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A recent study led by the NYU Grossman School of Medicine has introduced a novel technique, Long Range Cleavage sequencing (LORAX-seq), that precisely identifies the occurrence and extent of a molecular process known as "backtracking" across the genomes of various species. Published in Molecular Cell, this research provides compelling evidence that backtracking is a prevalent regulatory mechanism affecting numerous human genes, including those essential for cellular division and embryonic development.
Backtracking occurs when RNA polymerase II, a protein complex responsible for transcribing DNA into RNA, slips backward on the DNA strand it is reading. This phenomenon, first described in 1997 by Evgeny Nudler, Ph.D., and colleagues, can lead to transcriptional delays and necessitates repair mechanisms to prevent DNA damage. The current study builds on this foundation by employing LORAX-seq to uncover that backtracking events are more widespread and enduring than previously recognized, suggesting a significant role in gene regulation beyond DNA repair.
According to Nudler, the study's senior author, "The surprising stability of backtracking at longer distances makes it likely that it represents a ubiquitous form of genetic regulation in species from bacteria to humans." He further posits that expanding these findings could position backtracking as a fundamental regulatory mechanism, akin to epigenetics, which governs gene expression without altering the DNA sequence itself.
The research delves into the mechanics of RNA polymerase II's function, highlighting how it transcribes DNA into RNA. Under specific conditions, the polymerase backtracks, leading to the extrusion of the newly synthesized RNA segment from its active site. This backtracking, if prolonged, could result in transcriptional hindrances. The study illuminates how transcription factor IIS (TFIIS) assists in resolving these blockages by cleaving the backtracked RNA, thereby allowing transcription to resume.
The study team utilized a high concentration of purified TFIIS to precisely excise backtracked RNA from any location within the cell's genome, making these segments accessible for sequence analysis. This approach revealed that genes associated with histone regulation, which play a critical role in chromatin structure and gene expression, are particularly susceptible to persistent backtracking. The researchers suggest that the extent of backtracking on these genes could influence the timing of histone production during cell division and affect the transcription of developmentally important genes.
Kevin Yang, the study's first author, notes the dual nature of backtracking, stating, "Along with its potentially useful functions, persistent backtracking could also result in DNA damage and other genetic malfunctions that contribute to disease." The team speculates that analyzing backtracking in the context of aging or cancer could provide insights into cellular malfunctions and inform new therapeutic strategies. This study not only advances our understanding of backtracking as a regulatory mechanism but also opens avenues for further research into its implications in various biological processes and diseases.
Original Publication:
Yang, K. B., Rasouly, A., Epshtein, V., Martinez, C., Nguyen, T., Shamovsky, I., & Nudler, E. (2024). Persistence of backtracking by human RNA polymerase II. Molecular Cell.
Backtracking occurs when RNA polymerase II, a protein complex responsible for transcribing DNA into RNA, slips backward on the DNA strand it is reading. This phenomenon, first described in 1997 by Evgeny Nudler, Ph.D., and colleagues, can lead to transcriptional delays and necessitates repair mechanisms to prevent DNA damage. The current study builds on this foundation by employing LORAX-seq to uncover that backtracking events are more widespread and enduring than previously recognized, suggesting a significant role in gene regulation beyond DNA repair.
According to Nudler, the study's senior author, "The surprising stability of backtracking at longer distances makes it likely that it represents a ubiquitous form of genetic regulation in species from bacteria to humans." He further posits that expanding these findings could position backtracking as a fundamental regulatory mechanism, akin to epigenetics, which governs gene expression without altering the DNA sequence itself.
The research delves into the mechanics of RNA polymerase II's function, highlighting how it transcribes DNA into RNA. Under specific conditions, the polymerase backtracks, leading to the extrusion of the newly synthesized RNA segment from its active site. This backtracking, if prolonged, could result in transcriptional hindrances. The study illuminates how transcription factor IIS (TFIIS) assists in resolving these blockages by cleaving the backtracked RNA, thereby allowing transcription to resume.
The study team utilized a high concentration of purified TFIIS to precisely excise backtracked RNA from any location within the cell's genome, making these segments accessible for sequence analysis. This approach revealed that genes associated with histone regulation, which play a critical role in chromatin structure and gene expression, are particularly susceptible to persistent backtracking. The researchers suggest that the extent of backtracking on these genes could influence the timing of histone production during cell division and affect the transcription of developmentally important genes.
Kevin Yang, the study's first author, notes the dual nature of backtracking, stating, "Along with its potentially useful functions, persistent backtracking could also result in DNA damage and other genetic malfunctions that contribute to disease." The team speculates that analyzing backtracking in the context of aging or cancer could provide insights into cellular malfunctions and inform new therapeutic strategies. This study not only advances our understanding of backtracking as a regulatory mechanism but also opens avenues for further research into its implications in various biological processes and diseases.
Original Publication:
Yang, K. B., Rasouly, A., Epshtein, V., Martinez, C., Nguyen, T., Shamovsky, I., & Nudler, E. (2024). Persistence of backtracking by human RNA polymerase II. Molecular Cell.