Structure of an Intranucleosomal DNA Loop That Senses DNA Damage during Transcription

Abstract

Transcription through chromatin by RNA polymerase II (Pol II) is accompanied by the formation of small intranucleosomal DNA loops containing the enzyme (i-loops) that are involved in survival of core histones on the DNA and arrest of Pol II during the transcription of damaged DNA. However, the structures of i-loops have not been determined. Here, the structures of the intermediates formed during transcription through a nucleosome containing intact or damaged DNA were studied using biochemical approaches and electron microscopy. After RNA polymerase reaches position +24 from the nucleosomal boundary, the enzyme can backtrack to position +20, where DNA behind the enzyme recoils on the surface of the histone octamer, forming an i-loop that locks Pol II in the arrested state. Since the i-loop is formed more efficiently in the presence of SSBs positioned behind the transcribing enzyme, the loop could play a role in the transcription-coupled repair of DNA damage hidden in the chromatin structure.

Distinctive Participation of Transcription-Coupled and Global Genome Nucleotide Excision Repair of Pyrimidine Dimers in the Transcribed Strand of Yeast rRNA Genes

“…It was suggested that TC-NER overrides the impediments caused by nucleosomes in the TS, ,− although a complete understanding of the unequal modulation of NER in the two strands of nucleosomal DNA is lacking. One model indicates that transcription through a nucleosome by a single RNAPII is paired with the formation of an intra-nucleosome DNA loop. ,, Therefore, it is tempting to speculate that the NTS remains near the histone octamer surface restraining and modulating GG-NER, while the TS is looping out into the solvent where PDs can be homogeneously removed by TC-NER. The same model puts forward the possibility that transient eviction of a nucleosome during transcription elongation could depend on a local high density of RNAPII. , Comparable conditions are found along the rRNA gene coding regions that are densely loaded with RNAPI and largely devoid of nucleosomes, but they are transiently packed with nucleosomes as RNAPI are displaced from the damaged TS .…”

“…model indicates that transcription through a nucleosome by a single RNAPII is paired with the formation of an intra-nucleosome DNA loop. 22,23,47 Therefore, it is tempting to speculate that the NTS remains near the histone octamer surface restraining and modulating GG-NER, while the TS is looping out into the solvent where PDs can be homogeneously removed by TC-NER. The same model puts forward the possibility that transient eviction of a nucleosome during transcription elongation could depend on a local high density of RNAPII.…”

Structural Transition of the Nucleosome during Transcription Elongation

“…In particular, a small intra-nucleosomal DNA loop is reportedly formed more efficiently on a template containing a single-strand break in the non-template strand, than on the undamaged DNA template [118]. This finding suggested that RNAP may be arrested by template DNA looping at sites of DNA damage in chromatin [118]. The histone tails may facilitate this RNAP arrest [58].…”

“…A small DNA loop that does not affect nucleosome repositioning (called the zero-loop) has also been proposed [116,117]. To date, eukaryotic RNAPII, RNAPIII, bacteriophage SP6 RNAP, and Escherichia coli (E. coli) RNAPs have been reported to form template DNA loops in vitro [41,[113][114][115]118].…”

“…While the template DNA loop model has been proposed to function in nucleosome retention during transcription elongation, it has also been suggested that the formation of such an intra-nucleosomal loop may cause RNA polymerase arrest, probably due to the stable nature of the nucleosome with the template DNA loop [13,113,[118][119][120]. In particular, a small intra-nucleosomal DNA loop is reportedly formed more efficiently on a template containing a single-strand break in the non-template strand, than on the undamaged DNA template [118]. This finding suggested that RNAP may be arrested by template DNA looping at sites of DNA damage in chromatin [118].…”