Evidence that activities of coagulation factors VII and X are linked to chromosome 13 (q34)

In 7 patients with various anomalies of chromosome 13 coagulation, studies were performed. A 50% decrease of activities of factor VII and X were noted only in cases with deletion of 13 (q34) which supports the hypothesis that they are linked to this region.     

Structural genes of coagulation factors VII and X located on 13q34

From 7 cases of abnormalities involving chromosome 13, the structural gene(s) coding for coagulation factors VII and X were located in the region 13q34-13qter. Gene-dosage effects for these coagulation factors seem to act in both directions, causing a decrease when there is monosomy of segment 13q34, but also, as has not been demonstrated before, an increase when there is trisomy of this same segment.     

The structural gene for human coagulation factor X is located on chromosome 13q34

The gene coding for coagulation factor X was studied in a family segregating chromosomal abnormalities involving chromosomes 13 and 6. An individual monosomic for 13q34 was deficient in levels of clotting factors VII and X, while her brother, who is trisomic for 13q34, had elevated levels. DNA dosage studies with a cloned human factor X gene demonstrated that the low levels of factor X expression in the individual with the chromosome 13q34 deletion were due to the absence of one copy of the factor X structural gene. This confirms the assignment of the human gene coding for factor X to 13q34.     

Deficiency of coagulation factors VII and X associated with deletion of a chromosome 13 (q34). Evidence from two cases with 46,XY,t(13;Y)(q11;q34)

Summary Deficiency of coagulation factors VII and X was found in two patients (Erlangen and Nancy) who shared the same chromosomal aberration 46,XY,t(13;Y)(q11;q34) with probable loss of a terminal segment of 13q. Loci involved in synthesis or constitution of these factors may be located at 13 (q34).The paper is dedicated to Prof. Dr. G. Koch on the occasion of his 70th birthday     

The gene for clotting factor 10 is mapped to 13q32----qter

The structural gene for the human clotting factor 10 (F10) has been mapped to chromosome 13 with a cDNA probe hybridized to DNAs from a panel of human X hamster hybrids. In situ hybridization was used to assign F10 to region 13q32----qter of chromosomes from normal human lymphocytes.     

Enzyme-linked coagulation assay. III. Sensitive immunoassays for clotting factors II, VII, and X

The solid-phase clotting assay utilizing fibrinogen coated on the wells of a microtiter plate and peroxidase-fibrinogen in solution as a substrate for thrombin (enzyme-linked coagulation assay, ELCA) has been modified for use as an immunoassay. Direct inhibition of factors II, VII, and X by polyclonal (rabbit) antibodies and of factor X by monoclonal antibodies has been demonstrated at high dilution of these antibodies and detection of the specific factors using ELCA. Using plates coated with a second antibody (goat anti-mouse IgG) as well as fibrinogen, monoclonal antibodies to factors X and VII were measured by binding the active factor to the plate and detection of the bound factor using ELCA. The assay was very sensitive, permitting the detection of as little as 0.2 ng/ml (30 pg/assay) of monoclonal antibody, or less than 0.4 ng/ml (60 pg/assay) of factor Xa. When plates were coated with monoclonal antibody to factor X and fibrinogen, the assay permitted the identification of distinct epitope specificities for two monoclonal antibodies to factor X by distinct competition of the monoclonal antibodies added in the solution phase for binding of factor Xa to the plate. This assay could be applied generally for immunoassay of clotting factors, and could have application in general as an immunoassay amplification system.     

Proximal trisomy 13q and distal monosomy 8p in a dysmorphic and mentally retarded patient with an isodicentric chromosome 13q and a 13q/8p translocation chromosome

A 40 year-old dysmorphic and mentally retarded female is reported with a de novo unbalanced chromosomal rearrangement (karyotype: 46,XX,der(8)t(8;13)(p23;q123),idic(13)(pter-->q123: q123-->pter) resulting in an isodicentric chromosome 13 and a double aneusomy including partial trisomy 13 (13pter-q123) and distal monosomy 8p (8pter-p23). The main clinical findings consist of developmental/mental retardation, behavioural disturbances and minor congenital defects, not consistent with the clinical pattern of either of the two aneusomies. A mechanism for the chromosome rearrangement is proposed and the absence of specific physical findings in the present patient is discussed in the light of the available literature data.     

Large-scale mapping and chromosome jumping in the q27 region of the human X chromosome

We have used pulsed field gel electrophoresis for further physical mapping studies in the q27 region of the human X chromosome. We show that the DXS 102 locus and the F9 gene are separated by only 300 kb despite a genetic distance of 1.4 cM; this linkage orients our large-scale map and shows that the mcf.2 transforming sequence is telomeric to F9. A BssHII complete-digest jumping library was used to jump toward the DXS 105 locus; a 130-kb jump was achieved and the corresponding "linking clone" was obtained.     

Regional localization of the human platelet-derived endothelial cell growth factor (ECGF1) gene to chromosome 22q13.

Using in situ hybridization and a panel of human X rodent somatic cell hybrids, which discriminates between four different regions of human chromosome 22, we have localized the gene for human platelet-derived endothelial cell growth factor (ECGF1) to 22q13, placing ECGF1 distal to the PDGFB locus at 22q12.3----q13.1.     

Macrorestriction mapping of COL4A1 and COL4A2 collagen genes on human chromosome 13q34

The genes for the alpha-1 and alpha-2 chains of type IV collagen (COL4A1 and COL4A2) map to the same chromosomal band (13q34) and have a high degree of nucleotide homology. We have used pulsed field gel electrophoresis and cloned COL4A1 and COL4A2 DNA fragments as molecular probes to construct a 1200-kb macrorestriction map which encompasses both genes. The two genes are located within a 340-kb region with the 3' end of COL4A2 and the 5' region of COL4A1 separated by at least 100 kb but not more than 160 kb. These genes, therefore, are two members of a gene cluster on chromosome 13q34.     

Regional mapping of the FOS oncogene to rat chromosome 6q21----q23

Using a panel of rat x mouse somatic cell hybrids and in situ hybridization, we determined the chromosomal location of the rat FOS gene locus. Southern blot analysis assigned this oncogene to rat chromosome 6. In situ hybridization confirmed this finding and mapped the gene more precisely to 6q21----q2.     

Physical and cDNA mapping in the DBH region of human chromosome 9q34

Chromosome 9q34 has been extensively studied and mapped due to the presence of known disease genes, principally tuberous sclerosis 1 (TSC1), in this region. During the course of our mapping of this region we constructed a 555-kb contig beginning approximately 50 kb proximal to the dopamine-beta-hydroxylase (DBH) gene and extending, with one small deletion, distal to the D9S114 marker. The contig consists of 11 P1 clones, four PAC clones, one BAC clone and six cosmid clones and contains 27 new nonpolymorphic STSs. We have found the region to be unstable in P1, PAC and BAC cloning vehicles and have identified several deleted genomic clones. In addition, we have isolated and mapped the 3' portions of three putative genes located within or immediately distal to the DBH gene, including one large gene that runs on the opposite strand to DBH and utilizes portions of two DBH exons. The genomic clones of the contig, cDNAs and new STSs will be useful reagents for the further study and mapping of this region.     

Exclusion of human chromosome 13q34 as the site of the cystic fibrosis mutation.

We have studied a family in which both cystic fibrosis (CF) and an unbalanced translocation between chromosomes 6 and 13 are found. As CF occurs in the child who is effectively monosomic for the translocated part of the long arm of chromosome 13, it was suggested that the locus of the gene mutation causing CF is on chromosome 13q34. The gene for human coagulation factor X is located at 13q34, and we have found a restriction fragment length polymorphism (RFLP) that is revealed by a cloned cDNA coding for this protein. Linkage analysis in eight CF families shows no evidence of cosegregation between CF and the gene for factor X, strongly suggesting that the locus for the defect causing cystic fibrosis is not at 13q34.     

Preparation of vitamin K-dependent proteins,such as clotting factors II,VII, IX and X and clotting inhibitor protein C

A review is given of preparative methods for the isolation of the vitamin K-dependent clotting factors II, VII, IX, X and clotting inhibitor protein C, all derived from human plasma. Factor II, activated factor VII and activated protein C are also obtained from recombinant animal cells. The methods for their purification are described. The problem of difference in posttranslational modifications between plasma derived and recombinant protein is discussed with regard to therapeutic proteins.     

Regional localisation of the Friedreich ataxia locus to human chromosome 9q13----q21.1

We have previously assigned the Friedreich ataxia locus (FRDA) to chromosome 9; the current maximal lod score between FRDA and MCT112 (D9S15) is greater than 50 at a recombination fraction of theta = 0. The physical assignment of the locus defined by MCT112, and hence FRDA, has not been determined, although linkage analysis of MCT112 with other chromosome 9 markers inferred a location close to the centromere. We have used in situ hybridisation with MCT112, a corresponding cosmid MJ1, and DR47 (D9S5), coupled with mapping studies on hybrid cell panels, to define more precisely the location of the disease locus. The in situ location of all three probes is 9q13----q21.1, distal to the variable heterochromatin region. Physical assignment of FRDA will allow us to identify hybrid cell lines containing the mutated gene.     

Comparative mapping between human chromosome 3 and porcine chromosome 13.

Zoo-FISH and somatic cell hybrid panels have earlier demonstrated extended synteny conservation between human chromosome 3 (HSA3) and pig chromosome 13 (SSC13). In the present study, eight human genes viz., ADCY5, CASR, COL7A1, COL8A1, ITIH1, RHO, SIAT1 and XPC, spread along the length of HSA3, were chosen for expanding the comparative map between the two chromosomes. Using human and rat cDNAs, or human- and porcine-specific PCR products as probes, 8 porcine lambda clones were isolated. After subcloning and partial sequence determination, identity of the clones with regards to the specific genes was established. The eight type 1 markers thus obtained were biotin labeled and FISH mapped to pig metaphase spreads. All lambda clones localized to SSC13. In combination with the hitherto published mapping data of coding sequences on SSC13, a preliminary comparative status depicting the relative organization of this chromosome with respect to HSA3 was developed. The comparative map thus obtained bears significance in searching for candidate genes of economically important traits mapped to SSC13.     

Fine mapping of the diabetes-susceptibility locus, IDDM4, on chromosome 11q13.

Genomewide linkage studies of type 1 diabetes (or insulin-dependent diabetes mellitus [IDDM]) indicate that several unlinked susceptibility loci can explain the clustering of the disease in families. One such locus has been mapped to chromosome 11q13 (IDDM4). In the present report we have analyzed 707 affected sib pairs, obtaining a peak multipoint maximum LOD score (MLS) of 2.7 (lambda(s)=1.09) with linkage (MLS>=0.7) extending over a 15-cM region. The problem is, therefore, to fine map the locus to permit structural analysis of positional candidate genes. In a two-stage approach, we first scanned the 15-cM linked region for increased or decreased transmission, from heterozygous parents to affected siblings in 340 families, of the three most common alleles of each of 12 microsatellite loci. One of the 36 alleles showed decreased transmission (50% expected, 45.1% observed [P=.02, corrected P=.72]) at marker D11S1917. Analysis of an additional 1,702 families provided further support for negative transmission (48%) of D11S1917 allele 3 to affected offspring and positive transmission (55%) to unaffected siblings (test of heterogeneity P=3x10-4, corrected P=. 01]). A second polymorphic marker, H0570polyA, was isolated from a cosmid clone containing D11S1917, and genotyping of 2,042 families revealed strong linkage disequilibrium between the two markers (15 kb apart), with a specific haplotype, D11S1917*03-H0570polyA*02, showing decreased transmission (46.4%) to affected offspring and increased transmission (56.6%) to unaffected siblings (test of heterogeneity P=1.5x10-6, corrected P=4.3x10-4). These results not only provide sufficient justification for analysis of the gene content of the D11S1917 region for positional candidates but also show that, in the mapping of genes for common multifactorial diseases, analysis of both affected and unaffected siblings is of value and that both predisposing and nonpredisposing alleles should be anticipated.