Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q9NRP7 (fused)
 58,367 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mouse fibroblasts resistant to the drug rutamycin were isolated by selectively introducing BrdUrd into the mitochondrial genome of a line of mouse fibroblasts (clone 1 D) lacking a cytoplasmic thymidine kinase enzyme. The ATPase (ATP phosphohydrolase; EC 3.6.1.3) activity of mitochondria isolated from these cells was resistant to rutamycin. The rutamycin-resistant mutants were enucleated with cytochalasin B and fused with mouse A 9 cells resistant to 8-azaguanine and sensitive to rutamycin. Cytoplasmic hybrids, or cybrids, were selected as cells resistant to rutamycin and 8-azaguanine, and appeared at a high frequency. Other fusions between rutamycin-resistant nucleated cells and A 9 produced colonies at a much lower frequency. Finally, fusions between enucleated clone 1 D cells and A 9 cells produced no rutamycin-resistant colonies. These results indicate that rutamycin resistance is a cytoplasmically inherited characteristic in this cell line.
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PMID:Cytoplasmic inheritance of rutamycin resistance in mouse fibroblasts. 14 79

A procedure is described by which proteins can be rapidly and efficiently microinjected into large numbers of culture cells. Proteins were first introduced into mammalian red blood cells during hypotonic hemolysis, and the resealed red cells were subsequently fused to culture cells using Sendai virus. In seven experiments, thymidine kinase or 125I-BSA were transferred to culture cells during fusion. Although proteins were used in the present experiments, the microinjection procedure should work equally well for other macromolecules.
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PMID:Microinjection of thymidine kinase and bovine serum albumin into mammalian cells by fusion with red blood cells. 16 73

Although most mammalian cell lines can utilize either nicotinic acid or nicotinamide for the biosynthesis of nicotinamide adenine dinucleotide (NAD), thymidine kinase-deficient, mouse 3T3-4F cells are unable to utilize nicotinic acid. When 3T3-4E cells were fused with human D98/AH2 cells, autoradiography showed that the resultant heterokaryons synthesized NAD from nicotinic acid at rates comparable to the human parental cell. The rate of nicotinic acid utilization in heterokaryons remained unchanged over the four-day period of study following cell fusion. In contrast to the results observed with heterokaryons, nicotinic acid utilization was markedly reduced in hybrid cells. Of 100 hybrid clones examined at four or five days following cell fusion, 60 utilized nicotinic acid at rates less than one tenth that of the parental human cell. Similar results were observed in hybrid clones at nine or ten days following fusion. Uniformly high rates of NAD biosynthesis were observed in hybrid clones with nicotinamide as the precursor. This excludes the possibility that the reduction in nicotinic acid utilization in hybrid cells is due to a general metabolic dysfunction. The biochemical mechanism by which nicotinic acid utilization is markedly reduced has not been determined with certainty, however, several observations suggest genetic suppression.
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PMID:Autoradiographic studies of nicotinic acid utilization in human-mouse heterokaryons and inhibition of utilization in newly-formed hybrid cells. 17 18

GM54VA human cells transformed by simian virus 40 (SV40) were fused with peritoneal macrophages obtained from three different mouse strains. All 27 hybrid clones studied were positive for SV40 tumor antigen in 100% of their cells and contained human chromosome 17. Human chromosome 17 was the only human chromosome present in five of the hybrid clones. Fusion of GM54VA cells and either thymidine kinase (EC 2.7.1.75)-deficient mouse or Chinese hamster fibroblasts resulted in the growth in hypoxanthine-aminopterin-thymidine medium of hybrid clones positive and negative for SV40 tumor antigen. Counterselection of the hybrid clones positive for tumor antigen in medium containing 5-bromodeoxyuridine resulted in the growth of hybrid cells that were negative for tumor antigen. These experiments indicate that negative for tumor antigen. These experiments indicate that SV40 is integrated in only one of the two parental human chromosomes 17. Because the genome of SV40 has been assigned to human chromosome 7 in two other SV40-transformed human cell lines, at least two different integration sites for SV40 would seem to be present in human cells: one located in human chromosome 7 and the other located in human chromosome 17.
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PMID:Assignment of the integration site for simian virus 40 to chromosome 17 in GM54VA, a human cell line transformed by simian virus 40. 18 10

The mechanisms of normal cell differentiation in vivo may be related to some features of cellular aging in vitro in that both are considered to be under genetic control. Diploid rhesus choroidal melanocytes and purified peripheral lymphocytes were fused by means of inactivated Sendai virus with three long-term murine cell lines which lacked either hypoxanthine-guanine phosphoribosyl transferase or thymidine kinase. Cell hybrids were selected by their growth in medium containing hypoxanthine, aminopterin, thymidine, and glycine. G-banded chromosomes were analyzed and elements from both the rhesus and the established mouse cell lines were identified in all metaphases. Hybrids derived from choroid X mouse cells contained more than one chromosome set from the mouse, but those between lymphocytes and established cell lines had only one. However, in every combination continuously replicating hybrids were produced; most of them have undergone more than 40 subculture passages. Our results demonstrate not only that DNA synthesis can be re-initiated in postreplicative cells, but also that DNA continues to replicate in a manner consistent with the life span of the long-term cell line parent.
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PMID:Somatic cell hybrids derived from terminally differentiated rhesus cells and established mouse cell lines. 19 47

Mouse teratocarcinoma cells from the OTT6050 ascites tumor were established in tissue culture and selected for 5-bromodeoxyuridine (BrdUrd) resistance. The embryonal carcinoma cells grew without a feeder layer, remained deficient for thymidine kinase (EC 2.7.1.75), and differentiated like the original tumor into various tissues after subcutaneous injection into 129 mice. We fused the BrdUrd-resistant mouse teratocarcinoma cells with HT1080-6TG human diploid fibrosarcoma cells deficient in hypoxanthine phosphoribosyltransferase (EC 2.4.2.8) and selected for hybrid cells in hypoxanthine/aminopterin/thymidine medium. The resulting hybrid cells segregated human chromosomes quickly and retained one to three human chromosomes including chromosome 17 that carries the human genes for thymidine kinase and galactokinase (EC 2.7.1.6). Single hybrid cells from five independent clones containing human chromosome 17 were injected into mouse blastocysts bearing several genetic markers that affect the coat color phenotype and strain-specific enzyme variants in order to detect tissue differentiation derived from the injected cells. After the injection of single hybrid cells into a total of 103 experimental blastocysts that had been surgically transferred to pseudopregnant foster mothers, 49 mice were born and 2 of them clearly revealed coat mosaicism. In 2 of 17 mice thus far analyzed, the injected hybrid cells proved to be capable of participating substantially in development of seven different organs. However, human gene products have not yet been detected unequivocally in those tissues and weak human-specific galactokinase activity could be recovered only from two mosaic tissues. Our results demonstrate that, after in vitro culture and selection, at least some of the human-mouse hybrid cells still retain their in vivo potential to differentiate and become functionally integrated in the living organism. It now seems feasible to cycle mouse teratocarcinoma cells carrying human genetic material through mice via blastocyst injection to study human gene expression during differentiation.
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PMID:Chimeric mice derived from human-mouse hybrid cells. 20 75

To investigate chromosomal site(s) of integration of the herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) gene in biochemically transformed [HeLa(BU25)/KOS 8-1] cells, these human cells which had been transformed by ultraviolet light-irradiated HSV-1 were fused with TK-negative mouse LM(TK-) cells, human-mouse somatic cell hybrid clones (LH81 clones 1-20) were isolated by HATG-ouabain selection and their chromosomes and isozymes were analyzed. Electrophoretic and serological analyses showed that all 20 clones expressed type-specific HSV-1 TK. Isozyme analyses on 29 gene-enzyme systems representing 22 human chromosomes revealed that all of the HSV-1 TK-positive clones expressed human aminoacylase-1 (ACY-1) and esterase D (ESD), which have been mapped to human chromosomes 3 and 13, respectively. Other human isozymes were detected in only one to four clones or in none of the clones. Chromosome analyses showed that: (1) the hybrid clones retained only a few human chromosomes; (2) a marker chromosome, designated M7, consisting of a chromosome 17 translocated to the short arm of chromosome 3, occurred in 36 out of the 41 metaphases examined of LH81-4 clones 1 to 4 and in 31 out of the 33 metaphases examined of LH81-12 clone 10; (3) a modified M7 chromosome, (M7/m), in which the distal 2/3 of the long arm of M7 was translocated to a small acrocentric mouse chromosome, was the only human chromosome found in metaphases of LH81-13 clone 17; and (4) an intact human chromosome 13 was not present in LH81-12 clone 10 or LH81-13 clone 17 cells. Counterselection with BrdUrd resulted in the isolation of subclones lacking HSV-1 TK, human ACY-1 and ESD, and the human marker M7 chromosomes. The experiments indicate that the HSV-1 TK gene is probably associated in HeLa (BU25)/KOS 8-1 cells with marker chromosome M7, but the possibility is not excluded that the segment of human chromosome 13 which codes for ESD is involved.
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PMID:Isozyme studies on the association of the herpes simplex virus type 1 thymidine kinase gene with human chromosomes in somatic cell hybrids. 23 92

I have reviewed the current status of microinjection based on fusion of red blood cells and tissue culture cells. Macromolecules are introduced into red blood cells during hypotonic hemolysis, and the resealed red cells are then fused to tissue culture cells with Sendai virus. The procedure has been used to inject ferritin, thymidine kinase, bovine serum albumin, and transfer RNA molecules into large numbers of tissue culture cells. Physiologically significant amounts of various macromolecules can be transferred, and preliminary studies show that [125I]bovine serum albumin and transfer RNA are stable within recipient culture cells. Tissue culture cells remain viable following microinjection. Red cell-mediated microinjection should facilitate the study of various processes, such as macromolecular turnover and genetic regulation, that are not easily studied with conventional biochemical techniques.
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PMID:Red cell-mediated microinjection. 37 19

Incorporation of [3H]thymidine was observed in both parental nuclei in heterokaryons resulting from the fusion of post-mitotic, "senescent" human diploid cells and a thymidine kinase-deficient murine cell line (3T3der-4E). The senescent nuclei displayed a sudden increase of activity approximately 4--6 hours after fusion. A moderate increase of thymidine incorporation was observed in 3T3der-4E cells cocultivated with but not fused to senescent cells, consistent with metabolic cooperation. Chromosome preparations of cultures fixed approximately 24 hr after fusion revealed the presence of hybrid metaphase cells containing almost the entire human complement. All of the identifiable human chromosomes were bi-armed, suggesting that the senescent nuclei were stimulated to reinitiate replicative DNA synthesis rather than repair synthesis in these heterokaryons.
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PMID:Complementation in heterokaryons derived from a thymidine kinase deficient cell line and senescent human diploid cells. 42 67

Interspecific human-mouse and Chinese hamster-mouse hybrids were isolated from polyethylene glycol fused cells by a new half-selection technique employing a structurally modified polyene macrolide antibiotic, amphotericin B methyl ester (AME), and HAT media. Unfused parental cells were killed as a result of innate sensitivity to AME or their genetic deficiency, absence of thymidine kinase (TK-) or hypoxanthine guanine-phosphoribosyl transferase (HGPRT-). In contrast, hybrid colonies were isolated after two to three weeks growth in three or four changes of HAT-AME media and subsequent growth in HAT media alone. The ability of hybrid cells to proliferate using this selective protocol indicates that genetic complementation resulted, and polyene antibiotic resistance was expressed as a dominant phenotypic property in the hybrids. Hybrid selection was dependent on: (1) the number of cells of each parental cell type co-cultivated; (2) the level of polyene antibiotic administered; and (3) the time interval before selection was initiated. The half-selection technique described in this report is simple to use, very effective in eliminating unfused parental cells and increases the potential types of hybrids which can be formed. Only one parental cell type need contain a biochemical defect, whereas the second parental type can be genetically normal.
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PMID:Selecting interspecific human-mouse and Chinese hamster-mouse hybrids using a new half-selection technique with a polyene antibiotic. 54 Dec 68


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