Palindrome-Mediated and Replication-Dependent Pathogenic Structural Rearrangements within theNF1Gene

Abstract

Palindromic sequences can form hairpin structures or cruciform extrusions, which render them susceptible to genomic rearrangements. A 197-bp long palindromic AT-rich repeat (PATRR17) is located within intron 40 of the neurofibromatosis type 1 (NF1) gene (17q11.2). Through comprehensive NF1 analysis, we identified six unrelated patients with a rearrangement involving intron 40 (five deletions and one reciprocal translocation t(14;17)(q32;q11.2)). We hypothesized that PATRR17 may be involved in these rearrangements thereby causing NF1. Breakpoint cloning revealed that PATRR17 was indeed involved in all of the rearrangements. As microhomology was present at all breakpoint junctions of the deletions identified, and PATRR17 partner breakpoints were located within 7.1 kb upstream of PATRR17, fork stalling and template switching/microhomology-mediated break-induced replication was the most likely rearrangement mechanism. For the reciprocal translocation case, a 51 bp insertion at the translocation breakpoints mapped to a short sequence within PATRR17, proximal to the breakpoint, suggesting a multiple stalling and rereplication process, in contrast to previous studies indicating a purely replication-independent mechanism for PATRR-mediated translocations. In conclusion, we show evidence that PATRR17 is a hotspot for pathogenic intragenic deletions within the NF1 gene and suggest a novel replication-dependent mechanism for PATRR-mediated translocation.

Decoding NF1 Intragenic Copy-Number Variations

“…[43][44][45] Particularly, within intron 40 of NF1, a palindromic AT-rich repeat (PATRR17; 197 bp), especially the loop of the palindrome (7 bp), has been identified as an intragenic rearrangement hotspot. 41 In addition to non-B DNA structures, Alu elements were also shown in this study to be involved in recurrent genomic rearrangements. …”

“…In addition to causing deletions, serial replication stalling also leads to insertions and translocations, further supporting a significant contribution of the replication-based mechanism to genomic rearrangements. 20,41,[53][54][55] Other studies also indicate that the high level of microhomology observed at the breakpoint junctions suggests presence of a replication-based mechanism and a mitotic origin. 20,23,30,56 Given that microhomologous sequences are widely distributed in the genome, in addition to investigating sequence context, exploring the DNA spatial proximity and 3D genomic architecture close to the junction site is likely to reveal which DNA interval is susceptible to double-strand break or replication-fork collapse, as well as the accessibility of microhomologous sequences to enzymes associated with FoSTeS/ MMBIR.…”

“…41 However, in this study we identified a PATRR17-mediated deletion (UAB-68) that has no microhomology at the breakpoint-junction site, and the partner breakpoint is located in the 6.7 kb downstream of PATRR17, suggesting NHEJ, rather than FoSTeS/ MMBIR, was used to ligate the broken ends. Interestingly, UAB-68's breakpoint is also located in the loop of PATRR17, which is consistent with previously published PATRR17-mediated deletions and translocation.…”

“…In addition to playing a role in PATRR17-mediated deletion, a serial replication event is also involved in a PATRR17-mediated chromosomal translocation. 41 have mediated a recurrent NAHR, resulting in exon-2 deletion in three unrelated individuals (MUW-1, UAB-1, and UAB-2) with exactly the same breakpoints (chr17: 29,477,816-29,484,743), suggesting a strong recurrent Alu-mediated NAHR. The deletion in UAB-1 was proven to be absent in both unaffected parents; MUW-1 is from Austria and UAB-2 is from the US; therefore, we consider it unlikely that a founder effect could explain the recurrence of this deletion.…”

“…41 All of these 85 CNVs were submitted to LOVD2_NF1 (curated by Rick van Minkelen). Furthermore, we included two additional published NF1 CNVs (from IRO-1 and UNIMI-1).…”