Tissue- and age-specific DNA replication patterns at the CTG/CAG-expanded human myotonic dystrophy type 1 locus

Abstract

Myotonic dystrophy, caused by DM1 CTG/CAG repeat expansions, shows varying instability levels between tissues and across ages within patients. We determined DNA replication profiles at the DM1 locus in patient fibroblasts and tissues from DM1 transgenic mice of various ages showing different instability. In patient cells, the repeat is flanked by two replication origins demarcated by CTCF sites, with replication diminished at the expansion. In mice, the expansion replicated from only the downstream origin (CAG as lagging template). In testes from mice of three different ages, replication toward the repeat paused at the earliest age and was relieved at later ages-coinciding with increased instability. Brain, pancreas and thymus replication varied with CpG methylation at DM1 CTCF sites. CTCF sites between progressing forks and repeats reduced replication depending on chromatin. Thus, varying replication progression may affect tissue- and age-specific repeat instability.

Replication stalling and heteroduplex formation within CAG/CTG trinucleotide repeats by mismatch repair

“…Mapping of replication origins by nascent strand analysis in a mice model of myotonic dystrophy, showed that the expanded triplet repeat was replicated in the CAG orientation [63]. When the level of Msh2 or Msh3 was reduced in these same mice, expansions were reduced too.…”

Altered Replication in Human Cells Promotes DMPK (CTG)n · (CAG)n Repeat Instability

“…Transgenic and knock-in mouse models that recapitulate the expansion of the DMPK TNRs showed that the integration site and the amount of flanking human genomic sequence affect TNR instability (19,28,35,77,79). These findings suggest that cis-acting elements may play a crucial role in determining TNR stability during normal differentiation or pathological states.…”

“…Relevant to studies of the relationship between the DNA replication and (CAG) n · (CTG) n stability is a comparison of origin activity at the DMPK locus in human control and DM1 fetal fibroblasts and transgenic mice (19). In this work, two origins were identified, upstream and downstream of the DMPK (CAG) n · (CTG) n repeats, in control and DM1 human fibroblasts.…”

“…Recently, several DNA replication origins have been mapped close to the genomic sites of expandable microsatellite repeats at the FMR1 (9,37), HD, SCA7, SBMA (69), FMR2 (11), SCA10 (55), and DMPK (19) loci, which may reflect evolutionary selection for the position of DNA replication origins relative to microsatellite repeat tracts. At three disease loci, SCA7, HD, and SBMA, the results of origin mapping suggested that the unexpanded (CAG) · (CTG) sequence is replicated from a downstream initiation region (relative to the direction of transcription) at the wild-type SCA7 locus, from an upstream initiation region at the wild-type HD locus, and from either upstream or downstream initiation regions at the wild-type SMBA locus, leading to the conclusion that a simple relationship does not exist between (CAG) n · (CTG) n replication polarity and TNR instability when these loci are unaffected (69).…”

DNA Methylation and Trinucleotide Repeat Expansion Diseases

“…However, no effect of Dnmt1 deficit was seen on somatic instability (Dion et al, 2008). Another study, which investigated somatic instability of the DM1 CTG repeat in relation to replication and CTCF binding, has led to the suggestion that CpG methylation may regulate, in a tissue-specific manner, the role of CTCF in DNA replication and thereby CTG repeat instability (Cleary et al, 2010). Yet another connection between DNA methylation, CTCF and TNR expansion has been identified in studies of SCA7 transgenic mice, which revealed further destabilization of unstable expanded CAG repeats by CpG methylation of CTCF binding sites (Libby et al, 2008).…”