AnN-ethyl-N-nitrosourea screen for genes involved in variegation in the mouse

Abstract

We have developed a sensitized screen to identify genes involved in gene silencing, using random N-ethyl-N-nitrosourea mutagenesis on mice carrying a variegating GFP transgene. The dominant screen has produced six mutant lines, including both suppressors and enhancers of variegation. All are semidominant and five of the six are homozygous embryonic lethal. In one case, the homozygous lethality depends on sex: homozygous females die at midgestation and display abnormal DNA methylation of the X chromosome, whereas homozygous males are viable. Linkage analysis reveals that the mutations map to unique chromosomal locations. We have studied the effect of five of the mutations on expression of an endogenous allele known to be sensitive to epigenetic state, agouti viable yellow. In all cases, there is an effect on penetrance, and in most cases, parent of origin and sex-specific effects are detected. This screen has identified genes that are involved in epigenetic reprogramming of the genome, and the behavior of the mutant lines suggests a common mechanism between X inactivation and transgene and retrotransposon silencing. Our findings raise the possibility that the presence or absence of the X chromosome in mammals affects the establishment of the epigenetic state at autosomal loci by acting as a sink for proteins involved in gene silencing. The study demonstrates the power of sensitized screens in the mouse not only for the discovery of novel genes involved in a particular process but also for the elucidation of the biology of that process.

Double SMCHD1 variants in FSHD2: the synergistic effect of two SMCHD1 variants on D4Z4 hypomethylation and disease penetrance in FSHD2

“…[33][34][35][36][37] Female homozygous MommeD1 mice, which completely lack Smchd1 protein, die at midgestation because of a failure in X inactivation. 38 Family Rf385 shows that having two SMCHD1 copies with a missense variant is viable, both in males and females. This suggests that either SMCHD1 is not essential in human or, more likely, that the variants in family Rf385 only partially abrogate SMCHD1 protein function.…”

Uncovering the uncharacterized and unexpected: Unbiased phenotype‐driven screens in the mouse

“…A genome-wide screen identified six dominant mutant lines that alter the expression of a green fluorescent protein (GFP) transgene that is normally expressed in 55% of erythrocytes (Blewitt et al, 2005). As gene silencing in mammals is not restricted to transgenes, the mutant lines affecting GFP expression were subsequently tested for their effect on retrotransposon silencing using a visible coat color marker.…”

Mechanistic concepts in X inactivation underlying dosage compensation in mammals

“…A genetic screen for epigenetic modifiers in mice has identified a gene that is specifically required for maintenance of gene repression on the Xi (Blewitt et al, 2005). The SMC-hinge domain containing protein SmcHD1 (structural maintenance of chromosomes flexible hinge domain containing 1) has been shown to be required for DNA methylation of Xi-linked gene promoters and silencing on the Xi in mice (Blewitt et al, 2008).…”