH1 and MDH1 to chromosome 16; and GLA, G6PD, and HPRT to the X chromosome. High-resolution G-banding of human, cat, mink, and fox chromosomes containing homologous regions (according to genetic maps) revealed regions of putative homology. The results lend support to the suggestion that the most considerable karyotypic reorganization of the ancestral genome in the order Carnivora occurred during Canidae formation. The details of karyotypic evolution in mammals are discussed.     

A complete comparative chromosome map for the dog, red fox, and human and its integration with canine genetic maps

Cross-species reciprocal chromosome painting was used to delineate homologous chromosomal segments between domestic dog, red fox, and human. Whole sets of chromosome-specific painting probes for the red fox and dog were made by PCR amplification of flow-sorted chromosomes from established cell cultures. Based on their hybridization patterns, a complete comparative chromosome map of the three species has been built. Thirty-nine of the 44 synteny groups from the published radiation hybrid map and 33 of the 40 linkage groups in the linkage map of the dog have been assigned to specific chromosomes by fluorescence in situ hybridization and PCR-based genotyping. Each canine chromosome has at least one DNA marker assigned to it. The human-canid map shows that the canid karyotypes are among the most extensively rearranged karyotypes in mammals. Twenty-two human autosomal paints delineated 73 homologous regions on 38 canine autosomes, while paints from 38 dog autosomes detected 90 homologous segments in the human genome. Of the 22 human autosomes, only the syntenies of three chromosomes (14, 20, and 21) have been maintained intact in the canid genome. The dog-fox map and DAPI banding comparison demonstrate that the remarkable karyotype differences between fox (2n = 34 + 0-8 Bs) and dog (2n = 78) are due to 26 chromosomal fusion events and 4 fission events. It is proposed that the more easily karyotyped fox chromosomes can be used as a common reference and control system for future gene mapping in the DogMap project and CGH analysis of canine tumor DNA.     

Genome mapping in silver fox. Syntenic genes in Carnivora

Hamster X fox somatic cell hybrids segregating individual fox chromosomes in different combinations were used to assign seven structural loci to fox chromosomes. The gene for ME1 was mapped on the VFU1 chromosome, the genes for ADK and PP being located on the VFU4 chromosome. The gene for GSR was assigned to the VFU7 chromosome and the genes for MPI and COT1 were assigned to the VFU15 chromosome. Localization of these genes enhances the established fox genetic map and extends the known syntenic homologies between the fox and other mammalian. The comparison of data on gene mapping has provided basis for suggestion that there are significant differences in rates of karyotypic evolution in many mammalian taxa.     

Zoo-FISH with region-specific paints for mink chromosome 5q: delineation of inter- and intrachromosomal rearrangements in human, pig, and fox

Comparison of evolutionarily conserved mammalian chromosomes homologous to human chromosome 17, performed with microdissected painting probes, revealed rearrangements inside these chromosomes in mink and pig and a disruption of this conserved region in the fox. Detection of a homologous region on an Iberian shrew chromosome showed the efficiency of microdissected painting probes for delineation of homologous chromosome regions in species belonging to orders that diverged at least 100 million years ago.     

Association of MC3R gene polymorphisms with body weight in the red fox and comparative gene organization in four canids

There are five genes encoding melanocortin receptors. Among canids, the genes have mainly been studied in the dog (MC1R, MC2R and MC4R). The MC4R gene has also been analysed in the red fox. In this report, we present a study of chromosome localization, comparative sequence analysis and polymorphism of the MC3R gene in the dog, red fox, arctic fox and Chinese raccoon dog. The gene was localized by FISH to the following chromosome: 24q24‐25 in the dog, 14p16 in the red fox, 18q13 in the arctic fox and NPP4p15 in the Chinese raccoon dog. A high identity level of the MC3R gene sequences was observed among the species, ranging from 96.0% (red fox – Chinese raccoon dog) to 99.5% (red fox – arctic fox). Altogether, eight polymorphic sites were found in the red fox, six in the Chinese raccoon dog and two in the dog, while the arctic fox appeared to be monomorphic. In addition, association of several polymorphisms with body weight was analysed in red foxes (the number of genotyped animals ranged from 319 to 379). Two polymorphisms in the red fox, i.e. a silent substitution c.957A>C and c.*185C>T in the 3′‐flanking sequence, showed a significant association (P < 0.01) with body weight.     

Creation of a clone panel of fox x Chinese hamster somatic cell hybrids and chromosome mapping of genes for LDHA, LDHB, GPI, ESD, G6PD, HPRT, alpha-GALA in the silver fox

A clone panel of fox-hamster somatic cell hybrids which can be used for fox gene mapping was set up. Analysis of patterns of chromosome-enzyme segregation made it possible to assign gene GPI to chromosome 1, LDHA to chromosome 11, LDHB to chromosome 8, ESD to chromosome 6 and G6PD, HPRT, alpha-GALA to chromosome X.     

Phylogenetic implications of the 38 putative ancestral chromosome segments for four canid species.

Chromosome homologies between the Japanese raccoon dog (Nectereutes procyonoides viverrinus, 2n = 39 + 2-4 B chromosomes) and domestic dog (Canis familiaris, 2n = 78) have been established by hybridizing a complete set of canine paint probes onto high-resolution G-banded chromosomes of the raccoon dog. Dog chromosomes 1, 13, and 19 each correspond to two raccoon dog chromosome segments, while the remaining 35 dog autosomes each correspond to a single segment. In total, 38 dog autosome paints revealed 41 conserved segments in the raccoon dog. The use of dog painting probes has enabled integration of the raccoon dog chromosomes into the previously established comparative map for the domestic dog, Arctic fox (Alopex lagopus), and red fox (Vulpes vulpes). Extensive chromosome arm homologies were found among chromosomes of the red fox, Arctic fox, and raccoon dog. Contradicting previous findings, our results show that the raccoon dog does not share a single biarmed autosome in common with the Arctic fox, red fox, or domestic cat. Comparative analysis of the distribution patterns of conserved chromosome segments revealed by dog paints in the genomes of the canids, cats, and human reveals 38 ancestral autosome segments. These segments could represent the ancestral chromosome arms in the karyotype of the most recent ancestor of the Canidae family, which we suggest could have had a low diploid number, based on comparisons with outgroup species.     

Chromosomal evolution of the Canidae. II. Divergence from the primitive carnivore karyotype

The Giemsa-banding patterns of chromosomes from the arctic fox (Alopex lagopus), the red fox (Vulpes vulpes), the kit fox (Vulpes macrotis), and the raccoon dog (Nyctereutes procyonoides) are compared. Despite their traditional placement in different genera, the arctic fox and the kit fox have an identical chromosome morphology and G-banding pattern. The red fox has extensive chromosome arm homoeology with these two species, but has only two entire chromosomes in common. All three species share some chromosomes with the raccoon dog, as does the high diploid-numbered grey wolf (Canis lupus, 2n = 78). Moreover, some chromosomes of the raccoon dog show partial or complete homoeology with metacentric feline chromosomes which suggests that these are primitive canid chromosomes. We present the history of chromosomal rearrangements within the Canidae family based on the assumption that a metacentric-dominated karyotype is primitive for the group.     

Mapping of the silver fox genome. III. Determination of the chromosomal localization of the GOT2, AK1, ALDOC, ACP1, ITPA, PGP and BLVR genes

Evidence is presented for the chromosome localization of seven silver fox genes obtained with the help of panel of fox x Chinese hamster somatic cell hybrids. Thus, the AK1, GOT2 and ALDOC are assigned to chromosome VFU2, PGP to chromosome VFU3, BLVR to chromosome VFU5, ACP1 to chromosome VFU8 and ITPA to chromosome VFU14. The genetic map of 29 fox genes is compared with those reported for man and other animals. The results obtained support and extend our previous suggestion that formation of the Canidae branch of the Carnivora phylogenetic tree was associated with great increase in the rate of reorganization of the ancestral karyotype.     

Localization of human gene loci using spontaneous chromosome rearrangements in human-Chinese hamster somatic cell hybrids.

Analysis of human-Chinese hamster somatic cell hybrids with spontaneously derived chromosome structural changes has provided data for the regional and subregional localization of gene loci which have previously been assigned to human chromosomes 2, 12, and X. Correlation of the expression of human gene loci with the human chromosome complements present in somatic cell hybrids indicates that the cytoplasmic malate dehydrogenase (MDH1) locus is in the 2p23yields2pter region, and red cell acid phosphatase (AcP1) is at or adjacent to 2p23. The cytoplasmic isocitrate dehydrogenase (IDH1) locus is at or adjacent to 2q11, peptidase B (Pep B) is at or adjacent to 12q21, lactate dehydrogenase B (LDH B) is in the 12q21yiedls12pter region, glucose-6-phosphate dehydrogenase (G6PD) is in the Xq24yieldsXqter region, and the gene loci for phosphoglycerate kinase (PGK), alpha-galactosidase (alpha-gal), and hypoxanthine guanine phosphoribosyltransferase (GPRT) are in the Xp21yieldsXq24 region.     

Mapping of silver fox genes: chromosomal localization of the genes for GOT2, AK1, ALDOC, ACP1, ITPA, PGP, and BLVR.

Evidence is presented for the chromosome localization of seven silver fox genes by the use of a panel of fox x Chinese hamster somatic cell hybrids. AK1, GOT2, and ALDOC are assigned to chromosome VFU2, PGP to chromosome VFU38, BLVR to chromosome VFU5, ACP1 to chromosome VFU8, and ITPA to chromosome VFU14. The genetic map of 29 fox genes is compared with those reported for man and other mammals. The results we obtained support and extend our previous suggestion that the formation of the Canidae branch of the Carnivora phylogenic tree was associated with a great increase in the rate of reorganization of the ancestral karyotype.     

Molecular genetic linkage maps of mouse chromosomes 4 and 6.

We have generated a moderate resolution genetic map of mouse chromosomes 4 and 6 utilizing a (C57BL/6J x Mus spretus) F1 x Mus spretus backcross with RFLPs for 31 probes. The map for chromosome 4 covers 77 cM and details a large region of homology to human chromosome 1p. The map establishes the breakpoints in the mouse 4-human 1p region of homology to a 2-cM interval between Ifa and Jun in mouse and to the interval between JUN and ACADM in human. The map for mouse chromosome 6 spans a 65-cM region and contains a large region of homology to human 7q. These maps also provide chromosomal assignment and order for a number of previously unmapped probes. The maps should allow the rapid regional assignment of new markers to mouse chromosomes 4 and 6. In addition, knowledge of the gene order in mouse may prove useful in determining the gene order of the homologous regions in human.     

Identification of genes controlling collagen-induced arthritis in mice: striking homology with susceptibility loci previously identified in the rat.

The susceptibility to collagen-induced arthritis in the highly susceptible DBA/1 mouse has earlier been shown to be partly controlled by the MHC class II gene Aq. To identify susceptibility loci outside of MHC, we have made crosses between DBA/1 and the less susceptible B10.Q strain, both expressing the MHC class II gene Aq. Analysis of 224 F2 intercross mice with 170 microsatellite markers in a genome-wide scan suggested 4 quantitative trait loci controlling arthritis susceptibility located on chromosomes 6, 7, 8, and 10. The locus on chromosome 6 (Cia6), which was associated with arthritis onset, yielded a logarithm of odds score of 4.7 in the F2 intercross experiment and was reproduced in serial backcross experiments. Surprisingly, the DBA/1 allele had a recessive effect leading to a delay in arthritis onset. The suggestive loci on chromosomes 7 and 10 were associated with arthritis severity rather than onset, and another suggestive locus on chromosome 8 was most closely associated with arthritis incidence. The loci on chromosomes 7, 8, and 10 all appeared to contain disease-promoting alleles derived from the DBA/1 strain. Interestingly, most of the identified loci were situated in chromosomal regions that are homologous to regions in the rat genome containing susceptibility genes for arthritis; the mouse Cia6 locus is homologous with the rat Cia3, Pia5, Pia2, and Aia3; the locus on chromosome 7 (Cia7) is homologous with the rat Cia2; and the locus on chromosome 10 (Cia8) is homologous with the rat Cia4.     

Genomic homology of the domestic ferret with cats and humans

We used chromosome paints from both the domestic cat and humans to directly establish chromosomal homology between the genome of these species and the domestic ferret. The chromosome painting data indicate that the ferret has a highly conserved karyotype closer to the ancestral carnivore karyotype than that of the cat. The cat chromosome paints revealed 22 homologous autosomal regions in the ferret genome: 16 ferret chromosomes were hybridized by a single cat paint, while 3 ferret chromosomes were hybridized by two cat paints. In situ hybridization combined with banding showed that ferret Chromosome (Chr) 1 = cat A2p/C2, Chr 2 = F2/C1q, and Chr 3 = A2q/D2. Five ferret chromosomes are homologous to single arms of cat chromosomes: ferret 4 = A1q, 5 = B1q, 6 = C1p, 10 = A1p, and 12 = B1p. The human chromosome paints revealed 32 + XY homologous regions in the ferret genome: 9 ferret chromosomes were each hybridized by a single human paint, 7 by two paints, 3 by three paints. The 10 ferret chromosomes hybridized by multiple human paints produced the following associations: ferret 1 = human 19/3/21, 2 = 8q/2q, 3 = 10/7, 5 = 8/4, 8 = 15/14, 9 = 10/12/22, 11 = 20/2, 12 = 8/4, 14 = 12/22/18, 18 = 19/16. We present an index of genomic diversity, Z, based on the relative number of conserved whole chromosome and chromosome segments as a preliminary statistic for rapid comparison between species. The index of diversity between human-ferret (Z = 0.812) is slightly less than human-cat (Z = 0.843). The homology data presented here allow us to transfer gene mapping data from both cats and humans to the ferret. Received: 21 December 1999 / Accepted: 30 May 2000     

Polymorphism and chromosomal location of the MC4R (melanocortin-4 receptor) gene in the dog and red fox

The melanocortin-4 receptor (MC4R) is expressed in the hypothalamus and regulates energy intake and body weight. In silico screening of the canine chromosome 1 sequence and a comparison with the porcine MC4R sequence by BLAST were performed. The nucleotide sequence of the whole coding region and 3'- and 5'-flanking regions of the dog (1214 bp) and red fox (1177 bp) MC4R gene was established and high conservation of the nucleotide sequences was revealed (99%). Five sets of PCR primers were designed and a search for polymorphism was performed by the SSCP technique in a group of 31 dogs representing nineteen breeds and 35 farm red foxes. Sequencing of DNA fragments, representing the identified SSCP patterns, revealed three single nucleotide polymorphisms (including a missense one) in dogs and four silent SNPs in red foxes. An average SNP frequency was approx. 1/400 bp in the dog and 1/300 bp in the red fox. We mapped the MC4R gene by FISH to the canine chromosome 1 (CFA1q1.1) and to the red fox chromosome 5 (VVU5p1.2).     

A comparative analysis of MC4R gene sequence, polymorphism, and chromosomal localization in Chinese raccoon dog and Arctic fox

Numerous mutations of the human melanocortin receptor type 4 (MC4R) gene are responsible for monogenic obesity, and some of them appear to be associated with predisposition or resistance to polygenic obesity. Thus, this gene is considered a functional candidate for fat tissue accumulation and body weight in domestic mammals. The aim of the study was comparative analysis of chromosome localization, nucleotide sequence, and polymorphism of the MC4R gene in two farmed species of the Canidae family, namely the Chinese raccoon dog (Nycterutes procyonoides procyonoides) and the arctic fox (Alopex lagopus). The whole coding sequence, including fragments of 3'UTR and 5'UTR, shows 89% similarity between the arctic fox (1276 bp) and Chinese raccoon dog (1213 bp). Altogether, 30 farmed Chinese raccoon dogs and 30 farmed arctic foxes were searched for polymorphisms. In the Chinese raccoon dog, only one silent substitution in the coding sequence was identified; whereas in the arctic fox, four InDels and two single-nucleotide polymorphisms (SNPs) in the 5'UTR and six silent SNPs in the exon were found. The studied gene was mapped by FISH to the Chinese raccoon dog chromosome 9 (NPP9q1.2) and arctic fox chromosome 24 (ALA24q1.2-1.3). The obtained results are discussed in terms of genome evolution of species belonging to the family Canidae and their potential use in animal breeding.     

Sequences homologous to glutamic acid decarboxylase cDNA are present on mouse chromosomes 2 and 10

The chromosomal locations of mouse DNA sequences homologous to a feline cDNA clone encoding glutamic acid decarboxylase (GAD) were determined. Although cats and humans are thought to have only one gene for GAD, GAD cDNA sequences hybridize to two distinct chromosomal loci in the mouse, chromosomes 2 and 10. The chromosomal assignment of sequences homologous to GAD cDNA was determined by Southern hybridization analysis using DNA from mouse-hamster hybrid cells. Mouse genomic sequences homologous to GAD cDNA were isolated and used to determine that GAD is encoded by a locus on mouse chromosome 2 (Gad-1) and that an apparent pseudogene locus is on chromosome 10 (Gad-1ps). An interspecific backcross and recombinant inbred strain sets were used to map these two loci relative to other loci on their respective chromosomes. The Gad-1 locus is part of a conserved homology between mouse chromosome 2 and the long arm of human chromosome 2.     

The XRCC2 and XRCC3 repair genes are required for chromosome stability in mammalian cells.

The irs1 and irs1SF hamster cell lines are mutated for the XRCC2 and XRCC3 genes, respectively. Both show heightened sensitivity to ionizing radiation and particularly to the DNA cross-linking chemical mitomycin C (MMC). Frequencies of spontaneous chromosomal aberration have previously been reported to be higher in these two cell lines than in parental, wild-type cell lines. Microcell-mediated chromosome transfer was used to introduce complementing or non-complementing human chromosomes into each cell line. irs1 cells received human chromosome 7 (which contains the human XRCC2 gene) or, as a control, human chromosome 4. irs1SF cells received human chromosome 14 (which contains the XRCC3 gene) or human chromosome 7. For each set of hybrid cell lines, clones carrying the complementing human chromosome recovered MMC resistance to near-wild-type levels, while control clones carrying noncomplementing chromosomes remained sensitive to MMC. Fluorescence in situ hybridization with a human-specific probe revealed that the human chromosome in complemented clones remained intact in almost all cells even after extended passage. However, the human chromosome in noncomplemented clones frequently underwent chromosome rearrangements including breaks, deletions, and translocations. Chromosome aberrations accumulated slowly in the noncomplemented clones over subsequent passages, with some particular deletions and unbalanced translocations persistently transmitted throughout individual subclones. Our results indicate that the XRCC2 and XRCC3 genes, which are now considered members of the RAD51 gene family, play essential roles in maintaining chromosome stability during cell division. This may reflect roles in DNA repair, possibly via homologous recombination.     

Approaches to the analysis of reciprocal chromosome distribution in the bone marrow cells of mice

A study was made of G-banding chromosome preparations made of bone marrow cells of BALB/c mouse females, injected intraperitoneally with a single dose of the antibiotic adriamycin 23 hours before sacrifice. Frequencies of different chromosome fusions in pericentromeric regions were analysed. All chromosomes were found to be involved in the interchromosomal associations, different chromosomes participating in such fusions with different frequency. No associations of homologous chromosomes were observed. For each chromosome a limited number of preferential partners for fusion was revealed, the number of such partners for different chromosomes varying. Almost for a half of chromosomes, chromosome 15 was one of the most preferred partners. Possible significance of the data obtained is discussed for the analysis of principles of the spatial chromosome arrangement in murine bone marrow cells.