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Biomedical subjects

D Bootsma

Publications and source records attributed to D Bootsma.

At least 91 records · Page 5Linked to original sources
PubMed1987

Biological and biochemical consequences of the human ERCC-1 repair gene after transfection into a repair-deficient CHO cell line.

The consequences of the presence of the human gene ERCC1 in repair-deficient 43-3B cells were examined. The gene restores the sensitivity of this mutant not only to UV but also to 4NQO, N-Ac-AAF and alkylating agents to the normal level. Also, the frequency of mutation induction by UV at the Na+/K+-ATPase locus returns to the level of CHO wild-type cells. Additionally, the rate of cyclobutane pyrimidine dimer removal approaches that in wild-type CHO cells. The results obtained indicate that the human gene ERCC-1 restores the impaired functions in 43-3B, and that the gene is probably functionally homologous to the defective one in the 43-3B cell line. Some evidence was found for a difference between the human gene product and its rodent counterpart, as the restoration of normal sensitivity to 4NQO, ENU and N-Ac-AAF was complete whereas it was not for UV.

4-Nitroquinoline-1-oxide↗
PubMed1986

Molecular characterization of the human excision repair gene ERCC-1: cDNA cloning and amino acid homology with the yeast DNA repair gene RAD10.

The human excision repair gene ERCC-1 was cloned after DNA mediated gene transfer to the CHO mutant 43-3B, which is sensitive to ultraviolet light and mitomycin-C. We describe the cloning and sequence analysis of the ERCC-1 cDNA and partial characterization of the gene. ERCC-1 has a size of 15 kb and is located on human chromosome 19. The ERCC-1 precursor RNA is subject to alternative splicing of an internal 72 bp coding exon. Only the cDNA of the larger 1.1 kb transcript, encoding a protein of 297 amino acids, was able to confer resistance to ultraviolet light and mitomycin-C on 43-3B cells. Significant amino acid sequence homology was found between the ERCC-1 gene product and the yeast excision repair protein RAD10. The most homologous region displayed structural homology with DNA binding domains of various polypeptides.

Amino Acid Sequence↗
PubMed1986

Unscheduled DNA synthesis in xeroderma pigmentosum cells after microinjection of yeast photoreactivating enzyme.

Photoreactivating enzyme (PRE) from yeast causes a light-dependent reduction of UV-induced unscheduled DNA synthesis (UDS) when injected into the cytoplasm of repair-proficient human fibroblasts (Zwetsloot et al., 1985). This result indicates that the exogenous PRE monomerizes UV-induced dimers in these cells competing with the endogenous excision repair. In this paper we present the results of the injection of yeast PRE on (residual) UDS in fibroblasts from different excision-deficient XP-strains representing complementation groups A, C, D, E, F, H and I (all displaying more than 10% of the UDS of wild-type cells) and in fibroblasts from two excision-proficient XP-variant strains. In fibroblasts belonging to complementation groups C, F and I and in fibroblasts from the XP-variant strains UDS was significantly reduced, indicating that pyrimidine dimers in these cells are accessible to and can be monomerized by the injected yeast PRE. The UDS reduction in the XP-variant strains is comparable with the effect in wild-type cells. In cells from complementation groups C, F and I the reduction is less than in wild-type and XP-variant cells. Fibroblasts belonging to groups A, D, E and H did not show any reduction in UDS level after PRE injection and illumination with photoreactivating light. These results give evidence that the genetic repair defect in some XP-strains is probably due to an altered accessibility of the UV-damaged sites.

DNA Repair↗
PubMed1985

Interspecies complementation analysis of xeroderma pigmentosum and UV-sensitive Chinese hamster cells.

Complementation analysis was performed 24 h after fusion of UV-sensitive CHO cells (CHO 12 RO) with XP cells of complementation groups A, B, C, D, F and G. The parental cells are characterized by low levels of unscheduled DNA synthesis (UDS). In all combinations, the UDS levels observed in heterokaryons were higher than those in parental mutant cells, clearly indicating cooperation of human and Chinese hamster repair functions. In heterokaryons of CHO 12 RO with XP-A and XP-C cells, the UDS values reached about the normal human level, whereas in heterokaryons with XP-B, XP-D and XP-F, UDS was restored at a level approaching that in wild-type CHO cells. The results obtained after fusion of CHO cells with two representative cell strains from the XP-G group, XP 2 BI and XP 3 BR, were inconsistent. Fusion with XP 3 BR cells yielded UDS levels ranging from wild-type Chinese hamster to normal human, whereas fusion with XP 2 BI cells resulted in a slight increase in UDS which even after 48 h remained below the level found in wild-type CHO cells. The occurrence of complementation in these interspecies heterokaryons indicates that the genetic defect in the CHO 12 RO cells is different from the defects in the XP complementation groups tested.

Animals↗
PubMed1985

A ninth complementation group in xeroderma pigmentosum, XP I.

A new complementation group of excision-deficient xeroderma pigmentosum (XP) is described in 2 patients living in the F.R.G. Dermatological, ophthalmological and neurological symptoms of XP are presented together with DNA repair characteristics such as unscheduled DNA synthesis, colony-forming ability and alkaline elution studied in cultured fibroblasts. The results are compared to normal controls.

Adolescent↗
PubMed1985

Microinjected photoreactivating enzymes from Anacystis and Saccharomyces monomerize dimers in chromatin of human cells.

Photoreactivating enzymes (PRE) from the yeast Saccharomyces cerevisiae and the cyanobacterium Anacystis nidulans have been injected into the cytoplasm of repair-proficient human fibroblasts in culture. After administration of photoreactivation light, PRE-injected cells displayed a significantly lower level of UV-induced unscheduled DNA synthesis (UDS) than non-injected cells. This indicates that monomerization of the UV-induced pyrimidine dimers in the mammalian chromatin had occurred as a result of photoreactivation by the injected PRE at the expense of repair by the endogenous excision pathway. Purified PRE from yeast is able to reduce UDS to 20-25% of the UDS found in non-injected cells, whereas the in vitro more active PRE from A. nidulans gives a reduction to only 70%. This suggests that the eukaryotic enzyme is more efficient in the removal of pyrimidine dimers from mammalian chromatin than its equivalent purified from the prokaryote A. nidulans.

Cells, Cultured↗
PubMed1985

Translocation of oncogene c-sis from chromosome 22 to chromosome 11 in a Ewing sarcoma-derived cell line.

Somatic cell hybrids, obtained after fusion of translocation (11;22)-positive Ewing sarcoma cells and Chinese hamster fibroblasts, were assayed for the presence of immunoglobulin C lambda, Philadelphia chromosome breakpoint cluster region, and c-sis oncogene sequences. It was found that c-sis was translocated from chromosome 22 to chromosome 11 in the Ewing sarcoma cells used, indicating that the breakpoint must be proximal to this locus. Moreover, we found that the chromosome 22-linked C lambda and breakpoint cluster region sequences are not translocated. This result confirms an earlier cytogenetic observation that the Ewing sarcoma-associated breakpoint in chromosome 22 is distal to those observed in translocation (8;22)-positive Burkitt lymphoma and in Philadelphia chromosome-positive chronic myeloid leukemia.

Aconitate Hydratase↗
PubMed1984

Regional mapping of the human immunoglobulin lambda light chain to the Philadelphia chromosome in chronic myeloid leukaemia.

The lambda light chain immunoglobulin constant region (C lambda) locus was mapped on human chromosome 22. A DNA probe containing part of the C lambda locus was isolated from a human chromosome 22 genomic library, and a series of rodent X human somatic cell hybrids (each of which contained different translocated parts of chromosome 22) were constructed and characterized. The hybridization of the C lambda probe to DNA from these cell hybrids was then studied by Southern blot analysis. The results demonstrates that the C lambda locus is situated very close to the translocation breakpoint on human chromosome 22 which is characteristic of chronic myeloid leukaemia, and at least part if not at all of the locus is situated on the Philadelphia chromosome.

Chromosome Mapping↗
PubMed1984

Is the chromosomal region 9q34 always involved in variants of the Ph1 translocation?

Six variants of the Ph1 translocation are described. The clinical diagnoses were chronic myeloid leukemia (CML) in 5 cases (patients 1-5) and acute lymphocytic leukemia (ALL) in patient 6. Three Ph1 variants were clear complex translocations, involving chromosomes #9, #22, and a third chromosome, i.e., #16, #11, or #14. The other three Ph1 variants appeared as "simple" translocations between chromosome #22 and chromosome #19, #4, or #12 when G- or Q-banding were used. When studied with high resolution R-banding, a small deletion of the terminal part of one chromosome #9 was visible, strongly suggesting that these variants were also complex translocations, i.e., t(9;19;22)(q34;p13;q11),t(4;9;22) (p16;q34;q11), and t(9;12;22)(q34;p13;q11). In the latter two cases, using in situ hybridization techniques, we demonstrated the presence of c-abl sequences on the Ph1 chromosome. This proved the involvement of 9q34 in these two variants. Our proposal is that most, and probably all, variants of Ph1 are complex translocations involving part of 9q34 and that the conjunction of a specific region of 22q11 with a specific segment of 9q34 (carrying the c-abl protooncogene) is essential for the development of Ph1 + CML.

Chromosomes, Human, 21-22 and Y↗
PubMed1984

Werner's syndrome: proliferation in vitro of clones of cells bearing chromosome translocations.

Each of several cultures of Werner's syndrome (WS) fibroblasts and lymphoblasts examined was found to be composed of one or several clones of cells with mutated chromosome complements. Two "sister" fibroblasts cell lines (FCLs) that were derived from a mixture of explants cut from the same WS skin biopsy were found to have completely different rearranged chromosome complements. Daily observation of the skin explants from which these two sister FCLs were derived revealed not only that no more than a few fibroblasts ever migrated from a given explant but also that fibroblasts migrated from only a few of the explants. Two of three lymphoblastoid cell lines (LCLs), each probably developed as an independent clone from a different cell from the same WS blood sample, were mosaic, comprised of cells having both normal and rearranged chromosome complements. The third LCL studied, although nonmosaic, had a rearranged chromosome complement, but one that was completely different from those in the other two lines. Based on the observations described, hypotheses have been formulated to explain both the preponderance in long-term WS cultures of clones with mutated chromosome complements and the abbreviated lifespan characteristic of WS fibroblast cultures.

Adult↗
PubMed1983

c-sis is translocated from chromosome 22 to chromosome 9 in chronic myelocytic leukemia.

By analysis of a series of somatic cell hybrids derived by fusion of either mouse or Chinese hamster cells with leukocytes from different chronic myelocytic leukemia (CML) patients or from normal donors, we have localized the human oncogene, c-sis, on the q11 to qter segment of chromosome 22 and demonstrated its translocation from chromosome 22 to chromosome 9 (q34) in CML.

Animals↗
PubMed1983

Expression of human myeloid-associated surface antigens in human-mouse myeloid cell hybrids.

Hybrid cell lines were obtained after fusion of mouse myeloid cells (WEHI-TG) with leukocytes from two patients with chronic myeloid leukemia. A third fusion was carried out with leukocytes from a patient with acute lymphocytic leukemia. All three patients carried the Philadelphia chromosome (Ph1) in the leukemia cell population. Cytochemical analysis confirmed the myelo-monocytic nature of the hybrid cell lines. The presence of Ph1 translocation products could be established in most hybrids derived from the two chronic myeloid leukemic patients, which confirms that indeed human myeloid cells were fused. Several of these hybrid lines showed reactivity with monoclonal antibodies known to be specific for human myeloid cells, whereas interlineage Chinese hamster fibroblast-human chronic myeloid leukemia hybrids failed to react with these antibodies. Five independently obtained monoclonal antibodies--MI/NI, UJ-308, VIM-D5, FMC-10, and B4.3--showed very similar reactivity patterns when tested on the hybrid clones. This result substantiates the evidence obtained from other studies, that these five antibodies are directed against the same myeloid-associated antigen. The gene(s) for expression of the latter antigen could be assigned to human chromosome 11.

Animals↗
PubMed1982

A cellular oncogene is translocated to the Philadelphia chromosome in chronic myelocytic leukaemia.

The transforming genes of oncogenic retroviruses are homologous to a group of evolutionary conserved cellular onc genes. The human cellular homologue (c-abl) of the transforming sequence of Abelson murine leukaemia virus (A-MuL V) was recently shown to be located on chromosome 9. The long arm of this chromosome is involved in a specific translocation with chromosome 22, the Philadelphia translocation (Ph1), t(9; 22) (q34, q11), which occurs in patients with chronic myelocytic leukaemia (CML)3-5. Here we investigate whether the c-abl gene is included in this translocation. Using c-abl and v-abl hybridization probes on blots of somatic cell hybrids, positive hybridization is found when the 22q- (the Philadelphia chromosome), and not the 9q+ derivative of the translocation, is present in the cell hybrids. From this we conclude that in CML, c-abl sequences are translocated from chromosome 9 to chromosome 22q-. This finding is a direct demonstration of a reciprocal exchange between the two chromosomes and suggests a role for the c-abl gene in the generation of CML.

Chromosomes, Human, 21-22 and Y↗