Replication dependent and independent mechanisms of GAA repeat instability

Abstract

Trinucleotide repeat instability is a driver of human disease. Large expansions of (GAA)_n repeats in the first intron of the FXN gene are the cause Friedreich's ataxia (FRDA), a progressive degenerative disorder which cannot yet be prevented or treated. (GAA)_n repeat instability arises during both replication-dependent processes, such as cell division and intergenerational transmission, as well as in terminally differentiated somatic tissues. Here, we provide a brief historical overview on the discovery of (GAA)_n repeat expansions and their association to FRDA, followed by recent advances in the identification of triplex H-DNA formation and replication fork stalling. The main body of this review focuses on the last decade of progress in understanding the mechanism of (GAA)_n repeat instability during DNA replication and/or DNA repair. We propose that the discovery of additional mechanisms of (GAA)_n repeat instability can be achieved via both comparative approaches to other repeat expansion diseases and genome-wide association studies. Finally, we discuss the advances towards FRDA prevention or amelioration that specifically target (GAA)_n repeat expansions.

Keywords: DNA triplex; Friedreich’s ataxia; GAA repeat instability; H-DNA; Replication fork stalling; Trinucleotide repeats.

Figure 1:

Various types of triplex DNA structures formed by long (GAA)_n repeats. Each H-DNA conformation, YRY or RRY, can exists in two isoforms, depending on whether the 3’ or the 5’ of a strand is donated to the triplex. Homopurine strands are in orange, homopyrimidine strands are in blue. Dashes indicate Watson-Crick base-pairing, circles indicate Hoogsteen or reverse Hoogsteen base-pairing, asterisks indicate protonated cytosines. See Section 2 for more detail.

Figure 2:

Methods to detect in vivo formation of H-DNA in higher eukaryotes (see section 2). (A) Genome-wide H-DNA formation in cultured mouse B-cells (adapted from [43]). (B) Genome-wide H-DNA formation in primary mouse cells via S1-seq (adapted from [45]). (C) Genome-wide H-DNA formation in human cancer cell lines via S1-END Seq (adapted from [46] preprint).

Figure 3:

Models of (GAA)_n repeat instability. (A) (GAA)_n repeat instability in dividing yeast mainly occurs via replication-dependent mechanisms, such as template switching (TS) or flap ligation, leading to expansions (see 4.2) [65], [83]. Pol δ dissociation during lagging strand synthesis causes contractions (see 4.3) [64]. During transcription, formation of a triplex-stabilizing R-loop (H-loop) can trigger break-induced recombination leading to repeat expansions (see 5) [113]. (B) (GAA)_n repeat instability in non-dividing yeast is characterized by two different types of events, depending on whether the MMR machinery is functional. MMR drives incisions in H-DNA and sticky DNA structures, which are then converted into DSBs. DSB repair by HR leads to gene conversions events, while repair by NHEJ results in deletions. In MMR-deficient strains, nick repair leads to expansions (see 6.1). (C) Replication dependent (GAA)_n repeat expansions in human cells is initiated by triplex formation ahead of the fork leading to its regression. Repeats can expand upon strand slippage during the restoration of the regressed fork [56] (preprint) (see 4.2). (D) (GAA)_n repeat instability in non-dividing and somatic human cells is promoted by the formation of R-loops during transcription and/or upon transcription-replication collisions (TRCs), which are then recognized by MMR and converted into DSBs. Other sources of DSBs can initiate this process as well. Error-prone repair results in (GAA)_n repeat expansions (see 6.1 and 6.2) [52], [–116].