Germ-line transmission and expression of a human-derived yeast artificial chromosome

Jakobovits, Aya; Moore, Amy; Green, Larry L.; Vergara, German J.; Maynard-Currie, Catherine E.; Austin, Harry A.; Klapholz, Sue

doi:10.1038/362255a0

Cited by 146 publications

(44 citation statements)

References 19 publications

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“…Position-independent expression has rarely been achieved with conventional transgenes (35). However, among the four recent reports of transgenic mice carrying YAC (21)(22)(23)(24), two (21,22) have shown position-independent transgene expression, and three (21)(22)(23) Although the levels of mouse and human APP770 transcripts were comparable in the three ES cell lines the level of human APP751 transcript was slightly higher than the level of mouse APP751 transcript. This difference may reflect either a bias for processing of the primary human APP transcript toward the APP751 splice product or a difference in stability between mouse and human APP751 mRNAs.…”

Section: Methodsmentioning

confidence: 92%

“…Recently, methods for the introduction of yeast artificial chromosomes (YACs) into mice have been developed (21)(22)(23)(24). Because genomic-DNA-based transgenes permit expression of alternatively spliced transcripts and are more likely to be appropriately regulated than cDNA transgenes (25), we have introduced a 650-kb YAC containing the entire human APP gene into embryonic stem (ES) cells by the colipofection procedure (24).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Expression of the human beta-amyloid precursor protein gene from a yeast artificial chromosome in transgenic mice.

Pearson¹,

Choi²

1993

Proc. Natl. Acad. Sci. U.S.A.

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One hallmark of Alzheimer disease is the formation in the brain of amyloid plaques containing a small peptide derived from the fi-amyloid precursor protein (APP).The APP gene exhibits a complex pattern of expression in peripheral tissues and in the brain. The entire human APP gene was introduced into embryonic stem (ES) cells by co-lipofection of a 650-kb yeast artificial chromosome (YAC). Three ES lines containing an essentiafly intact YAC were isolated, and expression of human APP mRNAs at levels comparable to those of endogenous mouse APP transcripts was obtained. A transgenic mouse line was established by germ-line transmission of the APP YAC. RNase protection analysis of human APP mRNAs demonstrated appropriate splicing of the primary APP transcript in ES cells and in the brain of a transgenic animal. These mice may be useful for elucidating the function of the various APP isoforms in vivo.The senile plaques characteristic of Alzheimer disease (AD) consist of abnormal neurites around an amyloid core primarily composed of a 4-kDa polypeptide, /3-amyloid. The 3-amyloid peptide is derived from the cellular processing of ,B-amyloid precursor protein (APP). Mutations in the APP gene have been identified in certain cases of familial AD, suggesting a causal role for ,B-amyloid deposition in at least some forms of AD (1, 2). The human APP gene is located on chromosome 21, and Down syndrome (DS) individuals overexpress the wild-type APP protein (3). Those who survive past their thirties invariably develop an AD neuropathology (4), and immunoreactive f-amyloid depositions can be found prior to any detectable neuronal degeneration in the DS brain (5). Although APP is expressed at normal levels in non-DS cases of AD, the phenotype seen in DS individuals suggests that AD may be initiated or accelerated by overexpression of APP.The APP gene is composed of 18 exons distributed over several hundred kilobases (6). Alternative splicing of the primary transcript results in the production of at least five distinct transcripts. Of the three predominant APP isoforms, APP770 and APP751 contain a Kunitz-type protease inhibitor (KPI) domain, and are found at varying levels in many tissues (7,8). APP695, lacking the KPI domain, is the predominant splice form in the brain and is less abundant in the periphery (7). Other APP variants have been detected at lower levels (9-11). A possible role for the various isoforms in the progression of AD is suggested by changes in AD brains of the ratio of the KPI-containing APP variants to . However, it is clear that the APP gene exhibits complex tissue-and cell type-specific expression in human (10) and mouse (16) brain.The lack of a small-animal model for AD has hampered investigation of the early events in the progression of AD.

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Section: Methodsmentioning

confidence: 92%

mentioning

confidence: 99%

Expression of the human beta-amyloid precursor protein gene from a yeast artificial chromosome in transgenic mice.

Pearson¹,

Choi²

1993

Proc. Natl. Acad. Sci. U.S.A.

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“…Several studies have demonstrated efficient transgene expression through the transfer of YAC DNA into ES cells, suggesting that YACs can act as independent transcriptional units (21,36,54,61,65). Of particular relevance have been transgenic studies using YACs containing the human ␤-globin locus to examine the role of the LCR in controlling globin gene switching.…”

Section: Discussionmentioning

confidence: 99%

“…As an alternative approach, one may consider unbiased or "top-down" strategies to cis element dissection in which large genomic fragments are utilized at the outset in an effort to recapitulate correct developmental and quantitative gene expression. Such approaches are based on the demonstration that the stable transfer of yeast artificial chromosomes (YACs) containing large contiguous genomic DNA fragments into mammalian cells leads to proper transgene expression (36,61,65). In principle, the combined use of manipulation of YACs in yeast and transfer of YACs into embryonic stem (ES) cells provides a powerful strategy with which to investigate and identify novel distant regulatory domains within a gene locus.…”

mentioning

confidence: 99%

Regulation of the Myeloid-Cell-Expressed Human gp91-phox Gene As Studied by Transfer of Yeast Artificial Chromosome Clones into Embryonic Stem Cells: Suppression of a Variegated Cellular Pattern of Expression Requires a Full Complement of Distant cis Elements

Lien

Lee

Orkin

1997

Molecular and Cellular Biology

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“…We set out to determine whether we could recapitulate the transcriptional state of the maternal chromosome using a yeast artificial chromosome (YAC) that contains the two genes as well as the tightly linked Ins2 gene. YACs have proven useful in the analysis of large gene complexes in mammals, as they can be readily modified in yeast and then transferred into either tissue culture cells or mice (26,27,37,46,52). As Saccharomyces cerevisiae lacks the CpG methylation found in mammals, the YACs were expected to mimic the hypomethylated state of the maternal chromosome.…”

mentioning

confidence: 99%

The Absence of Enhancer Competition betweenIgf2 and H19 following Transfer into Differentiated Cells

Webber

Tilghman

1998

Molecular and Cellular Biology

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H19 and Igf2 are reciprocally imprinted genes that lie 90 kb apart on mouse chromosome 7. The two genes are coexpressed during development, with the H19 gene expressed exclusively from the maternal chromosome and Igf2 from the paternal chromosome. It has been proposed that their reciprocal imprinting is governed by a competition between the genes for a common set of enhancers. The competition on the paternal chromosome is influenced by extensive allele-specific methylation of the H19 gene and its 5 flank, which acts to inhibit H19 transcription and thus indirectly leads to the activation of the Igf2 gene. In contrast, no allele-specific methylation has been detected on the maternal chromosome, and the basis for the preference for H19 transcription on that chromosome is unresolved. In this investigation, the mechanism controlling the silencing of the Igf2 gene on the maternal chromosome was explored by studying the transcriptional activity of a yeast artificial chromosome (YAC) containing Igf2 and H19 following transfer into differentiated tissue culture cells. Contrary to expectations, both H19 and Igf2 were expressed from a single integrated copy of the YAC. Furthermore, Igf2 expression appeared to be independent of the H19 locus, based on deletions of the H19 gene promoter and its enhancers. These results suggest that an active process is responsible for the transcriptional bias toward H19 on the maternal chromosome and that the hypomethylated state of this chromosome cannot be viewed as a "default" state. Moreover, the active process is not reproduced in a differentiated cell and may require passage through the female germ line.

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Germ-line transmission and expression of a human-derived yeast artificial chromosome

Cited by 146 publications

References 19 publications

Expression of the human beta-amyloid precursor protein gene from a yeast artificial chromosome in transgenic mice.

Expression of the human beta-amyloid precursor protein gene from a yeast artificial chromosome in transgenic mice.

Regulation of the Myeloid-Cell-Expressed Human gp91-phox Gene As Studied by Transfer of Yeast Artificial Chromosome Clones into Embryonic Stem Cells: Suppression of a Variegated Cellular Pattern of Expression Requires a Full Complement of Distant cis Elements

The Absence of Enhancer Competition betweenIgf2 and H19 following Transfer into Differentiated Cells

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