Determinants of R-loop formation at convergent bidirectionally transcribed trinucleotide repeats.
R-loops have been described at immunoglobulin class switch sequences, prokaryotic and mitochondrial replication origins, and disease-associated (CAG)n and (GAA)n trinucleotide repeats. The determinants of trinucleotide R-loop formation are unclear. Trinucleotide repeat expansions cause diseases including DM1 (CTG)n, SCA1 (CAG)n, FRAXA (CGG)n, FRAXE (CCG)n and FRDA (GAA)n. Bidirectional convergent transcription across these disease repeats can occur. We find R-loops formed when CTG or CGG and their complementary strands CAG or CCG were transcribed; GAA transcription, but not TTC, yielded R-loops. R-loop formation was sensitive to DNA supercoiling, repeat length, insensitive to repeat interruptions, and formed by extension of RNA:DNA hybrids in the RNA polymerase. R-loops arose by transcription in one direction followed by transcription in the opposite direction, and during simultaneous convergent bidirectional transcription of the same repeat forming double R-loop structures. Since each transcribed disease repeat formed R-loops suggests they may have biological functions.
Program of Genetics & Genome Biology, The Hospital for Sick Children, 101 College Street, East Tower, 15-312 TMDT, Toronto, Ontario M5G 1L7, Canada.
Kaalak
K
Reddy
Mandy
M
Tam
Richard P
RP
Bowater
Miriam
M
Barber
Matthew
M
Tomlinson
Kerrie
K
Nichol Edamura
Yuh-Hwa
YH
Wang
Christopher E
CE
Pearson
D004247:DNA / Q000737:chemistry* / Q000648:ultrastructure; D004278:DNA, Superhelical / Q000737:chemistry; D012313:RNA / Q000737:chemistry* / Q000648:ultrastructure; D014158:Transcription, Genetic; D018911:Trinucleotide Repeats
D004278:DNA, Superhelical; D012313:RNA; D004247:DNA