Elsevier

Gene

Volume 395, Issues 1–2, 15 June 2007, Pages 125-134
Gene

Regional FMRP deficits and large repeat expansions into the full mutation range in a new Fragile X premutation mouse model

https://doi.org/10.1016/j.gene.2007.02.026Get rights and content

Abstract

Carriers of FMR1 alleles with 55–200 repeats in the 5′ UTR are at risk for Fragile X associated tremor and ataxia syndrome. The cause of the neuropathology is unknown but is thought to be RNA-mediated. Maternally transmitted premutation alleles are also at risk of expansion of the repeat tract into the “full mutation” range (> 200 repeats). The mechanism responsible for expansion is unknown. Full mutation alleles produce reduced amounts of the FMR1 gene product, FMRP, which leads to Fragile X mental retardation syndrome. We have developed a murine model for Fragile X premutation carriers that recapitulates key features seen in humans including a direct relationship between repeat number and Fmr1 mRNA levels, an inverse relationship with FMRP levels and Purkinje cell dropout that have not been seen in a previously described knock-in mouse model. In addition, these mice also show a differential deficit of FMRP in different parts of the brain that might account for symptoms of the full mutation that are seen in premutation carriers. As in humans, repeat instability is high with expansions predominating and, for the first time in a mouse model, large expansions into the full mutation range are seen that occur within a single generation. Thus, contrary to what was previously thought, mice may be good models not only for the symptoms seen in human carriers of FMR1 premutation alleles but also for understanding the mechanism responsible for repeat expansion, a phenomenon that is responsible for a number of neurological and neurodevelopmental disorders.

Introduction

The 5' UTR of the human Fragile X mental retardation 1 (FMR1) gene contains a CGG·CCG-repeat tract whose size varies from 5 to > 2000 repeats. Individuals with 55–200 repeats are said to be carriers of Fragile X premutation alleles. They are at risk for Fragile X associated tremor and ataxia syndrome (FXTAS) (Hagerman and Hagerman, 2004) and premature ovarian failure (Sherman, 2000). Premutation alleles produce elevated levels of FMR1 mRNA and this RNA is thought to have toxic effects (Jin et al., 2003, Handa et al., 2005). Premutation alleles are also prone to expand, with female carriers being at risk of having children with alleles with > 200 repeats. These carriers of so-called full mutation alleles are likely to have Fragile X mental retardation syndrome (FXS) (Verkerk et al., 1991, Yu et al., 1991). The symptoms of FXS result from a deficiency of the FMR1 gene product, Fragile X mental retardation protein (FMRP), which arises due to a combination of repeat-induced promoter silencing and difficulties in translating mRNA with large numbers of repeats (Feng et al., 1995). There is some overlap of symptoms in premutation and full mutation carriers probably because the negative effect of the repeat on translation is apparent even in the premutation range and results in a significant decrease in the level of FMRP (Kenneson et al., 2001, Primerano et al., 2002, Tassone et al., 2004).

The mechanism of expansion is not known and large expansions from a premutation-sized allele to an allele in the full mutation size range have not been previously described in mice. This has led to the idea that expansions do not occur in these animals and thus that they are not good models for studying the expansion mechanism. We describe here Knock-in (KI) mice we have generated that share key features of human premutation carriers not seen in a previous KI mouse model (Bontekoe et al., 2001, Willemsen et al., 2003). These animals also provide the first examples in mice of large repeat expansions that transform a premutation-sized allele into a full mutation sized allele in a single generation.

Section snippets

Generation of the Fragile X premutation KI mice

Mice were maintained in accordance with the guidelines of the NIDDK Animal Care and Use Committee and with the Guide for the Care and Use of Laboratory Animals (NIH publication no. 85-23, revised 1996). The outline of the strategy used to generate the targeting vector is shown in Fig. 1 and described in more detail in the figure legend. Briefly, the mouse Fmr1 gene was identified in a BAC clone (BAC 16411) derived from 129/SvJ mouse embryonic stem (ES) cells (Genome Systems, Inc, St. Louis, MO,

Generation of Fragile X premutation mice

Unlike the previous fragile X premutation mouse model which was generated by replacement of a region including the endogenous murine repeat tract with one derived from a yeast artificial chromosome (YAC) containing a human premutation allele (Bontekoe et al., 2001), the repeat tract in the premutation mice described here were generated by serial ligation of short, stable CGG·CCG-repeat tracts as previously described (Grabczyk and Usdin, 1999). The premutation-sized repeat tract was added as the

Acknowledgements

We would like to thank Lisa Garrett in the NHGRI Embryonic Stem Cell and Transgenic Core Facility, and Sonia Farmer and Maria Jorge in the NIDDK mouse facility. This research was supported in part by the Intramural Research Program of the NIDDK and NHGRI (NIH).

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  • Cited by (0)

    1

    Current address: Department of Surgery, Georgetown University, Washington DC, 20057, United States.

    2

    Current address: Department of Genetics, Louisiana State University Health Sciences Center, New Orleans, LA 70112, United States.

    3

    Current address: Department of Medicine, UCSF, San Francisco, CA 94943-0794, United States.

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