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.     

A human cell-surface antigen defined by a monoclonal antibody and controlled by a gene on chromosome 12

A murine monoclonal antibody 602-29, subclass IgG1, that recognizes an antigenic determinant expressed by most human cells is described. Immunoprecipitation and sodium dodecyl sulfate-gel electrophoresis analysis indicate that the antigenic determinant is carried by a protein with an apparent molecular weight of 21,000. The antigen is expressed by human-mouse somatic cell hybrids, and analysis of segregants that have lost human chromosomes indicates that the gene controlling expression of the 602-29 antigen is on chromosome 12.     

Assignment of the gene locus for human phosphoglucomutase 3 to chromosome 6q12-qter

Segregation of human PGM3 has been analyzed in somatic cell hybrids between mouse A9 cells and human fibroblasts carrying a reciprocal translocation: 46,XX, t(6;7) (q12;p14). The enzyme marker segregates with the 7p+ chromosome indicating that the PGM3 gene is located on 6q12 greater than qter.     

Chromosomal localization of two human zinc finger protein genes, ZNF24 (KOX17) and ZNF29 (KOX26), to 18q12 and 17p13-p12, respectively

Two members of the zinc finger Krüppel family, ZNF24 (KOX17) and ZNF29 (KOX26), have been localized by somatic cell hybrid analysis and in situ chromosomal hybridization to human chromosomes 18q12 and 17p13-p12, respectively. The mapping of ZNF29 together with the previously reported localization of ZFP3 suggests that a zinc finger gene complex is located on human chromosome 17p. ZNF29 maps centromeric to the human p53 tumor antigen gene (TP53). In the analogous murine position, the two mouse zinc finger genes Zfp2 and Zfp3 have recently been assigned to the distal region of mouse chromosome 11, the murine homolog of human chromosome 17. Both human zinc finger genes ZNF24 and ZNF29 are in chromosomal regions that have been noted to be deleted in neoplasms of the lung and of the central nervous system at chromosome 17p and in colorectal neoplasia at chromosomes 17p and 18q.     

In situ hybridization mapping and restriction fragment length polymorphism analysis of the porcine albumin (ALB) and transferrin (TF) genes

Summary
In situ hybridization analyses were conducted on porcine metaphase chromosomes using porcine liver albumin (ALB) and transferrin (TF) cDNA probes. The ALB gene was assigned to the q12 band of chromosome 8 and the TF gene to the q31 band of chromosome 13. For the latter, a statistically significant secondary peak was observed on the 6p15 band. However, the TF probe predominantly hybridized to the 13q31 band, indicating that this band is the most likely site of the TF gene. Since the TF gene belongs to linkage group V, this linkage group can now be assigned to chromosome 13. The TF and ALB probes were also used for restriction fragment length polymorphism (RFLP) analysis. A screening of 10 unrelated animals revealed Tag I RFLPs for both ALB and TF. Family studies indicated that the ALB and TF polymorphisms were controlled by three and two alleles, respectively.     

Chromosome localization of human ARH genes, a ras-related gene family

The human ARH genes (previously called RHO) share several properties with the ras gene family. Three members of the ARH family, the H6, H9, and H12 genes, have been localized to human chromosomes 2, 5, and 3, respectively. Analysis of DNAs from a rodent-human somatic cell hybrid panel demonstrates linkage of H6 to chromosome region 2p12----2pter and H9 to region 5q33----5qter. In situ chromosome hybridization also showed that the primary site for H9 is in the 5q31----qter region. The H12 gene was some-what difficult to localize using rodent-human hybrids because the probe detects a family of rodent genes as homologous to the human probe as in the human cognate gene. However, chromosome in situ hybridization revealed grains clustered in region 3p14----3p22 with a significant peak in band 3p21. We conclude that H6 is in 2p12----pter, H9 in 5q31----5qter, and H12 in 3p21.     

Monoclonal antibodies specific for human chromosome 5 obtained with a monochromosomal hybrid can be used to sort out cells containing the chromosome with a FACS

Using a human-mouse monochromosomal hybrid, BG15-6, that contains an intact human chromosome 5, we isolated four monoclonal antibodies, 2A10, 3H9, 5G9, and 6G12, as chromosome marker antibodies recognizing cell surface antigens specific for human chromosome 5. The binding patterns of these antibodies to BG15 subclones containing fragments of human chromosome 5 indicated that 2A10, 3H9, and 6G12 recognized the antigens produced by genes located on 5pterq22, and that 5G9 recognized the antigen produced by a gene located on 5q23. Cells containing human chromosome 5 were very effectively sorted in a fluorescence-activated cell sorter (FACS) using monoclonal antibody 6G12. This method for sorting cells containing human chromosome 5 or an appropriate fragment of this chromosome from among human-rodent hybrid cells should be very useful in studies on gene expression, gene cloning and gene mapping.by M. Trendelenburg     

Linkage studies do not confirm the cytogenetic location of incontinentia pigmenti on Xp11

Summary Linkage studies have been performed in 5 incontinentia pigmenti (IP) families totaling 29 potentially informative meioses. Ten probes of the Xp arm were used, six of them were precisely localized on the X chromosome, using hamster x human somatic cell hybrids containing a broken X chromosome derived from an incontinentia pigmenti patient carrying an X;9 translocation [46,XX,t(X;9)(p11.21;q34)]. The following order for probes is proposed: pter-(DXS7, DXS146, DXS255)-IP1-(DXS14, DXS90)-DXS106-qter. The negative lod scores obtained exclude the possibility that in the families studied, the gene for IP is located in Xp11 or in the major part of the Xp arm.     

Identification of a cell-surface antigen produced by a gene on human chromosome 3 (cen-q22) and not expressed by Rhnull cells.

A monoclonal antibody, 1D8, which recognizes a cell-surface antigen expressed by human chromosome 3 in Chinese hamster-human somatic-cell hybrids, has been produced. Testing of hybrids containing various deletions of chromosome 3 determines that the gene encoding the antigen is regionally localized to 3q (cen-22). This regional mapping is distinct from that elsewhere reported for two other cell-surface antigens assigned to chromosome 3--namely, the human transferrin receptor and the p97 melanoma-associated antigen. In addition, biochemical characterization is different from that elsewhere reported for other chromosome 3-encoded cell-surface antigens. When tested against a panel of rare-phenotype red blood cells, the only cells that failed to react were those of the Rhnull phenotype. The antibody reacts only weakly with homozygous -D- and fetal red cells, in contrast with a previously described antibody, R6A, which does not react with Rhnull cells. Furthermore, R6A does not recognize a cell-surface antigen expressed by chromosome 3 in Chinese hamster-human somatic-cell hybrids. Thus, the monoclonal antibody 1D8 recognizes a previously undescribed cell-surface antigen encoded by human chromosome 3 and not expressed on Rhnull cells. The gene on chromosome 3 regulating expression of this antigen may be that defective in Rhnull disease or may require the normal allele at an unlinked Rhnull locus for expression. Linkage studies will be required to further elucidate this matter.     

Localisation régionale des gènes humains LDHB,TPI, ENO2, PepB,PGK, αGALA,G6PD et HGPRT par l'hybridation cellulaire interspécifique

Summary Twenty-two independent man-mouse (A9, HGPRT^-) and manhamster (CH, HGPRT)-hybrids using male human with balanced reciprocal translocation XY t(X;12)(q2,3;q1,2) were analysed for human genes localized on chromosome 12 (LDH_B, PepB, ENO₂, TPI), on chromosome X (PGK, GAL_A, G6PD) and for human chromosomes in relation with the balanced reciprocal translocation (chromosome 12, 12q^-, Xq⁺). The human chromosome 12q^- was not analysed because of its morphology is similar to some rodent chromosomes.The following results were obtained:1.Man-mouse hybrids (12 independent hybrids):The chromosome 12 is absent. A positive correlation is observed between Xq⁺, PGK, and PepB. Four hybrids are Xq⁺+PGK+PepB+ and eight hybrids are Xq⁺-PGK-PepB-This result indicate that the genes for human PGK, PepB are on the chromosome Xq+. 2.Man-hamster hybrids (10 independent hybrids):A positive correlation is observed between Xq⁺, PGK, GAL_A. Two hybrids are Xq⁺+PGK+GAL_A+ and eight hybrids are Xq⁺-PGK-GAL_A-.A positive correlation is observed between Xq⁺, PGK, GAL_A, PepB with the seven hybrids missing the normal human chromosome 12. One hybrid is Xq⁺+PGK+GAL_A+PepB+ and six hybrids are Xq⁺-PGK-GAL_A-PepB-. These results indicate that the genes for human PGK, GAL_A, PepB are on the chromosome Xq⁺. 3.Man-mouse and man-hamster hybrids:The highest percentage of presence in the hybrids is observed for the following markers: G6PD (100%), LDH_B, TPI, ENO₂ (90%). This is explained by the fact that these hybrids selected in HAT medium, had to retain a segment of X (Xq⁺ or 12q^-) bearing the human HGPRT gene. The different results indicated that the segment of X retained in these hybrids must be on the 12q^- and not on the Xq⁺, for Xq⁺ is rarely present in man rodent hybrids.The different results implicate finally the following localisations:LDH_B, TPI, ENO₂ on 12 q12 12pter,PepB on 12 q12 12qter;PGK, GAL_A on X q23 Xpter;HGPRT, G6PD on X q23 Xqter.     

Gene for human CD59 (likely Ly-6 homologue) is located on the short arm of chromosome 11

The CD59 (MEM-43) antigen, which probably is a human homologue of mouse Ly-6 antigens, is a broadly expressedM _r 18000–25000 human leucocyte surface glycoprotein recognized by monoclonal antibody MEM-43. Ten mouse-human T-lymphocyte hybrids, carrying all mouse chromosomes and a limited number of human chromosomes, were analyzed for expression of CD59 by indirect immunofluorescence and immunoblotting with MEM-43 antibody. Karyotypic analysis of the tested clones showed that the presence of human chromosome 11 correlated with the expression of CD59 in all clones tested. Three other human chromosome 11-encoded antigens, 4F2 (Trop-4), Leu 7 (HNK-1, CD57), and lymphocyte homing receptor, were expressed concordantly with CD59. A more exact localization of the gene for CD59 was obtained by the study of Chinese hamster-human cell hybrids containing short or long arm deletions of human chromosome 11. CD59 segregated with hybrids containing part of the short arm of human chromosome 11, but not with the hybrids containing the long arm. Based on these studies we assign the gene for CD59 to regionP14–p13 of the short arm of chromosome 11.     

Regional mapping of the gene for human UDPGal 4-epimerase on chromosome 1 in mouse-human hybrids

Somatic cell hybrids between mouse and human cells containing two different reciprocal translocations involving human chromosome 1, 46,X,t(1;X)(q12;q26) and 47,XX,+21,t(1;17)(p32;p13), were studied for the expression of human uridine diphosphate galactose 4-epimerase (UDPGal 4-epimerase, E.C. 5.1.3.2) by starch-gel electrophoresis. Analysis of the hybrid clones for the expression of the enzyme and the presence of the translocation chromosome 1 has permitted the assignment of the gene for human UDPGal 4-epimerase to the pter yields p32 region of chromosome 1.     

Two human genes encoding zinc finger proteins,ZNF12 (KOX 3) and ZNF 26 (KOX 20), map to chromosomes 7p22-p21 and 12q24.33, respectively

Summary Two members of the human zinc finger Krüppel family, ZNF 12 (KOX 3) and ZNF 26 (KOX 20), have been localized by somatic cell hybrid analysis and in situ chromosomal hybridization. The presence of individual human zinc finger genes in mouse-human hybrid DNAs was correlated with the presence of specific human chromosomes or regions of chromosomes in the corresponding cell hybrids. Analysis of such mouse-human hybrid DNAs allowed the assignment of the ZNF 12 (KOX 3) gene to chromosome region 7p. The ZNF 26 (KOX 20) gene segregated with chromosome region 12q13-qter. The zinc finger genes ZNF 12 (KOX 3) and ZNF 26 (KOX 20) were localized by in situ chromosomal hybridization to human chromosome regions 7p22-21 and 12q24.33, respectively. These genes and the previously mapped ZNF 24 (KOX 17) and ZNF 29 (KOX 26) genes, are found near fragile sites.     

Chromosome 1 localization of the gene for CD34, a surface antigen of human stem cells.

CD34 is a surface antigen expressed on normal human hematopoietic stem cells, as well as on the blast cells of many patients with both lymphocytic and myelocytic leukemias. By Southern blot analysis of DNA from a panel of human x mouse somatic cell hybrids using a CD34 cDNA probe, we demonstrate that the gene for CD34 is located on human chromosome 1 in the 1q12----qter region.     

Separation of the immune response genes for LDH-B and MOPC-173. I. Description of an immune response defect in B10.BASR1

By using the intra-I-region recombinant mouse strain, B10.BASR1 (H-2as4), the immune response (Ir) genes for LDH-B and MOPC-173 were genetically and serologically separated, as assayed by T cell proliferation. Previous work demonstrated that the H-2s and H-2b strains respond to LDH-B and MOPC-173, whereas the H-2a and H-2k strains failed to respond due to haplotype-specific suppression of I-Ak-activated T helper cells by I-Ek-activated T suppressor cells. In the experiments reported here, B10.BASR1 mice, which lack I-Ek expression, mounted a significant T cell proliferative response to MOPC-173 but not to LDH-B. Separation of the Ia determinants used in restricting these two antigen responses was further confirmed when pretreatment of B10.S(9R) (A alpha sA beta sE beta sJk) macrophages with A.TL anti-B10.HTT (anti-A beta sE beta sJs) serum absorbed with B10.BASR1 spleen cells blocked the LDH-B response but not the MOPC-173 response. Unabsorbed serum blocked both antigen responses. The primary immunogenic determinant recognized by LDH-B or MOPC-173 immune T cells was not present on both antigens, as MOPC-173-primed T cells and LDH-B-primed T cells responded only to the priming antigen. Lastly, by using the A beta mutant strain, B6CH-2bm12, it was shown that the Ir gene and Ia determinants affected by this mutation had no effect on the LDH-B and MOPC-173 proliferative responses. These results suggest the possibility of an intragenic recombinatorial event in either the A alpha or A beta chain resulting in the separation of these two immune response gene functions.     

The use of cell surface antigens to characterize and select for fragments of human chromosomes retained by interspecies hybrids

We have used a mouse cell transformant generated by human chromosome-mediated gene transfer (CMGT) to explore the use of cell surface antigens in the identification of fragments of human chromosomes retained by somatic cell hybrids. The transformed line, 21-30b, contained an intact rear-ranged human chromosome, and could be shown by isozyme analysis to contain genetic material from chromosomes 9 and X. By using the transformant as an immunogen in mice, it was also possible to produce antiserum to human-specific surface antigens. Using genetically characterized human X rodent hybrid lines, the genes controlling expression of these antigens could be localized to 11per----11p13, segregating concordantly with surface antigen S3. These conclusions were possible despite the fact that the presence of chromosome 11 in the transformant was not detectable by the presence of chromosome specific isozyme LDH-A or surface antigens W6/34 and 4F2. Finally, the fluorescence-activated cell sorter (FACS) was used to fractionate the transformant cells into antigen positive and negative subpopulations. This resulted in the isolation and characterization of four additional chromosome rearrangements involving interspecies chromosome translocations. This work demonstrates the value of chromosome-specific surface antigens and the FACS in the evaluation of human chromosome fragments retained by interspecies hybrids.     

Regional localization of the gene for thyroid peroxidase to human chromosome 2pter----p12

A 2.0-kb thyroid peroxidase cDNA of human origin was used as probe for Southern blot hybridization of genomic DNA from human somatic cells and human-rodent somatic cell hybrids. The results showed that the gene coding for human thyroid peroxidase is located on chromosome. 2. Further analysis of hybrids derived from Burkitt lymphoma cells carrying a (2;8)(p12;q24) translocation revealed that the gene maps to the region 2pter----p12.     

Localisation régionale des gènes humains IDHS,MDHS, PGK, α-GAL,G6PD par l'hybridation cellulaire interspécifique

Summary 22 independent man-hamster (HGPRT^–) hybrids using male human cells with balanced reciprocal translocation t(X;2)(p22;q32) were analysed for human genes localized on chromosome 2 (IDH_S, MDH_S), on chromosome X (PGK, GAL, G6PD) and for the different chromosomes in relation with the balanced reciprocal translocation (chr.2, chr.2q^–, chr.Xp⁺).The following results were obtained:The chromosomes 2 and 2q^– are absent in the 22 hybrids.In 9 hybrids, the absence of MDH_S in spite of the presence of the chromosome Xp⁺ indicates that the gene for MDH_S is not localized on this chromosome (or that the gene for MDH_S is not on the segment 2q32-2qter translocated on X).In 14 hybrids, the three markers of X (PGK, GAL, G6PD) and IDH_S are expressed in the presence of the chromosome Xp⁺. This result indicates that the genes for these markers are on Xp⁺ or that the genes PGK, GAL, G6PD are on X without the Xp22-Xter segment, translocated on the chr.2, and that the gene for IDH_S is on the 2q32-2qter segment translocated on X.In 8 hybrids, in the absence of the intact chromosome Xp⁺, the higher percentage of the presence of G6PD (7 hybrids) and the lower percentage of the presence of IDH_S (3 hybrids) are explained by the fact that these hybrids selected in HAT medium had to retain a segment of Xp⁺ bearing the human gene HGPRT. G6PD appeared very close to HGPRT and IDH_S very distant from HGPRT.The study of the different correlations between the presence and the absence of these four markers on Xp⁺ in the different hybrids indicates the following order on the chromosome Xp⁺ from p to q: IDH_s — PGK — GAL — G6PD.

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Groupe INSERM: Directeur J. Frézal

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Groupe CNRS, ER, 149: Directeur J. de Grouchy     

The human phosphoglycerate kinase multigene family. HLA-associated sequences and an X-linked locus containing a processed pseudogene and its functional counterpart

Several phosphoglycerate kinase genes were previously detected in the human genome by blot hybridization with a phosphoglycerate kinase cDNA probe. Using subcloned fragments of the cDNA we estimate the presence of four independent phosphoglycerate kinase genes. These genes have been mapped to both the human X chromosome (band q13) and chromosome 6 (p12-21.1) using a panel of human-rodent somatic cell hybrids and by chromosomal in situ hybridization. The genomic distribution of phosphoglycerate kinase sequences is conserved in man and mouse, not only for the X chromosome, but also for linkage to the respective major histocompatibility complexes. Molecular cloning of X-linked phosphoglycerate kinase sequences led to the identification of a novel intronless phosphoglycerate kinase pseudogene which is localized proximal to the active gene on the X chromosome.