BIOLOGY OF REPRODUCTION 54, 273-278 (1996)
Genomic Imprinting in Mice: Its Function and Mechanism' Philip A. Leighton, Jennifer R. Saam, Robert S. Ingram, and Shirley M. Tilghman 2 HowardHughes Medical Institute and Departmentof Molecular Biology Princeton University, Princeton, New Jersey 08544 ABSTRACT Genomic imprinting is an epigenetic phenomenon by which the two parental alleles of a gene are differentially expressed. Although the function of genomic imprinting is not clear, it has been proposed that it evolved in mammals to regulate intrauterine growth. This proposal is consistent with experiments that were designed to reveal the mechanism and impact of genomic imprinting in a region of mouse chromosome 7 that contains four imprinted genes: Mash-2 (a transcription factor) and H19 (a noncoding RNA) are maternally expressed, whereas Insulin-2(lns-2) and Insulin-like growth factor2(lgf-2) are paternally expressed. Two targeted disruptions at the locus were generated in mice; these support the hypothesis that the function of the H19 gene is to set up the imprinting of both Igf-2 and Ins2.H19transcription on the maternal chromosome precludes transcription of the other two genes by a mechanism that involves competition for a common set of enhancers. On the paternal chromosome the H19 gene is silenced by DNA methylation, thus permitting the use of enhancers by the other genes.
INTRODUCTION Genomic imprinting is an epigenetic phenomenon by which the two parental alleles of a gene are differentially expressed [1, 2]. The existence of genomic imprinting in mice was first deduced from nuclear transplantation experiments in which diploid zygotes were generated with exclusive genetic contribution from either mothers or fathers [3, 4]. Neither kind of uniparental embryo was able to sustain normal development, and the conclusion was drawn that the lethality was due to the absence or overexpression of one or more gene products as the result of the unequal expression of genes in the maternal and paternal genomes. As of 1995 there are twelve genes known to be subject to genomic imprinting in mammals, three that are maternally expressed and nine that are paternally expressed (Table 1). However, on the basis of the mapping of imprinted regions in the mouse genome using classical genetics [5], it is evident that Table 1 contains only a subset of the imprinted genes in the genome. For most genes that have been tested, there is conservation of imprinting between mice and humans, although there is at least one exception, the Insulin-like growth factor 2 receptor (Igf-2r), which is maternally expressed in mice but biallelic in humans [6-8]. Some of the imprinted genes are organized in clusters, and two of the clusters, on mouse chromosomes 7 and 17, contain both maternally and paternally expressed genes [9, 10].
THE FUNCTION OF GENOMIC IMPRINTING With a growing list of imprinted genes in hand, the challenge is now to determine the function and mechanism of 1
Supported by a grant from the National Institutes of Health (GM51460). Correspondence: Shirley M. Tilghman, Department of Molecular Biology, Princeton University, Lewis Thomas Laboratory, Room 127, Washington Road, Princeton, NJ 08544. FAX: (609) 258-3345; e-mail:
[email protected] 2
allelic inactivation by imprinting. The function presumably must confer selective advantage to the organism, as it must counterbalance the obvious attendant risk of hemizygosity. Several explanations have been proposed for the acquisition of genomic imprinting in eutherian mammals, including protection against parthenogenesis and ovarian tumors as well as chromosome loss [11-14]. To date the most compelling model has been provided by Haig and his colleagues [15], who suggest that imprinting evolved in mammals because of the conflicting interests of maternal and paternal genes within a litter. In mammals, which are primarily nonmonogamous, the mother provides significant maternal resources to the offspring both during intrauterine life and during suckling after birth. Successful passage of paternal genes into the next generation is best ensured by having the embryos consume maternal resources, even if by so doing the fitness of her future litters is compromised. The mother's interests are best served by distributing her resources more equitably among litters. Haig's model predicts that paternally expressed genes will promote embryonic growth whereas maternal genes will act to restrain the use of maternal resources. In so doing, this model is in remarkably good agreement with the function of at least three imprinted genes: Insulin-like growthfactor2 (Igf-2), H19, and Igf-2r. Mouse embryos are exquisitely sensitive to the levels of the paternally expressed growth factor insulin-like growth factor II (IGFII). A complete loss of function of the gene encoding the growth factor, Igf-2, leads to a 40% reduction in birth weight [16]. Igf-2r, which is maternally expressed, encodes a bifunctional protein that acts as a sink for the growth factor. Fetuses that lack the receptor are approximately 30% larger than normal, have elevated circulating levels of IGFII, and die around birth [17, 18]. It has been argued that the lethality is the consequence of a gain of function of the effective 273
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LEIGHTON ET AL. TABLE 1. Chromosomal location if imprinted Gene
Expression
In mouse
In human
References
paternal maternal paternal maternal paternal paternal paternal maternal paternal paternal paternal paternal
distal 7 distal 7 distal 7 distal 7 central 7 central 7 not tested proximal 17 proximal 17 x proximal 11 proximal 6
11p15.5 11p15.5 not tested not tested 15q13-14 15q13-14 15q13-14 not imprinted not tested X not tested not tested
[16, 47 481 [28, 49, 50] [241 [22] [51, 521 R. Nicholls, personal comm. [451 16, 7, 81 191 [53, 54] [551 [561
Igf-2 H19 Insulin-2 Mash-2 Snrpn Znf-127 IPW Igf-2r Mas Xist U2afbp-rs Peg-I/Mest
.
Mash-2
IXf2
H19
Pat P at
Mat
Ins-2
Mat
_n~~n the 11tr\ ~ distnl ~t"I1 ~ nnrtinn Cu1 1 nf 1 mnlm 11 o rchrnmnnmo 1 1 11 1 1
._
_ ___________
[
FIG. 1. An imprinted region on the distal end of nnouse chromosome 7. The positions of the imprinted genes Mash-2, Ins-2, Igf- (Igf2 on figure), and H19 on mouse chromosome 7 [10, 221 are indicated. Mat, maternally expressed; Pat, paternally expressed.
concentration of circulating IGFII, orn the basis of the observation that the lethality can be abr
