Physical Mapping of Human 7q and 14q Subtelomeric DNA Sequences in the Great Apes

Phylogenetic divergence of the members of the Pongidae familyhas been based on genetic evidence. The terminal repeat array(T₂AG₃) has lately been considered as an additional basis toanalyze genomes of highly related species. The recent isolationof subtelomeric DNA probes specific for human (HSA) chromosomes7q and 14q has prompted us to cross-hybridize them to the chromosomesof the chimpanzee (PTR), gorilla (GGO) and orangutan (PPY) tosearch for its equivalent locations in the great ape species.Both probes hybridized to the equivalent telomeric sites ofthe long (q) arms of all three great ape species. Hybridizationsignals to the 7q subtelomeric DNA sequence probe were observedat the telomeres of HSA 7q, PTR 6q, GGO 6q and PPY 10q, whilehybridization signals to the 14q subtelomeric DNA sequence probewere observed at the telomeres of HSA 14q, PTR 15q, GGO 18qand PPY 15q. No hybridization signals to the chromosome 7-specificalpha satellite DNA probe on the centromeric regions of theape chromosomes were observed. Our observations demonstratesequence homology of the subtelomeric repeat families D7S427and D14S308 in the ape chromosomes. An analogous number of subtelomericrepeat units exists in these chromosomes and has been preservedthrough the course of differentiation of the hominoid species.Our investigation also suggests a difference in the number ofalpha satellite DNA repeat units in the equivalent ape chromosomes,possibly derived from interchromosomal transfers and subsequentamplification of ancestral alpha satellite sequences.     

Mapping of Aldose Reductase Gene Sequences to Human Chromosomes 1, 3, 7, 9, 11, and 13

Aldose reductase (alditol:NAD(P)+ 1-oxidoreductase; EC 1.1.1.21) (AR) catalyzes the reduction of several aldehydes, including that of glucose, to the corresponding sugar alcohol. Using a complementary DNA clone encoding human AR, we mapped the gene sequences to human chromosomes 1, 3, 7, 9, 11, 13, 14, and 18 by somatic cell hybridization. By in situ hybridization analysis, sequences were localized to human chromosomes 1q32-q42, 3p12, 7q31-q35, 9q22, 11p14-p15, and 13q14-q21. As a putative functional AR gene has been mapped to chromosome 7 and a putative pseudogene to chromosome 3, the sequences on the other seven chromosomes may represent other active genes, non-aldose reductase homologous sequences, or pseudogenes.     

Preferential loss of 5q14-21 in intestinal-type gastric cancer with DNA aneuploidy

BACKGROUND: Little is known about the genetic changes associated with DNA ploidy in gastric cancer (GC). The aim of this study was to identify recurrent or specific chromosomal regions of DNA sequence copy number aberrations (DSCNAs) that might harbor genes associated with DNA aneuploidy in GC. METHODS: We analyzed DSCNAs with comparative genomic hybridization and DNA ploidy by laser scanning cytometry in 16 primary intestinal-type GCs. RESULTS: All GCs examined showed at least one DSCNA (loss or gain); eight were DNA diploid (DD) tumors and eight were DNA aneuploid (DA) tumors. The frequent (>30%) DSCNAs were loss of 5q14-21 and gains of 7p11-14, 8q, 20q, and Xq25-26. Recurrent amplifications (>10%) were detected at chromosomal regions 6p, 7p, and 13q. The overall number of DSCNAs was significantly greater in DA than in DD tumors (P = 0.006). Furthermore, the number of aberrations was clearly greater with 5q loss than without 5q loss (P = 0.002). Losses of 5q14-21, 9p21-pter, 16q, and 18q21-qter were preferentially detected in DA tumors. CONCLUSION: The present observations indicate that there is a close relationship between DSCNA and DNA ploidy in intestinal-type GC and that gene(s) at 5q14-21, 9p21-pter, 16q, and/or 18q21-qter may play important roles in acquisition of DNA aneuploidy.     

The gene that encodes the human CD20 (B1) differentiation antigen is located on chromosome 11 near the t(11;14)(q13;q32) translocation site

The human CD20 gene (B1) encodes a B lymphocyte-specific, cell-surface molecule that is involved in B cell activation and differentiation. We report that the CD20 gene is located on human chromosome 11 at position q12-q13. The location of CD20 was determined by in situ hybridization and was further confirmed by Southern blot analysis of DNA from rodent/human hybrids that contained only portions of human chromosome 11. This localization places the CD20 gene near the site of the t(11;14)(q13;q32) translocation that is found in a subgroup of B cell-lineage malignancies. The site of this translocation has been previously identified by DNA cloning and termed bcl-1. The CD20 gene was found to lie on the centromeric side of bcl-1 on chromosome 11 and to be separated from bcl-1 by at least 50 kb of DNA. These results raise the possibility that alterations in the expression of the CD20 gene may result after the t(11;14) chromosomal alteration.     

Interleukin 4 is at 5q31 and interleukin 6 is at 7p15

Summary DNA probes to the human interleukin 4 (IL4) and interleukin 6 (IL6) genes have been used for in situ hybridization to normal human chromosomes and Southern blot analysis of a series of mouse-human hybrid cell lines. IL4 maps to 5q31, the same location as IL5 and other haemopoietic growth factor genes. IL6 maps to 7p15. The significance of these locations is discussed.     

用人染色体14q24.3区带探针池直接分离表达顺序

本文报道了从显微切割的人染色体区带直接分离区带专一性表达序列的方法和结果。     

Identification of Neurofibromatosis 1 (NF1) Homologous Loci by Direct Sequencing, Fluorescence in Situ Hybridization, and PCR Amplification of Somatic Cell Hybrids

Using fluorescence in situ hybridization (FISH), we have identified seven NF1-related loci, two separate loci on chromosome 2, at bands 2q21 and 2q33-q34, and one locus each on five other chromosomes at bands 14q11.2, 15q11.2, 18p11.2, 21q11.2-q21, and 22q11.2. Application of PCR using NF1 primer pairs and genomic DNA from somatic cell hybrids confirmed the above loci, identified additional loci on chromosomes 12 and 15, and showed that the various loci do not share homology beyond NF1 exon 27b. Sequenced PCR products representing segments corresponding to NF1 exons from these loci demonstrated greater than 95% sequence identity with the NF1 locus. We used sequence differences between bona fide NF1 and NF1 -homologous loci to strategically design primer sets to specifically amplify 30 of 36 exons within the 5′ end of the NF1 gene. These developments have facilitated mutation analysis at the NF1 locus using genomic DNA as template.     

Chromosomal localization of homeobox genes and associated markers on porcine Chromosomes 3, 5, 12, 15, 16 and 18: comparative mapping study with human and mouse

Four homeobox genes that belong to the four homeobox gene clusters known in mammals have been regionally assigned to four distinct porcine chromosomes in conserved regions between human and pig. HOXA11, HOXB6, HOXC8, and HOXD4 genes were mapped by radioactive in situ hybridization to porcine Chromosomes (Chrs) 18q21-24 (with a secondary signal in 16q14-21), 12p11-12, 5p11-12, and 15q22-23 respectively. Besides, we have also revealed the presence of a porcine homeobox (pig Hbx24) which, although showing DNA sequence homology with a mouse gene of HOXB cluster, was located on porcine Chr 3 (3p14-13) outside the Hox clusters. To support the identity of the homeobox gene clusters analyzed and in the light of the high sequence similarity among homeobox genes, we also localized markers known to be mapped near each Hox cluster in human. In this way, four genes were also mapped in pig: GAPD (5q12-21), GAD1 (15q21-22), INHBA (18q24), and IGFBP3 (18q24). Mapping of HOXA11, INHBA, and IGFBP3 on pig Chr 18 constitutes the first assignments of genes on this small chromosome. These new localizations extend the information on the conservation of four human chromosomal regions in the pig genome. Received: 7 August 1995 / Accepted: 16 October 1995     

Integration of DNA copy number alterations and transcriptional expression analysis in human gastric cancer

Background

Genomic instability with frequent DNA copy number alterations is one of the key hallmarks of carcinogenesis. The chromosomal regions with frequent DNA copy number gain and loss in human gastric cancer are still poorly defined. It remains unknown how the DNA copy number variations contributes to the changes of gene expression profiles, especially on the global level.

Principal Findings

We analyzed DNA copy number alterations in 64 human gastric cancer samples and 8 gastric cancer cell lines using bacterial artificial chromosome (BAC) arrays based comparative genomic hybridization (aCGH). Statistical analysis was applied to correlate previously published gene expression data obtained from cDNA microarrays with corresponding DNA copy number variation data to identify candidate oncogenes and tumor suppressor genes. We found that gastric cancer samples showed recurrent DNA copy number variations, including gains at 5p, 8q, 20p, 20q, and losses at 4q, 9p, 18q, 21q. The most frequent regions of amplification were 20q12 (7/72), 20q12–20q13.1 (12/72), 20q13.1–20q13.2 (11/72) and 20q13.2–20q13.3 (6/72). The most frequent deleted region was 9p21 (8/72). Correlating gene expression array data with aCGH identified 321 candidate oncogenes, which were overexpressed and showed frequent DNA copy number gains; and 12 candidate tumor suppressor genes which were down-regulated and showed frequent DNA copy number losses in human gastric cancers. Three networks of significantly expressed genes in gastric cancer samples were identified by ingenuity pathway analysis.

Conclusions

This study provides insight into DNA copy number variations and their contribution to altered gene expression profiles during human gastric cancer development. It provides novel candidate driver oncogenes or tumor suppressor genes for human gastric cancer, useful pathway maps for the future understanding of the molecular pathogenesis of this malignancy, and the construction of new therapeutic targets.     

Localization of subtelomeric sequences of human chromosomes 1q, 11p, 13q, and 16q in the higher primates

Relative phylogenetic divergence of the members of the Pongidae family has been based on genetic evidence. The recent isolation of subtelomeric probes specific for human (HSA) chromosomes 1q, 11p, 13q, and 16q has prompted us to cross hybridize these to the chromosomes of the chimpanzee (Pan troglodytes, PTR), gorilla (Gorilla gorilla, GGO), and orangutan (Pongo pygmaeus, PPY) to search for their equivalent locations in the great apes. Hybridization signals to the 1q subtelomeric DNA sequence probe were observed at the termini of human (HSA) 1q, PTR 1q, GGO 1q, PPY 1q, while the fluorescent signals to the 11p subtelomeric DNA sequence probe were observed at the termini of HSA 11p, PTR 9p, GGO 9p, and PPY 8p. Fluorescent signals to the 13q subtelomeric DNA sequence probe were observed at the termini of HSA 13q, PTR 14q, GGO 14q, and PPY 14q, and positive signals to the 16p subtelomeric DNA sequence probe were observed at the termini of HSA 16q, PTR 18q, GGO 17q, and PPY 19q. These findings apparently suggest sequence homology of these DNA families in the ape chromosomes. Obviously, analogous subtelomeric sequences exist in apes' chromosomes that apparently have been conserved through the course of differentiation of the hominoid species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Guinea pig (Cavio cambayo) 5S rRNA genes map to 7q2, 20q2 and 30q2 shown by an R-banded karyotype with PNA-FISH

A karyotype for the guinea pig (Cavio cambayo) is proposed based on an R-banding pattern. R-bands were obtained by BrdU incorporation into the cells followed by a combined DAPI and propidium iodide staining of the fixed metaphase spreads. In situ hybridization was performed with two biotinylated 18-mer PNA (peptide nucleic acid) probes complementary to sequences within the 5S rRNA gene. The 5S rRNA gene repeats map to chromosomes 7q2. 20q2 and 30q2, respectively.     

Chromosomal localization of the gene for human glucosamine-6-sulphatase to 12q14

Summary Glucosamine-6-sulphatase (G6S), a lysosomal enzyme found in all cells, is involved in the catabolism of heparin, heparan sulphate, and keratan sulphate. Deficiency of G6S results in the accumulation of undegraded substrate and the lysosomal storage disorder mucopolysaccharidosis type IIID (Sanfilippo D syndrome). Regional mapping by in situ hybridization of a ³H-labelled human G6S cDNA probe to human metaphase chromosomes indicated that the G6S gene is localized to chromosome 12 at q14. The localization of the G6S gene to chromosome 12 was confirmed using the G6S cDNA clone in Southern blot hybridization analysis of DNA from human x mouse hybrid cell lines.     

Molecular cytogenetic characterization of 17 rob(13q14q) Robertsonian translocations by FISH, narrowing the region containing the breakpoints.

We have characterized 17 rob(13q14q) Robertsonian translocations, using six molecular probes that hybridize to the repetitive sequences of the centromeric and shortarm regions of the five acrocentric chromosomes by FISH. The rearrangements include six de novo rearrangements and the chromosomally normal parents, five maternally and three paternally inherited translocations, and three translocations of unknown origin. The D21Z1/D13Z1 and D14Z1/D22Z1 centromeric alpha-satellite DNA probes showed all rob(13q14q) chromosomes to be dicentric. The rDNA probes did not show hybridization on any of the 17 cases studied. The pTRS-47 satellite III DNA probe specific for chromosomes 14 and 22 was retained around the breakpoints in all cases. However, the pTRS-63 satellite III DNA probe specific for chromosome 14 did not show any signals on the translocation chromosomes examined. In 16 of 17 translocations studied, strong hybridization signals on the translocations were detected with the pTRI-6 satellite I DNA probe specific for chromosome 13. All parents of the six de novo rob(13q14q), including one whose pTRI-6 sequence was lost, showed strong positive hybridization signals on each pair of chromosomes 14 and 13, with pTRS-47, pTRS-63, and pTRI-6. Therefore, the translocation breakpoints in the majority of rob(13q14q) are between the pTRS-47 and pTRS-63 sequences in the p11 region of chromosome 14 and between the pTRI-6 and rDNA sequences within the p11 region of chromosome 13.     

Chromosomal localization of ARSB,the gene for human N-acetylgalactosamine-4-sulphatase

Summary A deficiency of N-acetylgalactosamine-4-sulphatase (G4S, gene symbol ARSB), results in the accumulation of undegraded substrate and the lysosomal storage disorder, Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI). In situ hybridization using an ³H-labelled human G4S genomic DNA fragment to human metaphase chromosomes localized ARSB to chromosome 5q13–5q14. This location is consistent with, an refines, previous chromosomal assignments based on the expression of human G4S in somatic cell hybrids.     

Human Ovalbumin Serpin Evolution: Phylogenic Analysis, Gene Organization, and Identification of New PI8-Related Genes Suggest That Two Interchromosomal and Several Intrachromosomal Duplications Generated the Gene Clusters at 18q21–q23 and 6p25

The human ovalbumin (ov) serpins are associated with tumorigenesis, inflammation, and protection from autolysis by granule proteinases. Their genes are located at 18q21 or 6p25, falling into two structurally very similar but distinct categories depending on the presence or absence of a particular exon. Analysis of ov-serpin gene structure provides an opportunity to elucidate the mechanisms contributing to the formation of the larger serpin gene superfamily. Here we have identified a new gene (PI8L1) at 6p25 that is 72% identical to the 18q21 gene PI8. FISH analysis using the 3′ untranslated region of PI8 yielded an additional signal at 18q23, separable from the known 18q21.3 signal by the t(1;18)(p32;q23) chromosomal translocation. The presence of more than one PI8-related gene was confirmed by analysis of human genomic DNA using the same probe. Cloning and analysis of PI8 showed that its intron number and phasing are identical to those of the 6p25 genes PI6, PI9, and ELANH2, and it lacks the interhelical variable loop exon found in other 18q21 genes. PCR analysis demonstrated that PI5 at 18q21 also lacks this exon, indicating that it is organized identically to the 6p25 genes. By contrast, PI10 and megsin have this exon and resemble the other 18q21 genes, PLANH2, SCCA-1, and SCCA-2, in structure. Using these data with an ov-serpin phylogenic tree we have constructed, we propose that the ov-serpin gene clusters arose via interchromosomal duplication of PI5 (or a precursor) to 6p25, followed by duplication at 6p25, and a more recent interchromosomal duplication from 6p25 to 18q to yield PI8.     

Generation and fluorescent in situ hybridization mapping of yeast artificial chromosomes of 1p, 17p, 17q, and 19q from a hybrid cell line by high-density screening of an amplified library

A yeast artificial chromosome (YAC) library has been constructed from a somatic cell hybrid containing a t(1p;19q) chromosome and chromosome 17. After amplification, part of this library was analyzed by high-density colony filter screening with a repetitive human DNA probe (Alu). The human YACs distinguished by the screening were further analyzed by Alu fingerprinting and Alu PCR. Fluorescent in situ hybridization (FISH) was performed to localize the YACs to subchromosomal regions of chromosome 1p, 17, or 19q. We have obtained a panel of 123 individual YACs with a mean size of 160 kb, and 77 of these were regionally localized by FISH: 33 to 1p, 10 to 17p, 25 to 17q, and 9 to 19q. The YACs cover a total of 19.7 Mb or 9% of the 220 Mb of human DNA contained in the hybrid. No overlapping YACs have yet been detected. These YACs are available upon request and should be helpful in mapping studies of disease loci, e.g., Charcot-Marie-Tooth disease, Miller-Dieker syndrome, hereditary breast tumor, myotonic dystrophy, and malignant hyperthermia.     

Deletion of Chromosomes 13q and 14q Is a Common Feature of Tumors with BRCA2 Mutations

Introduction

Germline BRCA1 or BRCA2 mutations account for 20–30% of familial clustering of breast cancer. The main indication for BRCA2 screening is currently the family history but the yield of mutations identified in patients selected this way is low.

Methods

To develop more efficient approaches to screening we have compared the gene expression and genomic profiles of BRCA2-mutant breast tumors with those of breast tumors lacking BRCA1 or BRCA2 mutations.

Results

We identified a group of 66 genes showing differential expression in our training set of 7 BRCA2-mutant tumors and in an independent validation set of 19 BRCA2-mutant tumors. The differentially expressed genes include a prominent cluster of genes from chromosomes 13 and 14 whose expression is reduced. Gene set enrichment analysis confirmed that genes in specific bands on 13q and 14q showed significantly reduced expression, suggesting that the affected bands may be preferentially deleted in BRCA2-mutant tumors. Genomic profiling showed that the BRCA2-mutant tumors indeed harbor deletions on chromosomes 13q and 14q. To exploit this information we have created a simple fluorescence in situ hybridization (FISH) test and shown that it detects tumors with deletions on chromosomes 13q and 14q.

Conclusion

Together with previous reports, this establishes that deletions on chromosomes 13q and 14q are a hallmark of BRCA2-mutant tumors. We propose that FISH to detect these deletions would be an efficient and cost-effective first screening step to identify potential BRCA2-mutation carriers among breast cancer patients without a family history of breast cancer.     

Molecular cytogenetic characterization of pancreas cancer cell lines reveals high complexity chromosomal alterations

Karyotype analysis can provide clues to significant genes involved in the genesis and growth of pancreas cancer. The genome of pancreas cancer is complex, and G-band analysis cannot resolve many of the karyotypic abnormalities seen. We studied the karyotypes of 15 recently established cell lines using molecular cytogenetic tools. Comparative genomic hybridization (CGH) analysis of all 15 lines identified genomic gains of 3q, 8q, 11q, 17q, and chromosome 20 in nine or more cell lines. CGH confirmed frequent loss of chromosome 18, 17p, 6q, and 8p. 14/15 cell lines demonstrated loss of chromosome 18q, either by loss of a copy of chromosome 18 (n = 5), all of 18q (n = 7) or portions of 18q (n = 2). Multicolor FISH (Spectral Karyotyping, or SKY) of 11 lines identified many complex structural chromosomal aberrations. 93 structurally abnormal chromosomes were evaluated, for which SKY added new information to 67. Several potentially site-specific recurrent rearrangements were observed. Chromosome region 18q11.2 was recurrently involved in nine cell lines, including formation of derivative chromosomes 18 from a t(18;22) (three cell lines), t(17;18) (two cell lines), and t(12;18), t(15;18), t(18;20), and ins(6;18) (one cell line each). To further define the breakpoints involved on chromosome 18, YACs from the 18q11.2 region, spanning approximately 8 Mb, were used to perform targeted FISH analyses of these lines. We found significant heterogeneity in the breakpoints despite their G-band similarity, including multiple independent regions of loss proximal to the already identified loss of DPC4 at 18q21.     

Porcine alpha-1-antitrypsin (PI): cDNA sequence, polymorphism and assignment to chromosome 7q2.4- > q2.6

A cDNA clone encoding the complete coding sequence for porcine alpha-1-antitrypsin (or α₁-protease inhibitor, PI) was isolated and its DNA sequence determined. The cDNA is assumed to encode alpha-1-antitrypsin on the basis of its sequence similarity to the corresponding cDNAs for human, baboon, rat, mouse, sheep and cow. The porcine cDNA clone was used in conjunction with BamHI, KpnI, MspI, SacI, TuqI and XbaI to develop restriction fragment length polymorphism-based genetic markers for linkage mapping in pigs. The cDNA has also been used to map the porcine PI locus to chromosome 7q2.4- > q2.6 by radioactive in situ hybridization. Thus, the PI locus has been added to the developing physical and genetic maps of the porcine genome.