A strain-specific modifier on mouse chromosome 4 controls the methylation of independent transgene loci

Engler, Peter; Haasch, Deanna L.; Pinkert, Carl A.; Doglio, Lynn; Glymour, M. Maria; Brinster, Ralph L.; Storb, Ursula

doi:10.1016/0092-8674(91)90546-b

Cited by 205 publications

(135 citation statements)

References 27 publications

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“…This imprinting was independent of the genetic background in which it was measured, as the imprinting was observed in both progeny outcrossed to CD1 mice, as well as mice that had been inbred for three generations into DBA/2J (data not shown). Thus, the imprinting cannot be explained by the presence of a single dominant genetic modifier, as has been observed for other transgenes (Hadchouel et al 1987;Sapienza et al 1989;Allen et al 1990;Engler et al 1991), although the presence of complex genetic modifications cannot be ruled out until the transgenes are on completely inbred backgrounds. The appropriate imprinting of the H19 transgene confirms that the elements required for its allele-specific expression are localized to a small region surrounding the structural gene.…”

Section: Discussionmentioning

confidence: 99%

Epigenetic mechanisms underlying the imprinting of the mouse H19 gene.

Bartolomei¹,

Webber²,

Brunkow³

et al. 1993

Genes Dev.

456

324

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The expression of the H19 gene is governed by parental imprinting in mammals. H19, an unusual gene encoding an RNA with no known function, is exclusively expressed from the maternal chromosome. In mouse, it lies 90 kb downstream from the lgf2 gene, which encodes a fetal-specific growth factor, insulin-like growth factor II, and is expressed primarily from the paternally inherited chromosome. In this report we have utilized interspecific hybrid mice to identify male-specific DNA methylation of a 7-to 9-kb domain surrounding the 1-119 gene and its promoter. This allele-specific methylation could function as a mark to suppress transcription of the H19 paternal allele. Consistent with this proposal, the H19 promoter displayed an open chromatin conformation only on the relatively unmethylated active maternal allele. In contrast, a cell type-specific enhancer that lies outside the methylation domain is hypersensitive to restriction enzyme digestion in nuclei on both maternal and paternal chromosomes. That the allele-specific methylation domain, coupled to the two H19 enhancers, contains all the information necessary for its imprinting was tested by examining two transgenic lines containing an internally deleted H19 transgene. Both displayed paternal-specific methylation of the transgene and maternal-specific expression. Although neither line has been tested in an inbred genetic background, and therefore the action of complex modifiers cannot be formally excluded, the result suggests that the sequences necessary for the imprinting of H19 have been identified.

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Section: Discussionmentioning

confidence: 99%

Epigenetic mechanisms underlying the imprinting of the mouse H19 gene.

Bartolomei¹,

Webber²,

Brunkow³

et al. 1993

Genes Dev.

456

324

View full text Add to dashboard Cite

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“…22,23 It has been postulated that under-methylated satellite sequences confer abnormal chromatin structures and predispose to chromosomal instability by enhancing chromosome recombinations. 24 A high frequency of chromosome rearrangements with pericentromeric or heterochromatic breakpoints has been observed in many tumors, which may suggest a relationship between satellite hypomethylation, chromosome instability and carcinogenesis. 25,26 Our current finding supported a relationship between hypomethylated Sat2 sequences and recurrent aberrant 1q formation.…”

Section: Discussionmentioning

confidence: 99%

Hypomethylation of Chromosome 1 Heterochromatin DNA Correlates with q-Arm Copy Gain in Human Hepatocellular Carcinoma

Wong

Lam

Lai

et al. 2001

The American Journal of Pathology

122

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Using comparative genomic hybridization (CGH) analysis, we, and others, have shown that there is a high and consistent incidence of chromosome 1q copy gain in human hepatocellular carcinoma (HCC). Chromosome 1 rearrangements, that involved pericentromeric breakpoints, have also been frequently reported in karyotypic studies of HCC. Satellite DNA hypomethylation has been postulated as the mechanism underlying the induction of chromosome 1 pericentromeric instability in many human cancers and in individuals with the rare recessive disorder ICF (immunodeficiency, centromeric heterochromatin instability, facial anomalies). In this study, we have investigated the role of DNA hypomethylation in 1q copy gain in HCC by examining the methylation status of chromosome 1 heterochromatin DNA (band 1q12). Thirty-six histologically confirmed samples of HCC were studied (24 paired tumor and adjacent nontumorous liver tissues, and 12 tumor only). Hypomethylation of satellite 2 (Sat2) DNA in 1q12 was analyzed by Southern blotting using methyl-sensitive enzyme digestion. In parallel, all cases were analyzed by CGH. A strong correlation between hypomethylated Sat2 sequences and 1q copy gain with a 1q12 breakpoint was found (P < 0.001). We postulate that such hypomethylation alters the interaction between the CpG-rich satellite DNA and chromatin proteins, resulting in heterochromatin decondensation, breakage and aberrant 1q formation.

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“…In our effort to transfect candidate BACs into mouse ES cells for the functional complementation screen of Ssm1, a mouse gene responsible for strain-specific transgene methylation (Engler et al 1991), we explored various strategies of BAC retrofitting. We decided to pursue strategies based on cre/loxP site-specific recombination because of its precise specificity.…”

Section: Rationale and The Structure Of Pretroesmentioning

confidence: 99%

An Efficient Method for High-Fidelity BAC/PAC Retrofitting with a Selectable Marker for Mammalian Cell Transfection

Wang¹,

Engler²,

Longacre³

et al. 2001

Genome Res.

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Large-scale genomic sequencing projects have provided DNA sequence information for many genes, but the biological functions for most of them will only be known through functional studies. Bacterial artificial chromosomes (BACs) and P1-derived artificial chromosomes (PACs) are large genomic clones stably maintained in bacteria and are very important in functional studies through transfection because of their large size and stability. Because most BAC or PAC vectors do not have a mammalian selection marker, transfecting mammalian cells with genes cloned in BACs or PACs requires the insertion into the BAC/PAC of a mammalian selectable marker. However, currently available procedures are not satisfactory in efficiency and fidelity. We describe a very simple and efficient procedure that allows one to retrofit dozens of BACs in a day with no detectable deletions or unwanted recombination. We use a BAC/PAC retrofitting vector that, on transformation into competent BAC or PAC strains, will catalyze the specific insertion of itself into BAC/PAC vectors through in vivo cre/loxP site-specific recombination.Bacterial artificial chromosomes (BACs) and P1-derived artificial chromosomes (PACs), with their capacity for large inserts, stability, and lack of chimerism, play important roles in genome mapping and sequencing (Osoegawa et al. 2000). For these reasons and the fact that virtually every gene will have one or more corresponding BAC/PAC clones, they are the first choice for functional complementation or dose-effect studies in differentiated or embryonic stem (ES) cells. Mouse ES cells are of special interest for gene function studies because of their capacity for in vitro differentiation into different lineages (O'Shea 1999) and the ability to generate transgenic animals through germline transmission. The lack of a universal selection marker for mammalian cell transfection makes it difficult to include any particular marker in the BAC/PAC vectors. Therefore, the vectors for most current BAC or PAC libraries do not contain any mammalian selection marker for transfection studies. This necessitates the development of efficient ways to insert such selection markers into individual clones chosen from BAC/PAC libraries, a "retrofitting" operation.Several strategies have been developed to retrofit BACs/PACs on the basis of restriction digestion, transposition, homologous recombination, or site-specific recombination. Certain rare-cutter restriction enzymes have been used to linearize both the BAC clone and a retrofitting cassette. These sticky ends were then ligated to form retrofitted BACs with the selectable marker inserted (Mejia and Monaco 1997;Hejna et al. 1998). For this strategy to work, it has to be certain that there is no such restriction site in the genomic insert, which often is not the case with large inserts. It also requires subsequent transformation of the retrofitted BAC/PAC into Escherichia coli, which is both very inefficient and deleterious to the genomic insert. Recombination-based approaches allow the direct...

show abstract

A strain-specific modifier on mouse chromosome 4 controls the methylation of independent transgene loci

Cited by 205 publications

References 27 publications

Epigenetic mechanisms underlying the imprinting of the mouse H19 gene.

Epigenetic mechanisms underlying the imprinting of the mouse H19 gene.

Hypomethylation of Chromosome 1 Heterochromatin DNA Correlates with q-Arm Copy Gain in Human Hepatocellular Carcinoma

An Efficient Method for High-Fidelity BAC/PAC Retrofitting with a Selectable Marker for Mammalian Cell Transfection

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