RESUMO
Autosomal dominant Emery-Dreifuss muscular dystrophy (EDMD) is characterized by muscle wasting and is caused by mutations in the LMNA gene encoding A-type lamins. Overexpression of the EDMD lamin A R453W mutation in C2C12 myoblasts impairs myogenic differentiation. We show here the influence of stable expression of the R453W and of the Dunnigan-type partial lipodystrophy R482W mutation of lamin A in C2C12 cells on transcription and epigenetic regulation of the myogenin (Myog) gene and on global chromatin organization. Expression of R453W-, but not R482W-lamin A, impairs activation of Myog and maintains a repressive chromatin state on the Myog promoter upon induction of differentiation, marked by H3 lysine (K) 9 dimethylation and failure to hypertrimethylate H3K4. Cells expressing WT-LaA also fail to hypertrimethylate H3K4. No defect occurs at the level of Myog promoter DNA methylation in any of the clones. Expression of R453W-lamin A and to a lesser extent R482W-lamin A in undifferentiated C2C12 cells redistributes H3K9me3 from pericentric heterochromatin. R453W-lamin A also elicits a redistribution of H3K27me3 from inactive X (Xi) and partial decondensation of Xi, but maintains Xist expression and coating of Xi, indicating that Xi remains inactivated. Our results argue that gene-specific and genome-wide chromatin rearrangements may constitute a molecular basis for laminopathies.
Assuntos
Epigênese Genética , Lamina Tipo A/genética , Distrofia Muscular de Emery-Dreifuss/genética , Mutação de Sentido Incorreto , Mioblastos/metabolismo , Miogenina/genética , Substituição de Aminoácidos , Animais , Arginina/genética , Diferenciação Celular , Linhagem Celular , Núcleo Celular/enzimologia , Metilação de DNA , Histonas/química , Histonas/metabolismo , Humanos , Lamina Tipo A/metabolismo , Metilação , Camundongos , Mioblastos/citologia , Miogenina/biossíntese , Regiões Promotoras Genéticas , Triptofano/genética , Regulação para Cima , Cromossomo X/enzimologiaRESUMO
In this paper, we describe a hidden two-compartment stochastic process used to model the kinetics of feline hematopoietic stem cells (HSCs) in continuous time. Because of the experimental design and data collection scheme, the inferential task presents numerous challenges. While the hematopoietic process evolves in continuous time, the observations are collected only at discrete irregular times and are a probabilistic function of the state of the process. In addition, the animals go through an experimental procedure such that their reserve of HSCs is severely depleted at the start of the observation period. This impedes any approximation of the hematopoietic process with a continuous state-space process (normal approximation of the transition probabilities would be inaccurate when the state of the process, i.e. the number of stem cells, is small). We implement a Markov chain Monte Carlo algorithm that allows us to estimate the posterior distribution of the parameters of the hematopoietic process while maintaining its state-space discrete (i.e. without using any approximation). We show the performance of the algorithm on simulated data. Finally, we apply the algorithm to data on multiple experimental cats and provide estimates of the rates of the fates of feline HSCs. The obtained estimates are in agreement with the estimates obtained with different methods published in the medical literature. However, the proposed approach makes a more efficient use of the data and hence the parameter estimates are much more accurate than the one obtained with the methods previously proposed.
Assuntos
Teorema de Bayes , Gatos/fisiologia , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Modelos Biológicos , Algoritmos , Animais , Gatos/sangue , Simulação por Computador , Feminino , Glucosefosfato Desidrogenase/sangue , Células-Tronco Hematopoéticas/citologia , Cadeias de Markov , Método de Monte Carlo , Processos Estocásticos , Cromossomo X/enzimologiaRESUMO
We describe here the characterization of the so far identified human DNase I family DNases, DNase I, DNase X, DNase gamma, and DNAS1L2. The DNase I family genes are found to be expressed with different tissue specificities and suggested to play unique physiological roles. All the recombinant DNases are shown to be Ca(2+)/Mg(2+)-dependent endonucleases and catalyze DNA hydrolysis to produce 3'-OH/5'-P ends. High activities for DNase I, DNase X, and DNase gamma are observed under neutral conditions, whereas DNAS1L2 shows its maximum activity at acidic pH. These enzymes have also some other peculiarities: different sensitivities to G-actin, aurintricarboxylic acid, and metal ions are observed. Using a transient expression system in HeLa S3 cells, the possible involvement of the DNases in apoptosis was examined. The ectopic expression of each DNase has no toxic effect on the host cells; however, extensive DNA fragmentation is observed only in DNase gamma-transfected cells after the induction of apoptosis. Furthermore, DNase gamma is revealed to be located at the perinuclear region in living cells, and to translocate into the nucleus during apoptosis. Our results demonstrate that DNase I, DNase X, DNase gamma, and DNAS1L2 have similar but unique endonuclease activities, and that among DNase I family DNases, DNase gamma is capable of producing apoptotic DNA fragmentation in mammalian cells.
Assuntos
Apoptose , Desoxirribonuclease I/química , Endodesoxirribonucleases/química , Endodesoxirribonucleases/metabolismo , Família Multigênica , Actinas/farmacologia , Transporte Ativo do Núcleo Celular/genética , Adulto , Sequência de Aminoácidos , Apoptose/genética , Linhagem Celular , Desoxirribonuclease I/antagonistas & inibidores , Desoxirribonuclease I/genética , Desoxirribonuclease I/metabolismo , Endodesoxirribonucleases/antagonistas & inibidores , Endodesoxirribonucleases/biossíntese , Endodesoxirribonucleases/genética , Ativação Enzimática/genética , Inibidores Enzimáticos/farmacologia , Células HeLa , Humanos , Hidrólise , Dados de Sequência Molecular , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transfecção , Cromossomo X/enzimologiaRESUMO
JIL-1 is a novel chromosomal kinase that is upregulated almost twofold on the male X chromosome in Drosophila. Here we demonstrate that JIL-1 colocalizes and physically interacts with male specific lethal (MSL) dosage compensation complex proteins. Furthermore, ectopic expression of the MSL complex directed by MSL2 in females causes a concomitant upregulation of JIL-1 to the female X that is abolished in msl mutants unable to assemble the complex. Thus, these results strongly indicate JIL-1 associates with the MSL complex and further suggests JIL-1 functions in signal transduction pathways regulating chromatin structure.
Assuntos
Cromatina/enzimologia , Proteínas de Drosophila , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Cromossomo X/enzimologia , Animais , Proteínas de Ligação a DNA , Drosophila , Feminino , Dosagem de Genes , Regulação Enzimológica da Expressão Gênica/fisiologia , Histonas/metabolismo , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Masculino , Fosforilação , Ligação Proteica/genética , Transcrição GênicaRESUMO
Ornithine transcarbamylase (OTC) deficiency is the most common urea cycle disorder. It is X-linked and hemizygous new-born males usually suffer fatal hyperammonemia. In contrast, carrier females manifest variable phenotypes, ranging from asymptomatic carriers to those with severe hyperammonemia. In order to understand the correlation between X-inactivation status and the clinical phenotype of carrier females with this disorder, we analyzed the X-inactivation pattern of peripheral blood leukocytes in a family consisting of a clinically normal mother and two daughters with severe manifestation. In addition, we obtained tissue samples from various parts of the liver of one of these daughters and analyzed X-inactivation patterns and the residual OTC activities. The X-inactivation of peripheral blood leukocytes was nearly random in these carrier females and showed no correlation with the disease phenotype. However, the X-inactivation of the liver was much more skewed and correlated well with the OTC activity of all samples. Interestingly, the degree of X-inactivation varied considerably, even within the same liver.
Assuntos
Mecanismo Genético de Compensação de Dose , Fígado/enzimologia , Doença da Deficiência de Ornitina Carbomoiltransferase , Cromossomo X/genética , Criança , Pré-Escolar , DNA/análise , DNA/genética , Regulação Enzimológica da Expressão Gênica , Humanos , Lactente , Fígado/metabolismo , Transplante de Fígado , Masculino , Ornitina Carbamoiltransferase/genética , Linhagem , Compostos de Amônio Quaternário/sangue , Cromossomo X/enzimologiaRESUMO
Cytogenetic studies have shown that the Y chromosome is eliminated from many somatic cell types of the bandicoot Isoodon macrourus, an Australian marsupial. Molecular techniques allow examination of a greater range of tissue types than that possible using cytogenetic techniques. The presence or absence of the Y chromosome was established using partial sequences of the Y-linked SRY and UBE1Y genes in I. macrourus, with the X-linked gene G6PD as a control. We show that a very small proportion of cells comprising hematopoietic tissues, and even fewer cells in peripheral blood, retain the Y chromosome. The Y chromosome is retained in most brain, liver, kidney, and lung cells and in cardiac and skeletal muscle. We also show that the bandicoot Y chromosome is retained in some cell types within tissues previously believed to completely eliminate the Y chromosome.
Assuntos
Aberrações Cromossômicas/genética , Marsupiais/genética , Proteínas Nucleares , Cromossomos Sexuais/genética , Fatores de Transcrição , Cromossomo Y/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Southern Blotting , DNA/química , DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Fibroblastos/química , Fibroblastos/citologia , Fibroblastos/metabolismo , Marcadores Genéticos/genética , Glucosefosfato Desidrogenase/genética , Glucosefosfato Desidrogenase/metabolismo , Cariotipagem , Ligases/genética , Ligases/metabolismo , Masculino , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Proteína da Região Y Determinante do Sexo , Distribuição Tecidual , Ubiquitina-Proteína Ligases , Cromossomo X/enzimologia , Cromossomo X/genética , Cromossomo Y/químicaRESUMO
PURPOSE: To evaluate the role of TIMP-1 in inherited retinal degeneration. METHODS: The genomic structure of the TIMP-1 gene was established and male patients with x-linked retinitis pigmentosa 2 from five families were screened for sequence alterations by direct sequencing in all exons, exon-intron boundaries, and the 5' upstream region of the gene. RESULTS: TIMP-1 appears to be expressed in the retina at low levels and consists of six exons spanning a genomic region of approximately 4.5 kb on Xp11.23. No disease-specific sequence alterations were identified. A site substitution in exon 5 was observed in samples from control subjects and patients, but it did not alter the amino acid sequence of the protein product. CONCLUSIONS: The results of this study exclude mutations in the TIMP-1 coding sequence, splice sites, and the 5' upstream region as a cause of retinal degeneration in x-linked retinitis pigmentosa 2. However, an as yet unidentified regulatory element that lies outside these intervals may be implicated. The role of this tightly regulated protein in the normal functioning of the retina has yet to be determined.
Assuntos
Ligação Genética , Glicoproteínas/genética , Inibidores de Proteases , Retinose Pigmentar/enzimologia , Cromossomo X/enzimologia , Sequência de Bases , DNA/análise , Análise Mutacional de DNA , Primers do DNA/química , Éxons , Humanos , Íntrons , Masculino , Dados de Sequência Molecular , Retina/enzimologia , Retinose Pigmentar/etiologia , Inibidores Teciduais de MetaloproteinasesRESUMO
X chromosome inactivation is associated with a highly asynchronous pattern of DNA replication at most X-linked loci in females. We studied the human HPRT locus, which is subject to X inactivation and expressed from only the active homolog, with the goal of comparing replication properties between the active and inactive homologs in this region using a fluorescence in situ hybridization approach. We found that in normal female lymphoblasts this locus is replicated in a highly asynchronous manner across a broad, discrete 500-600 kb zone with earliest replication appearing at the gene coding sequence. This general timing profile is maintained in normal male lymphoblasts, as well as in hamster x human hybrid cells containing the active human X chromosome. However, the inactive human X chromosome in the hamster cell background does not appear to function in a fully equivalent manner to the normal inactive X chromosome in female cells. Furthermore, reactivation of the inactive human X chromosome in a hamster x human hybrid system by 5-azacytidine treatment and HAT selection restores early replication at the HPRT gene itself, but does not change the overall domain behavior.
Assuntos
Replicação do DNA , Hipoxantina Fosforribosiltransferase/genética , Cromossomo X/enzimologia , Animais , Azacitidina/farmacologia , Linhagem Celular Transformada , Cromatina/química , Clonagem Molecular , Cricetinae , Replicação do DNA/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Células Híbridas , Hipoxantina Fosforribosiltransferase/efeitos dos fármacos , Masculino , Fatores de Tempo , Cromossomo X/efeitos dos fármacosRESUMO
Norrie disease is a rare disease of newborn males caused by prenatal or perinatal retinal detachment, which may be associated with mental retardation, psychosis, and/or hearing loss. DXS7 (L1.28) and MAO A and B loci have been linked to the ND locus on the short arm of the X chromosome. Sequences homologous to OAT also have been mapped to the short arm of the X chromosome. We performed linkage analyses between the ND locus and one of the OAT-like clusters of sequences on the X chromosome (OATL1), using a ScaI RFLP in a ND family, and increased the previously calculated lod score (z) to over 3 (3.38; theta = 0.05). Similarly, we calculated a lod score of 4.06 (theta = 0.01) between the OATL1 and DXS7 loci. Alone, the OATL1 ScaI RFLP system is expected to be informative in 48% of females. If this system were used in combination with the DXS7 TaqI polymorphism, 71% of females would be informative for at least one of the markers and 21% would be informative for both. Because the OATL1 ScaI RFLP is a relatively common polymorphism, this system should be useful for the identification of ND carriers and affected male fetuses and newborns.
Assuntos
Ligação Genética , Ornitina-Oxo-Ácido Transaminase/genética , Retina/anormalidades , Cromossomo X/enzimologia , DNA/análise , Sondas de DNA , Desoxirribonucleases de Sítio Específico do Tipo II , Humanos , Escore Lod , Masculino , Linhagem , Polimorfismo de Fragmento de Restrição , Descolamento Retiniano/genéticaRESUMO
The measurement of the activity of the X-linked enzyme HPRT has been widely used as an indicator of X-chromosome activity during preimplantation development in the mouse. More recently, the concomitant measurement of the activity of the autosomally-encoded enzyme APRT has been used in an attempt to decrease the variability inherent in the measurement of enzyme activity from minute samples such as preimplantation embryos. In this study the use of the HPRT-deficient mouse mutant, Hprtb-m3, allowed the unequivocal identification of the parental origin of HPRT activity measured in embryos derived from crosses between wild-type mice, and mice which were homozygous or hemizygous for the Hprtb-m3 allele. Results were similar to those of a previous study, where oocyte-encoded HPRT activity accounted for about 10% of total HPRT activity at 76 hours post human chorionic gonadotrophin injection and the paternally-derived Hprt allele was shown to be transcriptionally active by the late 2-cell stage. In contrast to other studies, differential expression of the two Hprt alleles was detected during the preimplantation period, in embryos derived from crosses between wild-type and HPRT-deficient mice. Evidence was also found for the existence of an X-linked locus which influences the amount of APRT activity in the unfertilized oocyte. We propose that the expression pattern of this locus may be influenced by its parental origin.
Assuntos
Adenina Fosforribosiltransferase/genética , Blastocisto/fisiologia , Desenvolvimento Embrionário e Fetal/genética , Expressão Gênica/fisiologia , Hipoxantina Fosforribosiltransferase/genética , Adenina Fosforribosiltransferase/metabolismo , Animais , Blastocisto/enzimologia , Desenvolvimento Embrionário/fisiologia , Feminino , Hipoxantina Fosforribosiltransferase/efeitos dos fármacos , Hipoxantina Fosforribosiltransferase/metabolismo , Masculino , Camundongos , Camundongos Mutantes , Oócitos/enzimologia , Gravidez , Cromossomo X/enzimologiaRESUMO
Northern analysis using 32P-labeled subfragments of human liver MAO-A and B cDNA clones detected 5Kb and 3Kb transcripts, respectively in fetal tissues and adult brains. The tissue distribution of these transcripts was determined. Small intestine and placenta express, in addition to the MAO-A 5Kb transcript, a 2Kb transcript determined to lack a 3' flanking region. MAO-A appeared prior to MAO-B in the fetal brain, whereas both MAO-A and B were found in adult brain.
Assuntos
Monoaminoxidase/genética , Northern Blotting , Encéfalo/enzimologia , Encéfalo/ultraestrutura , Sondas de DNA , Feminino , Feto/metabolismo , Regulação Enzimológica da Expressão Gênica , Humanos , Intestino Delgado/enzimologia , Placenta/enzimologia , Gravidez , RNA Mensageiro/metabolismo , Frações Subcelulares/metabolismo , Transcrição Gênica , Cromossomo X/enzimologiaRESUMO
Cloned cDNA sequences representing exons from the Duchenne/Becker muscular dystrophy (DMD/BMD) gene were used for deletion screening in a population of 287 males males affected with DMD or BMD. The clinical phenotypes of affected boys were classified into three clinical severity groups based on the age at which ambulation was lost. Boys in group 1 had DMD, losing ambulation before their 13th birthday; those in group 2 had disease of intermediate severity, losing ambulation between the ages of 13 and 16 years; and boys in group 3 had BMD, being ambulant beyond 16 years. A fourth group consisted of patients too young to be classified. Clinical group allocation was made without previous knowledge of the DNA results. A gene deletion was found in 124 cases where the clinical severity group of the affected boy was known. The extent of the deletions was delineated using cDNA probes. There were 74 different deletions. Fifty-five of these were unique to individual patients, but the other 19 were found in at least two unrelated patients. The different clinical groups showed generally similar distributions of deletions, and the number of exon bands deleted (that is, deletion size) was independent of phenotype. Some specific deletion types, however, correlated with the clinical severity of the disease. Deletion of exons containing HindIII fragments 33 and 34 and 33 to 35 were associated with BMD and were not found in patients with DMD. Deletions 3 to 7 occurred in four patients with the intermediate phenotype and one patient with BMD. Other shared deletions were associated with DMD, although in four cases patients with disease of intermediate severity apparently shared the same deletion with boys with DMD. The range of phenotypes observed, and the overlap at the genetic level between severe and intermediate and mild and intermediate forms of dystrophy, emphasizes the essential continuity of the clinical spectrum of DMD/BMD. There were no characteristic deletions found in boys with mental retardation or short stature which differed from deletions in affected boys without these features.
Assuntos
Deleção Cromossômica , Distrofias Musculares/genética , Índice de Gravidade de Doença , Cromossomo X/enzimologia , Adolescente , Estatura/genética , Criança , Desoxirribonuclease HindIII , Éxons , Humanos , Masculino , Distrofias Musculares/diagnóstico , Distrofias Musculares/fisiopatologia , Desempenho Psicomotor/fisiologiaRESUMO
In an animal model for Lesch-Nyhan syndrome, the affected male embryos, as well as the carrier female embryos, have been successfully identified by biochemical microassay of a sample of trophectoderm cells taken from the mouse embryos at the blastocyst stage. The embryos were removed from the uterus, diagnosed and returned to the uterus within 2 days without the need for cryopreservation. The diagnosis was confirmed at 14 days gestation by analysis of the hypoxanthine phosphoribosyl transferase (HPRT) status of the fetuses. Live young were obtained from biopsied embryos after transfer.
Assuntos
Blastocisto/enzimologia , Modelos Animais de Doenças , Desenvolvimento Embrionário , Hipoxantina Fosforribosiltransferase/metabolismo , Síndrome de Lesch-Nyhan/diagnóstico , Animais , Biópsia , Blastocisto/ultraestrutura , Células Cultivadas , Transferência Embrionária , Feminino , Triagem de Portadores Genéticos , Ligação Genética , Hipoxantina Fosforribosiltransferase/deficiência , Hipoxantina Fosforribosiltransferase/genética , Técnicas In Vitro , Síndrome de Lesch-Nyhan/enzimologia , Síndrome de Lesch-Nyhan/genética , Camundongos , Camundongos Mutantes , Gravidez , Cromossomo X/enzimologiaRESUMO
Previous evidence had shown that, when a mammalian X-chromosome is broken by a translocation, only one of the two X-chromosome segments shows cytological signs of X-inactivation in the form of late replication or Kanda staining. In the two mouse X-autosome translocations T(X;4)37H and T(X;11)38H the X-chromosome break is in the A1-A2 bands; in both, the shorter translocation product fails to exhibit Kanda staining. By in situ hybridization, the locus of ornithine carbamoyltransferase (OCT) was shown to be proximal to the breakpoint (i.e. on the short product) in T37H and distal to the breakpoint in T38H. Histochemical staining for OCT showed that in T38H the locus of OCT undergoes random inactivation, as in a chromosomally normal animal, whereas in T37H the OCT locus remains active in all cells. The interpretation is that, when a segment of X-chromosome is physically separated from the X-inactivation centre, it fails to undergo inactivation. This point is important for the understanding of the mechanism of X-inactivation, since it implies that inactivation is a positive process, brought about by some event that travels along the chromosome. It is also relevant to the interpretation of the harmful effects of X-autosome translocations and the abnormalities seen in individuals carrying such translocations.
Assuntos
Mecanismo Genético de Compensação de Dose , Cromossomo X , Animais , Bandeamento Cromossômico , Marcadores Genéticos , Histocitoquímica , Camundongos , Ornitina Carbamoiltransferase/metabolismo , Translocação Genética , Cromossomo X/enzimologiaRESUMO
Experimental conditions are discussed that are necessary for a useful application of genetically marked laboratory animals to distinguish between clonal and nonclonal origin of induced tumors: quantitative discrimination of biochemical markers, definition of the "patch" sizes, evaluation of possible admixture of "contaminating" cells and the approach to congenicity of parental animal strains.
Assuntos
Isoenzimas/metabolismo , Neoplasias Mamárias Experimentais/enzimologia , Fosfoglicerato Quinase/metabolismo , Cromossomo X/enzimologia , Alelos , Animais , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Isoenzimas/genética , Fígado/enzimologia , Camundongos , Fosfoglicerato Quinase/genéticaRESUMO
The immunodeficiency in CBA/N mice is reflected by abnormal development of a subset of B lymphocytes. However, it is not clear how xid, the mutant gene in CBA/N mice, affects the development of this subset. Specifically, it is not known if the xid gene influences the development of the B cell subset directly or indirectly by providing the improper developmental milieu through effects on other cells. We investigated this question using female mice heterozygous for two x chromosomal genes, xid and Pgk-1 (phosphoglycerate kinase-1). Since females are mosaic because of x chromosome inactivation, their lymphocytes can be studied for the choice of the x chromosome, using the two PGK-1 isoenzymes as the cytological marker. We find that B lymphocytes in the spleen prefer the x chromosome without xid while the remaining splenocytes and cells from other tissues do not. This suggests that xid affects B lymphocytes directly and not through their developmental milieu. Furthermore, our data suggest that the precursors for IgG1- and IgG3-producing cells may be both few and different.
Assuntos
Linfócitos B/imunologia , Síndromes de Imunodeficiência/imunologia , Camundongos Endogâmicos CBA/genética , Mosaicismo , Animais , Encéfalo/enzimologia , Feminino , Ligação Genética , Células-Tronco Hematopoéticas/enzimologia , Hibridomas/imunologia , Imunoglobulina G/biossíntese , Síndromes de Imunodeficiência/genética , Isoenzimas/metabolismo , Fígado/enzimologia , Camundongos , Fosfoglicerato Quinase/metabolismo , Aberrações dos Cromossomos Sexuais/genética , Aberrações dos Cromossomos Sexuais/imunologia , Baço/citologia , Cromossomo X/enzimologiaRESUMO
Human--hamster somatic cell hybrids have been obtained by fusion of a CHO line (NA31) doubly deficient in hypoxanthine guanine phosphoribosyltransferase and glucose 6-phosphate dehydrogenase (G6PD) with normal G6PD(+) human fibroblasts. Analysis of NA31 extracts has revealed that, although G6PD activity is nearly absent, significant activity can be detected with 2-deoxyglucose 6-phosphate as substrate, so that the mutant and normal forms of the enzyme can both be easily detected. The cell hybrids obtained express human G6PD. The human G6PD subunits are distributed in homodimeric molecules as well as in human--hamster heterodimeric molecules. However, whereas the amount of hamster G6PD subunits present in the hybrid is similar to that in the hamster parental cells, the amount of human G6PD subunits is decreased by 3- to 10-fold when compared to the human parental cell. These results indicate that either the expression of the G6PD gene or the stability of the gene product is altered in the hybrid. By mutagenesis and selection in diamide (a substance that oxidizes intracellular glutathione), we have isolated a clone with a 3- to 5-fold increase in human G6PD activity. This derivative may have an increased rate of expression of the human G6PD structural gene.