Heritable DNA Research Articles

Prenatal diagnosis of a hypermethylated full fragile X mutation in chorionic villi of a male fetus.

Fragile X syndrome, one of the most common human genetic diseases, is characterised by a unique genetic mechanism which involves dynamic mutation because of a heritable unstable DNA sequence and abnormal DNA methylation. Direct detection of the dynamic mutation and its methylation status at the DNA level would facilitate reliable tests for prenatal and postnatal diagnosis of the disease and for carrier detection. However, it has been suggested that DNA methylation can not be used as the basis for prenatal diagnosis as the CpG island is not always methylated in chorionic villus DNA. We report here a male fetus exhibiting both extensive somatic heterogeneity and abnormal hypermethylation of the full fragile X mutation in chorionic villus DNA as well as in fetal tissue DNA. Our results indicate that both somatic heterogeneity and hypermethylation of the full fragile X mutation are events that are clearly detectable in the 11th to 12th week of pregnancy.

A 43-base-pair complementary DNA sequence homology and triplet repeat motif among putative polymeric immunoglobulin receptor messenger RNAs in regenerating rat liver

HepatologyVolume 18, Issue 1 p. 226-228 CorrespondenceFree Access A 43-base-pair complementary DNA sequence homology and triplet repeat motif among putative polymeric immunoglobulin receptor messenger RNAs in regenerating rat liver K. S. Koch PH.D., K. S. Koch PH.D. Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorR. G. Fletcher, R. G. Fletcher Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorM. P. Grond, M. P. Grond Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorH. L. Leffert M. D., H. L. Leffert M. D. Department of Pharmacology and Center for Molecular Genetics University of California, San Diego La Jolla, California 92093–0636Search for more papers by this author K. S. Koch PH.D., K. S. Koch PH.D. Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorR. G. Fletcher, R. G. Fletcher Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorM. P. Grond, M. P. Grond Department of Pharmacology University of California, San Diego La Jolla, California 92093–0636Search for more papers by this authorH. L. Leffert M. D., H. L. Leffert M. D. Department of Pharmacology and Center for Molecular Genetics University of California, San Diego La Jolla, California 92093–0636Search for more papers by this author First published: July 1993 https://doi.org/10.1002/hep.1840180138Citations: 2AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat References 1 Lu XP, Koch KS, Lew DJ, Dulic V, Pines J, Reed SI, Hunter T. Induction of cyclin mRNA and cyclin-associated histone H1 kinase during liver regeneration. J Biol Chem 1992; 267: 2841– 2844. 2 Koch KS, Lu XP, Brenner DA, Fey GH, Martinez-Conde A, Leffert HL. Mitogens and hepatocyte growth control in vivo and in vitro. In Vitro Cell Dev Biol 1990; 26: 1011– 1023. 3 Lew DJ, Dulic V, Reed SI. Isolation of three novel human cyclins by rescue of G1 cyclin (Cln) function in yeast. Cell 1991; 66: 1197– 1206. 4 Schuler GD, Altschul SF, Lipman DJ. A workbench for multiple alignment construction and analysis. Proteins Struct Funct Genet 1991; 9: 180– 190. 5 Banting G, Brake B, Braghetta P, Luzio JF, Stanley KK. Intracellular targetting signals of polymeric immunoglobulin receptors are highly conserved between species. FEBS Lett 1989; 254: 177– 183. 6 Karlin S, Altschul SF. Methods for assessing the statistical significance of molecular sequence features by using general scoring schemes. Proc Natl Acad Sci USA 1990; 87: 2264– 2268. 7 Karlin S, Dembo A, Kawabata T. Ann Stat 1990; 18: 571– 581. 8 Huling S, Fournier GR, Feren A, Chuntharapai A, Jones AL. Ontogeny of the secretory immune system: maturation of a functional polymeric immunoglobulin receptor regulated by gene expression. Proc Natl Acad Sci USA 1992; 89: 4260– 4264. 9 Proudfoot NJ, Whitelaw E. Chapter 3. In: BD Hames, DM Glover, eds. Transcription and Splicing. Oxford: IRL Press, 1988; 97– 129; and Richards RI, Sutherland GR. Heritable DNA sequences. Nature Genetics 1992; 1: 7– 9. Citing Literature Volume18, Issue1July 1993Pages 226-228 ReferencesRelatedInformation

The fragile X syndrome: no evidence for any recent mutations.

Fragile X (fra(X)) syndrome, the most common form of familial mental retardation, is caused by heritable unstable DNA composed of CGG repeats. As reproductive fitness of fra(X) patients is severely compromised, a high mutation rate has been proposed to explain the high prevalence. However, we have been unable to show any new mutation for 84 probands referred to us to date. We show here the same fra(X) gene in five fra(X) probands with common ancestors married in 1747. The lack of new fra(X) mutations implies that there must be many more fra(X) gene carriers in the population than previously realised. As it is now possible to detect asymptomatic fra(X) gene carriers by DNA analysis, extended family studies for any new proband are recommended. A family illustrating the importance of fra(X) carriership determination is reported.

Heritable unstable DNA sequences and hypermethylation associated with fragile X syndrome in Japanese families.

Fragile X syndrome, associated with the fragile site at Xq27.3 (FRAXA), is the most common form of familial mental retardation. The fragile X mutation has recently been characterized as a heritable unstable DNA sequence, p(CCG)n/p(CGG)n, in the FRAXA locus. In the present study, a correlation between fragile X-genotypes in the FRAXA locus and hypermethylation of an adjacent CpG island was examined in four Japanese families with fragile X syndrome. We show here that the heritable unstable DNA sequences in the fragile X chromosome usually increase in size when transmitted by female carriers, and that the degree of methylation in the CpG island correlated with the increased sizes of the unstable DNA sequences. When a hypermethylated full mutation was transmitted by a male to his daughters, both the size of the unstable DNA sequence and the degree of the methylation reduced to the premutation range. Our observations suggest that female meiosis has a greater potential for amplifying unstable DNA sequences and that amplified DNA sequences can be transmitted through germ cells, while male germ cells seem not to be able to tolerate highly amplified unstable DNA sequences.

The mechanism of fission yeast mating-type interconversion: evidence for two types of epigenetically inherited chromosomal imprinted events.

Mating types of the fission yeast S. pombe interchange such that only one cell among four granddaughters of a cell ever switches, and the sister of the newly switched cell switches efficiently in consecutive cell divisions, thereby producing chains of recurrent switching. The developmental program is known to be mediated through specific parental chromosomal DNA-strand inheritance at the mating-type locus (mat1). A heritable DNA strand- and sequence-specific imprinting event was previously postulated to be required to cleave the chromosome at mat1 to initiate recombination required for switching. The pedigree analysis presented here demonstrates that swi1, swi3, and swi7 genes are required for the DNA cleavage step directly, and not for the imprinting function, since mutations in these genes reduce the efficiency of initial (i.e., switching of one in four granddaughters) and recurrent switching equally. However, when the mat1 gene sequence is placed adjacent to the indigenous mat1 locus, apparently it is imprinted inefficiently, since recurrent switches of the inserted locus occur nearly five times more often than the initial switches. This "runaway switching" is the first evidence formally demonstrating both the existence of and the requirement for an imprinted mat1 DNA, making it cleavable and consequently swichable. Second, the double-stranded break constitutes another imprinted event as it is evidenced to be inherited from the parental cell.

Heritable unstable DNA sequences

Heritable unstable DNA sequences

Fragile X syndrome: molecular analysis reveals a new mechanism of mutation in human genetic diseases.

The fragile X syndrome belongs to the most common genetic diseases and has a prevalence of one in every 2000 children. The syndrome is named after the fragile site in q27.3 on the X chromosome. The molecular cloning of the DNA containing the fragile site has resulted in the identification of a heritable unstable DNA sequence revealing a new mechanism of mutation in human genetic disorders. This DNA sequence significantly facilitates the diagnosis and provides a rapid method for carrier detection and prenatal diagnosis. The unstable element is located within a candidate gene, FMR1. The FMR1 protein is not made in fragile X patients and nothing is known about its function. We will have to await studies on this protein to be able to understand the variable phenotype of this disease.

Mutations and epimutations in mammalian cells

Early studies on heritable variation in cultured mammalian cells suggested that both mutation and epigenetic events might be involved. The importance of mutations has subsequently been fully documented, but only recently has an alternative form of inheritance been uncovered. This is based on the post-synthetic methylation of cytosine in regulatory regions of genes. The pattern of methylation is heritable, and in almost all cases studied, methylation of a region is associated with lack of gene expression. Such silent genes can be reactivated by the powerful demethylating agent 5-azacytidine (5-aza-CR). Changes in heritable DNA methylation which alter phenotype are referred to as epimutations. It now seems very likely that the well known ‘functional hemizygosity’ in CHO cells and other near diploid cell lines is due to the existence of one active and one silent gene at many autosomal loci. It is clear that permanent cell lines inactive genes by de novo methylation, whereas normal diploid cells do not have this activity. This has important implications for our understanding of cellular transformation, tumor progression, and the increase in chromosome number frequently associated with these cellular changes. It is likely that both mutations and epimutations are important in the emergence of fully transformed tumorigenic cells. Agents which increase or reduce DNA methylation in cells can be regarded as epimutagens, although in many cases the mechanisms of inducing hypo- or hyper-methylation are not understood. Two exceptions are 5-aza-CR which inhibits the normal DNA maintenance methylase activity, and 5-methyldeoxycytidine triphosphate which is incorporated into cellular DNA following electroporation and has been shown to silence genes.

Hereditary unstable DNA: a new explanation for some old genetic questions?

Fragile X syndrome, associated with the fragile X chromosome, is the most common cause of familial mental retardation. The condition is characterised by a heritable DNA sequence that consists of an abnormal number of CCG repeats, and which is unstable in both mitosis and meiosis. We suggest that such heritable unstable DNA sequences could be present in other parts of the genome and that these might explain a number of genetic events that are not well understood in terms of classic genetic mechanisms. Such poorly explained observations include anticipation, incomplete penetrance, variable expression, and possibly imprinting, variegation, and multifactorial inheritance.

Genomic imprinting and allelic exclusion

In diploid cells, allelic exclusion reduces genes to functional haploidy, because only one of two alleles is active. It is best known in cells producing immunoglobulins, but other examples also exist. X-chromosome inactivation in female mammals is related to allelic exclusion, but in this case the dosage compensation mechanism extends to the whole chromosome. Functional hemizygosity in some mammalian cell lines is probably also due to allelic exclusion, where one autosomal allele is active and the other is methylated and inactive. In early development, it may be important to have only one functional copy of specific regulatory genes. If one considers the possible mechanisms whereby genes are switched from an active to an inactive form, or vice versa, complications arise if the same type of switch operates in two homologous chromosomes segregating independently at mitosis. This complication is avoided if one of the genes is totally inactive. It is therefore suggested that important regulatory gene are subject to allelic exclusion and that this provides a basis for genomic imprinting. Male or female gametes complement in the zygote, because they may have different inactive genes, and the active allele in each case is then functionally haploid in the zygote and developing embryo. These haploid genes would be those involved in critical switches of gene activity during the developmental process. Allelic exclusion imposed by imprinting might be based on the heritable DNA methylation of the regulatory regions of silent genes.

A further assessment of factors influencing measurements of thioguanine-resistant mutant frequency in circulating T-lymphocytes

We have used the T-Lymphocyte cloning technique as a method of monitoring the human population for somatic cell mutant frequency. We present a statistical analysis of the experimental factors which may influence the observed mutant frequency. We have obtained consistently high plating efficiencies of T-cells from the mononuclear cell fraction from donor blood samples (mean of 56%, based on 123 observations from 70 individuals). Nevertheless, an inverse correlation of mutant frequency with plating efficiency was observed, and some experimental factors (serum and interleukin-2 batch, and worker) may have a significant effect on the observed mutant frequency. We discuss the difficulties that these possible effects present in establishment of a reference database and design of long-term studies. No significant effect of donor sex on mutant frequency was observed, but age (1.3% increase per year for normal adults) and smoking (56% increase over normal non-smokers) both significantly increased the mutant frequency. We discuss the utility of the assay for the monitoring of populations for heritable DNA damage, and we compare the results to those obtained with lymphocytes using other endpoints, e.g. chromosome aberrations, micronuclei and sister-chromatid exchange.

Restoration of u.v.-induced excision repair in Xeroderma D cells transfected with the denV gene of bacteriophage T4.

The heritable DNA repair defect in human Xeroderma D cells, which results in failure to incise at u.v. light-induced pyrimidine dimers, has been partially but stably corrected by transfection of immortalised cells with the denV pyrimidine dimer glycosylase gene of bacteriophage T4. Transfectants selected either for a dominant marker on the mammalian vector carrying the prokaryotic gene or for the dominant marker plus resistance to killing by u.v. light, have been shown to express the denV gene to varying degrees. denV expression results in significant phenotypic change in the initially repair-deficient, u.v.-hypersensitive cells. Increased resistance to u.v. light and more rapid recovery of replicative DNA synthesis following u.v. irradiation have been correlated both with improved repair DNA synthesis and with a novel dimer incision capability present in denV transfected Xeroderma cells but not as evident in transfected normal cells. Most of the transfectants contain a single integrated copy of the denV gene; increase in denV copy number does not result in either increased gene expression or enhanced survival to u.v. light. These results show that expression of a heterologous prokaryotic repair gene can partially compensate for the genetic defect in a human Xeroderma D cell.

Dermatologic needs in drugs and instrumentation

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Bacterial resistance to antibiotics.

Effective antibacterial drugs have been available for nearly 50 years. After the introduction of each new such drug, whether chemically synthesized or a naturally occurring antibiotic, bacterial resistance to it has emerged. The genetic mechanisms by which bacteria have acquired resistance were quite unexpected; a new evolutionary pathways has been revealed. Although some antibiotic resistance has resulted from mutational changes in structural proteins--targets for the drugs' action--most has resulted from the acquisition of new, ready-made genes from an external source--that is, from another bacterium. Vectors of the resistance genes are plasmids--heritable DNA molecules that are transmissible between bacterial cells. Plasmids without antibiotic-resistance genes are common in all kinds of bacteria. Resistance plasmids have resulted from the insertion of new DNA sequences into previously existing plasmids. Thus, the spread of antibiotic resistance is at three levels: bacteria between people or animals; plasmids between bacteria; and transposable genes between plasmids.

Heritable Nuclear DNA Changes in Doubled Haploid Plants Obtained by Pollen Culture of Nicotiana Sylvestris

Heritable quantitative and qualitative changes can be observed in the nuclear DNA of doubled-haploid (D.H.) plants obtained by pollen culture of Nicotiana sylvestris . D.H. plants resulting from consecutive androgenic cycles contain on the average increasing amounts of total DNA and increasing proportions of highly repeated sequences. All the results suggest that both A-T rich and G-C rich regions are amplified in the D.H. genome. Some hypotheses on the origin of the changes are discussed.