Fine mapping of chromosome 22 breakpoints within the breakpoint cluster region (bcr) implies a role for bcr exon 3 in determining disease duration in chronic myeloid leukemia.

The chromosomal translocation that fuses the phl gene with the c-abl proto-oncogene appears to be a pivotal step in the pathogenesis of some leukemias. In chronic myeloid leukemia (CML) the breakage within the phl gene is largely confined to a 5.8-kb segment referred to as the breakpoint cluster region (bcr). To determine whether the presence of specific bcr exons on the Philadelphia chromosome has any clinical significance, we have analyzed the bcr breakpoints in 134 patients with CML. As many as five probes were used in this analysis, including a synthetic oligonucleotide probe homologous to the bcr exon 3 (phl exon 14) region. The distribution of breakpoints indicates that, in fact, breakage is largely confined to a 3.1-kb segment lying between bcr exon 2 and exon 4 (phl exons 13-15). In 61 CML patients analyzed within 1 year of diagnosis, the distribution of breakpoints appeared to be random within the 3.1-kb region. However, a significant excess of 5' breakpoints was observed in the total population studied, consistent with previous data showing that patients with 3' breakpoints have shorter disease durations. Analysis using the bcr exon 3 sequence probe indicated it was probably the presence or absence of bcr exon 3 on the Philadelphia chromosome that accounts for some of the variability in disease duration seen in CML. The data suggest that the phl/abl protein product may influence the timing of the onset of blast crisis and imply a continuing role for this protein during the evolution of the disease.     

Genes and chromosomal breakpoints in the Langer-Giedion syndrome region on human chromosome 8

The tricho-rhino-phalangeal syndrome type II (TRPS II, or Langer-Giedion syndrome) is an example of contiguous gene syndromes, as it comprises the clinical features of two autosomal dominant diseases, TRPS I and a form of multiple cartilaginous exostoses caused by mutations in the EXT1 gene. We have constructed a contig of cosmid, lambda-phage, PAC, and YAC clones, which covers the entire TRPS I critical region. Using these clones we identified a novel submicroscopic deletion in a TRPS I patient and refined the proximal border of the minimal TRPS1 gene region by precisely mapping the inversion breakpoint of another patient. As a first step towards a complete inventory of genes in the Langer-Giedion syndrome chromosome region (LGCR) with the ultimate aim to identify the TRPS1 gene, we analyzed 23 human expressed sequence tags (ESTs) and four genes (EIF3S3, RAD21, OPG, CXIV) which had been assigned to human 8q24.1. Our analyses indicate that the LGCR is gene-poor, because none of the ESTs and genes map to the minimal TRPS1 gene region and only two of these genes, RAD21 and EIF3S3, are located within the shortest region of deletion overlap of TRPS II patients. Two genes, OPG and CXIV, which are deleted only in some patients with TRPS II may contribute to the clinical variability of this syndrome.     

Restriction fragments homologous to mitochondrial plasmid-like DNAs are located within limited chromosomal regions on the rice nuclear genome

Summary The shifted multiplicative model (SHMM) is used with a cluster method to identify subsets of sites in an international maize (Zea mays L.) trial without genotypic rank-change. For cluster analysis, distance between two sites is defined as the residual sum of squares after fitting SHMM with one multiplicative term (SHMM₁) if SHMM₁ does not show genotypic rank-change. However, if SHMM₁ does show genotypic rank-change, the distance between two sites is defined as the smaller of the sums of squares owing to genotypes within each of the two sites. Calculation of distance between two sites is facilitated by using the site regression model with one multiplicative term (SREG₁), which can be reparameterized as SHMM₁ when only two sites are considered. The dichotomous splitting procedure, used on the dendrogram obtained from cluster analysis, will first perform SHMM analyses on each of the last two cluster groups to join (end of the dendrogram). If SHMM₁ does not give an adequate fit, the next step is to move down the branches of the tree until groups of sites (clusters) are found to which SHMM₁ provides an adequate fit and primary effects of sites are all of the same sign. Five final groups of sites to which SHMM₁ provides an adequate fit and primary effects of sites are all of the same sign were obtained. The procedure appears to be useful in identifying subsets of sites in which genotypic rank-change interactions are negligible.Research reported in this paper (Journal article no. 91-3-218) is part of a project of the Kentucky Agricultural Experiment Station, published with the approval of the Director     

Isolation and mapping of cosmid markers on human chromosome 22, including one within the submicroscopically deleted region of DiGeorge syndrome

A genomic cosmid library was constructed from a Chinese hamster/human hybrid cell containing human intact chromosome 22 as its only human component. Of 1000 cosmids with inserts derived from human chromosome 22, 191 were tested for restriction fragment length polymorphisms (RFLPs). As a result, 64 clones detected RFLPs, including five variable number of tandem repeats systems. Of the remaining 127 cosmids, 111 detected a single copy sequence on human chromosome 22. Five somatic cell hybrids allowed us to assign all of the 64 polymorphic cosmids and 44 non-polymorphic cosmids to four different regions of human chromosome 22. In two patients with DiGeorge syndrome, one of the cosmids that had been sublocalized to 22pter-q11 detected hemizygosity. These 108 cosmid markers regionally assigned to human chromosome 22 should be useful for the construction of long-range physical maps and the identification of genetic alterations on the chromosome.     

A 1.5-Mb contig within the cat eye syndrome critical region at human chromosome 22q11.2

We have constructed a 1.5-Mb contig spanning the distal half of the critical region for cat eye syndrome on human chromosome 22 from D22S543 to D22S181. The contig consists of 20 P1 artificial chromosome (PAC) clones and 11 bacterial artificial chromosome (BAC) clones screened from 2 BAC and 2 PAC libraries. Continuous overlap between the clones was confirmed using vectorette PCR and riboprobes. Despite the instability of this region in a previous YAC contig, only 1 BAC showed a minor instability and then in only one isolation. This contig is now providing the basis for genomic sequencing and gene identification in the cat eye syndrome critical region.     

WNT10A and WNT6, clustered in human chromosome 2q35 region with head-to-tail manner, are strongly coexpressed in SW480 cells

Human WNT10A and WNT6 were cloned and characterized. WNT10A encoded a 417-amino-acid polypeptide with WNT core domain, and WNT6 encoded a 365-amino-acid polypeptide with N-terminal signal peptide, WNT core domain, and RGD motif. WNT10A and WNT6 genes were clustered in the head-to-tail manner with an interval less than 7.0 kb in human chromosome 2q35 region. Among human WNT family, WNT10A was most homologous to WNT10B (59.2% amino-acid identity), and WNT6 was most homologous to WNT1 (47.4% amino-acid identity). WNT10B and WNT1 genes were also clustered in human chromosome 12q13 region. Two WNT gene clusters in human chromosome 2q35 and 12q13 regions might be generated due to duplication of ancestral gene cluster. The 3.0- and 2.4-kb WNT10A mRNAs were expressed in fetal kidney, placenta, adult spleen and kidney. The 2.0-kb WNT6 mRNA was coexpressed with WNT10A in placenta and adult spleen. WNT10A and WNT6 were strongly coexpressed in SW480 (colorectal cancer). In addition to SW480, WNT10A was strongly expressed in HL-60 (promyelocytic leukemia) and Raji (Burkitt's lymphoma), and WNT6 in HeLa S3 (cervical cancer). Overexpression WNT10A and WNT6 might play key roles in human carcinogenesis through activation of WNT-beta-catenin-TCF signaling pathway, just like Wnt10b and Wnt1.     

Molecular cytogenetic investigations define a subgroup of thyroid adenomas with 2p21 breakpoints clustered to a region of less than 450 kb

Structural rearrangements involving chromosome band 2p21 characterize a cytogenetic subgroup of benign thyroid tumors. To narrow down the breakpoints of these aberrations, we established two cell lines from benign thyroid tumors showing translocations involving 2p21. These two cell lines and one additional primary tumor were used for FISH-studies with 18 BAC clones. All breakpoints were mapped to a cluster of about 450 kb.     

Microsatellite motifs with moderate GC content are clustered around genes on Arabidopsis thaliana chromosome 2

Microsatellites, arrays of 1-6 bp sequences, are abundant in almost all the eukaryotic genomes. Their distribution in the genome is widely accepted to be differential and non random along the axis of the chromosomes. Arabidopsis thaliana genome is dominated by mononucleotide repeats, (A)n being the most abundant motif. In total, 39 microsatellite motifs extended to more than 100 bp in length. Of these, 8 loci are devoid of any gene in their proximity. (AG)n is the most abundant motif among longer repeats. The non-random distribution of microsatellite in the genome is reflected as occurrence of microsatellite clusters in the genome. In total, 3400 microsatellite clusters have been identified in the Arabidopsis genome. Chromosome 2, which is 19.7 Mb long, harbors 550 clusters accommodating 29% of all the microsatellites present on this chromosome. Further, 409 of the 6239 genes on chromosome 2 are associated with 323 microsatellite clusters. Motifs like (AGG)n and (ACT)n, show preferential accommodation in clusters that overlap with genes. Among all the microsatellite clusters that show an overlap with genes, 80% of the clusters show an overlap in such a way that the cluster ends beyond the 3'-end of the gene or starts before the 5'-end of a gene. Genes with diverse functions show association with the clusters. However, not all members of a gene family show similar associations.     

Four chromosomal breakpoints and four new probes mark out a 10-cM region encompassing the fragile-X locus (FRAXA)

We report the validation and use of a cell hybrid panel which allowed us a rapid physical localization of new DNA probes in the vicinity of the fragile-X locus (FRAXA). Seven regions are defined by this panel, two of which lie between DXS369 and DXS296, until now the closest genetic markers that flank FRAXA. Of those two interesting regions, one is just distal to DXS369 and defined by probe 2-71 (DXS476), which is not polymorphic. The next one contains probes St677 (DXS463) and 2-34 (DXS477), which are within 130 kb and both detect TaqI RFLPs. The combined informativeness of these two probes is 30%. We cloned from an irradiation-reduced hybrid line another new polymorphic probe, Do33 (DXS465; 42% heterozygosity). This probe maps to the DXS296 region, proximal to a chromosomal breakpoint that corresponds to the Hunter syndrome locus (IDS). The physical order is thus Cen-DXS369-DXS476-(DXS463,DXS477)-(DXS296, DXS465)-IDS-DXS304-tel. We performed a linkage analysis for five of these markers in both the Centre d'Etude du Polymorphisme Humain families and in a large set of fragile-X families. This establishes that DXS296 is distal to FRAXA. The relative position of DXS463 and DXS477 with respect to FRAXA remains uncertain, but our results place them genetically halfway between DXS369 and DXS304. Thus the DXS463-DXS477 cluster defines presently either the closest proximal or the closest distal polymorphic marker with respect to FRAXA. The three new polymorphic probes described here have a combined heterozygosity of 60% and represent a major improvement for genetic analysis of fragile-X families, in particular for diagnostic applications.     

Closing in on the Rieger syndrome gene on 4q25: mapping translocation breakpoints within a 50-kb region.

Rieger syndrome (RGS) is an autosomal dominant disorder of morphogenesis affecting mainly the formation of the anterior eye chamber and of the teeth. RGS has been localized to human chromosome 4q25 by linkage to epidermal growth factor (EGF). We have constructed a detailed physical map and a YAC contig of the genomic region encompassing the EGF locus. Using FISH, several YACs could be shown to cross the breakpoint in two independent RGS patients with balanced 4q translocations. Alu- and LINE-fragmentation of a 2.4-Mb YAC generated a panel of shorter YACs ranging in size from 2.4 Mb to 75 kb. Several fragmentation YACs were subcloned in cosmids, which were mapped to specific subregions of the original YAC by hybridization to the fragmentation panel to further refine the localization of the translocation breakpoints, allowing mapping of the breakpoints to within the most-telomeric 200 kb of the original 2.4-Mb YAC. FiberFISH of cosmids located in this 200-kb region mapped the two translocation breakpoints within a 50-kb region approximately 100-150 kb centromeric to D4S193, significantly narrowing down the candidate region for RGS. The mapping data and resources reported here should facilitate the identification of a gene implicated in Rieger syndrome.     

The order of loci in the pericentric region of chromosome 17, based on evidence from physical and genetic breakpoints.

Previous genetic analyses of chromosome 17 markers and NF1 (Fain et al. 1987) were extended in an attempt to order marker loci that map physically to 17cen----17q12. Three additional markers (HHH202, CRI-L581, and CRI-L946) were included in the analyses. Recombinants within the cluster of seven unordered marker loci were identified by pairwise analyses for each family and by examining the within-sibship segregation patterns for different markers. Changes in the segregation pattern for different loci define genetic breakpoints. Given that interference is complete in the region, markers with the same segregation pattern lie on one side of the breakpoint, while markers with different segregation patterns lie on opposite sides of the breakpoint. If the order of boundary markers is known, markers on each side of a breakpoint can be oriented in relation to the centromere. The order cen-(HHH202/NF1)-(EW207)-(EW203/CRI-L581)- (CRI-L946)-(HOX-2/NGFR)-qter was inferred by combining information from physical breakpoints in a panel of mouse/human hybrids and information from genetic breakpoints found in 16 NF1 families.     

Assemblages of bats are phylogenetically clustered on a regional scale

Phylogenetically related species are assumed to be ecologically similar. Ecological similarity might lead to competition and to low distributional overlap. Therefore, if competitive interactions drive assemblages, we expect a decrease in distributional overlap with increasing phylogenetic relatedness and phylogenetic over-dispersion in assemblages. We tested this hypothesis by evaluating the mean phylogenetic distance of bat assemblages within grid cells of ≈36 km² across Bavaria, Germany (887 grids; 20,023 records). To calculate phylogenetic distance between species, we used a phylogenetic tree derived from sequences of three mitochondrial genes (cytb, COI, ND1), two nuclear-protein-encoding genes (vWF, RAG2) and the genes encoding16S rRNA, 12S rRNA and tRNA-Val. Overall, bat species co-occurring within grid cells were more similar than expected from null models (phylogenetic clustering). This suggests that on the considered scale, bat assemblages are triggered more by environmental filters than by competition. Furthermore, mean phylogenetic distance decreased with the amount of anthropogenic habitats within grids. This contrasts with species richness of bats, which increased with anthropogenic habitats.     

Characterization of a translocation within the von Recklinghausen neurofibromatosis region of chromosome 17

The genetic defect causing von Recklinghausen neurofibromatosis (NF1) has been mapped to the proximal long arm of chromosome 17 by linkage analysis. Flanking markers have been identified, bracketing NF1 in 17q11.2 and laying the foundation for isolating the disease gene. Recently, a family in which a mother and her two children show both the symptoms of NF1 and the presence of a balanced translocation, t(1;17)(p34.3;q11.2), has been identified. We have examined the possibility that the translocation has occurred in or near the NF1 gene by constructing a somatic cell hybrid line containing the derivative chromosome 1 (1qter-p34.3::17q11-qter). On chromosome 1, the breakpoint occurred between SRC2 and D1S57, which are separated by 14 cM. The translocation breakpoint was localized on chromosome 17 between D17S33 and D17S57, markers that also flank NF1 within a region of 4 cM. These data are consistent with the possibility that the translocation event is the cause of NF1 in this pedigree. Consequently, the isolation of the translocation breakpoint, by approach from either the chromosome 1 or the chromosome 17 side, may facilitate the identification of the NF1 gene.     

The mouse C1q genes are clustered on chromosome 4 and show conservation of gene organization

Mouse complement component C1q is a serum glycoprotein which consists of six A chains, six B chains and six C chains. The three polypeptides are 223, 228, and 217 residues long, respectively, and are encoded by three genes. DNA probes for mouse C1q A, B, and C chains were hybridized to Southern blots of DNA obtained from various inbred mouse strains. On the basis of fragment length polymorphisms, two different alleles of each of the genes could be identified. The distribution of these alleles was determined in the BXD and LXPL recombinant inbred strain series. Comparison with previously reported strain distribution patterns shows that the genes encoding mouseClq map to the same locus on distal chromosome 4. Overlapping clones spanning the entire gene cluster ofClq were isolated from genomic libraries using specific cDNA probes. The three genesClqA, ClqB, andClqC are closely arranged on a 19 kilobase stretch of DNA in the 5 to 3 orientation A-C-B. Each gene consists of two exons separated by one intron. Sequence comparison of Clq from three different species have shown that the B chains have the strongest similarity. Southern blot analysis of chromosomal DNA from 14 vertebrate species demonstrated highest similarity between theClqB genes, followed byClqC and finallyClqA.The nucleotide sequence data reported in this paper have been submitted to the EMBL, GenBank, and DDBJ nucleotide sequence databases and have been assigned the accession numbers X92958 (ClqA), X92959 (ClqB), and X92960 (ClqC)