Mouse chromosome 19 and distal rat chromosome 1: a chromosome segment conserved in evolution

Through a combination of radiation hybrid mapping and studies by FISH and zoo-FISH we have made a comparative investigation of the distal portion of rat chromosome 1 (RNO1) and the entire mouse chromosome 19 (MMU19). It was found that homologous segments of RNO1 and MMU19 are similar in banding morphology and in length as determined by several different methods, and that the gene order of the 46 genes studied appears to be conserved across the homologous segments in the two species. High-resolution zoo-FISH techniques showed that MMU19 probes highlight only a continuous segment on RNO1 (1q43-qter), with no detectable signals on other rat chromosomes. We conclude that these data suggest the evolutionary conservation of a chromosomal segment from a common rodent ancestor. This segment now constitutes the entire MMU19 and a large segment distally on RNO1q in the mouse and rat, respectively.     

Comparative mapping of seven genes in mouse, rat and Chinese hamster chromosomes by fluorescence in situ hybridization

By fluorescence in situ hybridization (FISH) using mouse probes, we assigned homologues for cathepsin E (Ctse), protocadherin 10 (Pcdh10, alias OL-protocadherin, Ol-pc), protocadherin 13 (Pcdh13, alias protocadherin 2c, Pcdh2c), neuroglycan C (Cspg5) and myosin X (Myo10) genes to rat chromosomes (RNO) 13q13, 2q24-->q25, 18p12-->p11, 8q32.1 and 2q22.1-->q22.3, respectively. Similarly, homologues for mouse Ctse, Pcdh13, Cspg5 and Myo10 genes and homologues for rat Smad2 (Madh2) and Smad4 (Madh4) genes were assigned to Chinese hamster chromosomes (CGR) 5q28, 2q17, 4q26, 2p29-->p27, 2q112-->q113 and 2q112-->q113, respectively. The chromosome assignments of homologues of Ctse and Cspg5 reinforced well-known homologous relationships among mouse chromosome (MMU) 1, RNO 13 and CGR 5q, and among MMU 9, RNO 8 and CGR 4q, respectively. The chromosome locations of homologues for Madh2, Madh4 and Pcdh13 genes suggested that inversion events were involved in chromosomal rearrangements in the differentiation of MMU 18 and RNO 18, whereas most of MMU 18 is conserved as a continuous segment in CGR 2q. Furthermore, the mapping result of Myo10 and homologues suggested an orthologous segment of MMU 15, RNO 2 and CGR 2.     

Chromosome evolution of MMU16 and RNO11: conserved synteny associated with gene order rearrangements explicable by intrachromosomal recombinations and neocentromere emergence

Comparative mapping between the rat and mouse genomes has shown that some chromosomes are entirely or almost entirely conserved with respect to gene content. Such is the case of rat chromosome 11 (RNO11) and mouse chromosome 16 (MMU16). We determined to what extent such an extensive conservation of synteny is associated with a conserved gene order. Therefore, we regionally localized several genes on RNO11. The comparison of the gene map of RNO11 and MMU16 unambiguously shows that the gene order has not been conserved in the Murinae lineage, thereby implying the occurrence of intrachromosomal evolutionary rearrangements. The transition from one chromosome configuration to the other one can be explained either by two intrachromosomal recombinations or by a single intrachromosomal recombination accompanied by neocentromere emergence.     

Evolutionary aspects of the genomic organization of rat chromosome 10

Using FISH and RH mapping a chromosomal map of rat chromosome 10 (RNO10) was constructed. Our mapping data were complemented by other published data and the final map was compared to maps of mouse and human chromosomes. RNO10 contained segments homologous to mouse chromosomes (MMU) 11, 16 and 17, with evolutionary breakpoints between the three segments situated in the proximal part of RNO10. Near one of these breakpoints (between MMU17 and 11) we found evidence for an inversion ancestral to the mouse that was not ancestral to the condition in the rat. Within each of the chromosome segments identified, the gene order appeared to be largely conserved. This conservation was particularly clear in the long MMU11-homologous segment. RNO10 also contained segments homologous to three human chromosomes (HSA5, 16, 17). However, within each segment of conserved synteny were signs of more extensive rearrangements. At least 13 different evolutionary breakpoints were indicated in the rat-human comparison. In contrast to what was found between rat and mouse, the rat-human evolutionary breaks were distributed along the entire length of RNO10.     

Further insights into the ancestral murine karyotype: the contribution of the Otomys-Mus comparison using chromosome painting

The African vlei rat, Otomys irroratus, comprises several distinct chromosomal races that may be grouped into two major cytogenetic clades. Recognition of these clades is underpinned by a complex chromosomal rearrangement involving three different autosomes in the unfused state. We have used unidirectional fluorescence in situ hybridization (FISH) of mouse chromosome-specific painting probes to molecularly define the components of this rearrangement as well as to establish the chromosomal homologies between the mouse and the vlei rat genomes. This has allowed for the detection of 41 autosomal segments of conserved synteny. Nine mouse chromosomes were conserved in toto (MMU3, 4, 6, 7, 11, 12, 14, 18, 19) with a further seven (MMU2, 5, 8, 9, 10, 13, 16) showing homology to two discrete regions in the vlei rat genome. Two mouse autosomes (MMU15, 17) correspond to three regions in O. irroratus with MMU1 being the most fragmented showing five sites of hybridization in this species. By mapping these data to published sequence-based phylogenies we are able to confirm most of the published putative ancestral murine chromosomal states. Our data further indicate that MMU15a+ MMU13b+MMU10b+MMU17b was present in the murine ancestral karyotype suggesting an ancestral 2n = 52 rather than the 2n = 54 previously postulated.     

Analysis of chromosomal aberrations involving chromosome 1q31-->q53 in a DMBA-induced rat fibrosarcoma cell line: amplification and overexpression of Jak2

In a study of DMBA-induced rat fibrosarcomas we repeatedly found deletions and/or amplifications in the long arm of rat chromosome 1 (RNO1). Comparative genome hybridization showed that there was amplification involving RNO1q31-->q53 in one of the DMBA-induced rat fibrosarcoma tumors (LB31) and a cell culture derived from it. To identify the amplified genes we physically mapped rat genes implicated in cancer and analyzed them for signs of amplification. The genes were selected based on their locations in comparative maps between rat and man. The rat proto-oncogenes Ccnd1, Fgf4, and Fgf3 (HSA11q13.3), were mapped to RNO1q43 by fluorescence in situ hybridization (FISH). The Ems1 gene was mapped by radiation hybrid (RH) mapping to the same rat chromosome region and shown to be situated centromeric to Ccnd1 and Fgf4. In addition, the proto-oncogenes Hras (HSA11p15.5) and Igf1r (HSA15q25-->q26) were mapped to RNO1q43 and RNO1q32 by FISH and Omp (HSA11q13.5) was assigned to RNO1q34. PCR probes for the above genes together with PCR probes for the previously mapped rat genes Bax (RNO1q31) and Jak2 (RNO1q51-->q53) were analyzed for signs of amplification by Southern blot hybridization. Low copy number increases of the Omp and Jak2 genes were detected in the LB31 cell culture. Dual color FISH analysis of tumor cells confirmed that chromosome regions containing Omp and Jak2 were amplified and were situated in long marker chromosomes showing an aberrant banding pattern. The configuration of the signals in the marker chromosomes suggested that they had arisen by a break-fusion-bridge (BFB) mechanism.     

Karyotypic evolution of hapalomys inferred from chromosome painting: a detailed characterization contributing new insights into the ancestral murinae karyotype

We report on the construction of a comparative chromosome map between the emblematic laboratory rat, Rattus norvegicus (RNO), and Delacour's Marmoset rat, Hapalomys delacouri (HDE), based on cross-species fluorescence in situ hybridization with R. norvegicus painting probes. Sixteen R. norvegicus chromosomes (RNO 3-6, 8, 10-15, 17-20, and X) were retained in their entirety (as a conserved block or as a single chromosome) in the H. delacouri genome. The remaining 5 R. norvegicus chromosomes (RNO 1, 2, 7, 9, and 16) produced 2 signals in the H. delacouri karyotype. Our analysis allowed the detection of an X-autosome translocation between RNO X and 11 that occurred convergently in an unrelated species, Bandicota savilei, and a single B chromosome that accounts for the 2n = 48 karyotype observed in this specimen. In total, the rat chromosome paints revealed 27 segments of conserved synteny in H. delacouri. The analysis showed 7 NOR bearing pairs in H. delacouri (HDE 1, 3, 6, 7, 8, 10, and 13) and the occurrence of an interstitial telomeric signal at the centromeric regions of 8 H. delacouri chromosomes (HDE 3, 10, 11, 12, 13, 16, 19, and 22). These data, together with published comparative maps, enabled a revision of the previously postulated murine ancestral condition suggesting that it probably comprised a wholly acrocentric karyotype with 2n = 46-50.     

Comparative mapping of human Chromosome 14q11.2-q13 genes with mouse homologous gene regions

An examination of the synteny blocks between mouse and human chromosomes aids in understanding the evolution of chromosome divergence between these two species. We comparatively mapped the human (HSA) Chromosome (Chr) 14q11.2-q13 cytogenetic region with the intervals of orthologous genes on mouse (MMU) chromosomes. A lack of conserved gene order was identified between the human cytogenetic region and the interval of orthologs on MMU 12. The evolutionary breakpoint junction was defined within 2.5 Mb, where the conserved synteny of genes on HSA 14 changes from MMU 12 to MMU 14. At the evolutionary breakpoint junction, a human EST (GI: 1114654) with identity to the human and mouse BCL2 interacting gene, BNIP3, was mapped to mouse Chr 3. New gene homologs of LAMB1, MEOX2, NRCAM, and NZTF1 were identified on HSA 7 and on the proximal cytogenetic region of HSA 14 by mapping mouse genes recently reported to be genetically linked within the relevant MMU 12 interval. This study contributes to the identification of homology relationships between the genes of HSA 14q11.2-q13 and mouse Chr 3, 12, and 14. Received: 16 March 2000 / Accepted: 16 June 2000     

The gene for cystathionine beta-synthase (CBS) maps to the subtelomeric region on human chromosome 21q and to proximal mouse chromosome 17

The human gene for cystathionine beta-synthase (CBS), the enzyme deficient in classical homocystinuria, has been assigned to the subtelomeric region of band 21q22.3 by in situ hybridization of a rat cDNA probe to structurally rearranged chromosomes 21. The homologous locus in the mouse (Cbs) was mapped to the proximal half of mouse chromosome 17 by Southern analysis of Chinese hamster X mouse somatic cell hybrid DNA. Thus, CBS/Cbs and the gene for alpha A-crystalline (CRYA1/Crya-1 or Acry-1) form a conserved linkage group on human (HSA) chromosome region 21q22.3 and mouse (MMU) chromosome 17 region A-C. Features of Down syndrome (DS) caused by three copies of these genes should not be present in mice trisomic for MMU 16 that have been proposed as animal models for DS. Mice partially trisomic for MMU 16 or MMU 17 should allow gene-specific dissection of the trisomy 21 phenotype.     

Localization of the mouse thymidine kinase gene to the distal portion of chromosome 11

We report the cytogenetic mapping of the thymidine kinase (tk-1) gene in the mouse using two complementary and independent analyses: (1) investigation of chromosome aberrations associated with tk-1 gene inactivation in the L5178Y TK+/- -3.7.2C cell line, and (2) fluorescence in situ molecular hybridization of cloned tk-1 cDNA probes to mitotic chromosomes of this cell line. The consensus location from both analyses is 11E1-E2. Consideration of the mouse tk-1 gene localization, along with evidence that the homologous human TK1 gene is located distally on the large arm of chromosome 17, appears to extend the region of homology between MMU11 and HSA17 to the distal end of both chromosomes.     

Towards identification of individual homologous chromosomes: comparative genomic hybridization and spectral karyotyping discriminate between paternal and maternal euchromatin in Mus musculus x M. spretus interspecific hybrids

We have developed an in situ technique to label individual euchromatic chromosome arms in interspecific crosses between Mus musculus (MMU) and M. spretus (MSP). The MMU and MSP genomes diverged 2-3 million years ago and show an overall sequence divergence of approximately 1%. Comparative hybridization of MMU versus MSP DNA and subsequent spectral analysis of the euchromatic hybridization profiles discriminated between maternal (MMU) and paternal (MSP) chromosomes in F(1) hybrids. Dispersed repetitive DNA elements were the preferred hybridization target of MMU DNA on maternal chromosomes and of MSP DNA on paternal chromosomes. Differences in centromeric satellite DNAs were detected by conventional fluorescence in situ hybridization and served as internal controls. Our experiments suggest that it is possible, in principle, to discriminate between paternal and maternal chromosomes on the basis of sequence differences.     

Comparative mapping using somatic cell hybrids

Summary Comparative mapping, or ascertaining the gene linkage relationships between different species, is rapidly developing. This is possible because new techniques in chromosome identification and somatic cell hybridization, such as the generation of hybrids preferentially segregating chromosomes of any desired species including rodents, and the development of gene transfer techniques have yielded new information about the human and rodent gene maps. In addition, the discovery and characterization of mouse subspecies has generated new mouse sexual genetic linkage data. The following picture is emerging. Several X-linked genes in man are X-linked in all mammalian species tested. The linkage relationships of several tightly linked genes, less than 1 map unit apart, are also conserved in all mammalian species tested. Ape autosomal genes are assigned to ape chromosomes homologous to their human counterparts indicating extensive conservation in the 12 million years (MYR) of evolution from apes to man. Similarly, mouse and rat, 10 MYR apart in evolution, have several large autosomal synteny groups conserved. In comparing the mouse and human gene maps we find that human genes assigned to different arms of the same human chromosome are unlinked in the mouse; mouse genes large map distances (20 to 45 cM) apart are very likely to be unlinked in the human. However, several autosomal synteny groups 10 to 20 cM apart, including thePgd, Eno-1, Pgm-1 group on human chromosome arm lp, are conserved in mice and man. This suggests that homology mapping, the superimposition of one species gene map on the homologous conserved portion of another species genome may be possible, and that ancestral autosomal synteny groups should be detectable. Presented in the formal symposium on Somatic Cell Genetics at the 27th Annual Meeting of the Tissue Culture Association, Philadelphia, Pennsylvania, June 7–10, 1976.     

Genomic structure,conservation and FISH mapping of the Rattus norvegicus Adprt gene

Poly(ADP-ribose) polymerase 1 (PARP-1) lies at the basis of a DNA-interacting protein family that maintains genome integrity. Here we describe the genomic organisation of rat PARP-1 gene (Adprt), refine its assignment to rat chromosome (RNO) 13q25-->q26 by FISH and compare its genomic organisation between rat, mouse and human. It appears that in human, mouse and rat Adprt consists of 23 similar-sized exons with well-conserved intron and exon borders. Adprt orthologs map to homologous chromosome regions at the termini of the q-arms of human and mouse chromosomes 1 and rat 13, with gene order being conserved between the rodents. Kimura protein distance comparison with human PARP-1 as reference revealed the bovine protein to be the least conserved with 10.3 substitutions per 100 amino acids, followed by rat (8.6) and mouse (8.4).     

Chromosomal localization of ceruoplasmin and transferrin genes in laboratory rats,mice and in man by hybridization with specific DNA probes

Fragments of the natural rat ceruloplasmin (Cp) gene and cDNA copies of rat Cp and transferring (Tf) mRNAs highly labelled by nick translation with ¹²⁵I-dCTP were used as specific probes for assignment of these genes to the metaphase chromosomes of rat, mouse and man by in situ hybridization. Both Cp and Tf genes were found to be syntenic in rodents, occupying with high probability the regions 9D and 9F1–3 in mice and 7q11–13 and 7q31–34 in rats respectively. The significant increase in silver grain count over chromosome 15 in rats after hybridization with both the Cp and Tf probes suggests the presence of a related pseudogene cluster on this particular chromosome and thus favours its partial homeology to chromosome 7. The localization of silver grains in metaphase chromosome of man indicates subregional assignment of the Tf gene to 3q21. Use of the rat Cp DNA probe does not indicate synteny of the Cp and Tf genes in man and suggests the existence of a related DNA sequence in 15q11–13. The potential and limitations of the in situ hybridization technique with heterologous DNA probes for gene mapping in mammalian species are discussed.     

High-resolution comparative physical mapping of mouse Chromosome 10 in the region of homology with human Chromosome 21

Comparative mapping of human and mouse chromosomes can be used to predict locations of homologous loci between the species, provides the substrate to examine the process of chromosomal evolution, and facilitates the continuing development of mouse genetic models for human disorders. A YAC contig of the region of mouse Chromosome (Chr) 10 (MMU10) that demonstrates conserved linkage with the distal portion of human Chr 21 (HSA21) has been constructed. The contig contains all known genes mapped in both species, defines the proximal region of homology between MMU10 and HSA22, and contains the evolutionary junction between HSA21 and HSA22 on MMU10. It consists of 23 YACs and 2 PACs, and covers 3.2 Mb of MMU10. The average marker density for this region is 1 marker/69 kb. Nine of 22 expressed sequences are mapped here for the first time in mouse, and two are newly characterized expressed sequences. The contig also contains 12 simple sequence repeats (SSRs) and 16 YAC and PAC endclone markers. YAC fragmentation analysis was used to create a physical map for the proximal 2.2 Mb of the contig. Cloning of the corresponding region of HSA21 has proven difficult, and the mouse contig includes segments absent from previously described sequence ready maps of HSA21. Received: 22 July 1998 / Accepted: 13 November 1998     

Chromosomal assignment of retinoic acid receptor (RAR) genes in the human, mouse, and rat genomes.

The human genes encoding the alpha and beta forms of the retinoic acid receptor are known to be located on chromosomes 17 (band q21.1:RARA) and 3 (band p24:RARB). By in situ hybridization, we have now localized the gene for retinoic acid receptor gamma, RARG, on chromosome 12, band q13. We also mapped the three retinoic acid receptor genes in the mouse, by in situ hybridization, on chromosomes 11, band D (Rar-a); 14, band A (Rar-b); and 15, band F (Rar-g), respectively, and in the rat, using a panel of somatic cell hybrids that segregate rat chromosomes, on chromosomes 10 (RARA), 15 (RARB), and 7 (RARG), respectively. These assignments reveal a retention of tight linkage between RAR and HOX gene clusters. They also establish or confirm and extend the following homologies: (i) between human chromosome 17, mouse chromosome 11, and rat chromosome 10 (RARA); (ii) between human chromosome 3, mouse chromosome 14, and rat chromosome 15 (RARB); and (iii) between human chromosome 12, mouse chromosome 15, and rat chromosome 7 (RARG).     

Cloning, mapping and expression of UBL3, a novel ubiquitin-like gene.

A novel Drosophila melanogaster gene UBL3 was characterized and shown to be highly conserved in man and Caenorhabditis elegans (C. elegans). The human and mouse homologues were cloned and sequenced. UBL3 is a ubiquitin-like protein of unknown function with no conserved homologues in yeast. Mapping of the human and mouse UBL3 genes places them within a region of shared gene order between human and mouse chromosomes on human chromosome 13q12-13 and telomeric mouse chromosome 5 (MMU5).     

Use of comparative physical and sequence mapping to annotate mouse chromosome 16 and human chromosome 21

Distal mouse chromosome 16 (MMU16) shares conserved linkage with human chromosome 21 (HSA21), trisomy for which causes Down syndrome (DS). A 4.5-Mb physical map extending from Cbr1 to Tmprss2 on MMU16 provides a minimal tiling path of P1 artificial chromosomes (PACs) for comparative mapping and genomic sequencing. Thirty-four expressed sequences were positioned on the mouse map, including 19 that were not physically mapped previously. This region of the mouse:human comparative map shows a high degree of evolutionary conservation of gene order and content, which differs only by insertion of one gene (in mouse) and a small inversion involving two adjacent genes. "Low-pass" (2.2x) mouse sequence from a portion of the contig was ordered and oriented along 510 kb of finished HSA21 sequence. In combination with 68 kb of unique PAC end sequence, the comparison provided confirmation of genes predicted by comparative mapping, indicated gene predictions that are likely to be incorrect, and identified three candidate genes in mouse and human that were not observed in the initial HSA21 sequence annotation. This comparative map and sequence derived from it are powerful tools for identifying genes and regulatory regions, information that will in turn provide insights into the genetic mechanisms by which trisomy 21 results in DS.     

Comparative genomic in situ hybridization discloses recurrent gain of chromosome 4 in experimental gliomas of the rat.

The genetic characterization of experimental tumors is essential in order to evaluate their relevance as appropriate animal models for human neoplasms. We have used flow cytometry and a recently established Comparative Genomic in situ Hybridization (CGH) protocol for the rat (Kappler et al., 1998) to investigate chromosome copy number changes in five ethylnitrosourea induced gliomas of the rat. Flow cytometry showed aneuploid DNA indices in three of the tumors investigated. CGH analysis of primary tumors revealed whole chromosome and subchromosomal gains of rat chromosomes (RNO) 1, 2, 4, 6, 7, 10, 11, 12, and 13. Loss of RNO 5q23-->q35 was apparent in one tumor. High level copy number gains were not observed using CGH as well as semiquantitative PCR with Tgfa, Met and Hbb primers. Low copy number gain of RNO 4 represents the most common aberration, since it was detected in four of five tumors investigated. Three tumors showed gain of RNO 7, while two tumors showed gains of RNO 10q31-->qter and RNO 12q. Deletion of RNO 5q23-->q35 and gain of RNO 4 occurred mutually exclusively. Therefore, we conclude that these two alterations may represent different pathways in the pathogenesis of experimental gliomas in the rat. Findings are discussed in analogy to human gliomas.     

The physical map ofMus musculus chromosome 11 reveals evolutionary relationship with different syntenic groups of genes inHomo sapiens

Summary The physical localization of sequences homologous to three cloned genes was determined by in situ hybridization to metaphase chromosomes. Previous work had assigned the skeletal myosin heavy chain gene cluster (Myh), the functional locus for the cellular tumor antigen p53 (Trp53-1), and the cellular homologue of the viral erb-B oncogene (Erbb) toMus musculus chromosome 11 (MMU11). Our results provide regional assignments ofMyh andTrp53-1 to chromosome bands B2C, and ofErbb to bands A1A4. Taken together with in situ mapping of three other loci on MMU 11 (Hox-2 homeobox-containing gene cluster, theSparc protein, and theColla-1 collagen gene), which have been reported elsewhere, these data allowed us to construct a physical map of MMU11 and to compare it with the linkage map of this chromosome. The map positions of the homologous genes on human chromosomes suggest evolutionary relationships of distinct regions of MMU11 with six different human chromosome arms: 1p, 5q, 7p, 16p, 17p, and 17q. The delineation of conserved chromosome regions has important implications for the understanding of karyotype evolution in mammalian species and for the development of animal models of human genetic diseases.