Newly Discovered Technique Uncovers Additional Layer of Human Gene Regulation
A technique has been developed that can, for the first time, determine how frequently and exactly where a molecular event known as “backtracking” occurs across the genetic material (genome) of any species, according to a new study.
Published online on February 9 in Molecular Cell, the study results provide support for the theory that backtracking represents a widespread form of gene regulation. This form of regulation influences thousands of human genes, including many involved in basic life processes like cell division and development in the womb.
The study conducted by researchers at NYU Grossman School of Medicine revolves around genes, the stretches of DNA molecular “letters” arranged in a certain sequence to encode the blueprints for most organisms. In both humans and bacteria, the first step in a gene’s expression, transcription, occurs as a protein “machine” called RNA polymerase II reads genetic instructions in one direction while ticking down the DNA chain.
The study was led by researchers from NYU Grossman School of Medicine. The study’s results suggest that backtracking represents a widespread form of gene regulation, which influences thousands of human genes, including many involved in basic life processes like cell division and development in the womb.
For a long time, studies have shown that backtracking occasionally takes place in living cells soon after RNA polymerase begins RNA synthesis or when it encounters damaged DNA to make room for incoming repair enzymes. Subsequent work also suggested that the backsliding and repair machinery had to work quickly and dissipate, or it might collide with DNA polymerase to cause cell-death-inducing breaks in DNA chains.
Now a team led by Nudler’s team at NYU Langone Health reveals that their new technique, Long Range Cleavage sequencing (LORAX-seq), can directly detect where backtracking events begin and end. This method reveals that many such events move backward further than previously thought and also last longer. The results also suggest that persistent backtracking occurs frequently throughout genomes, happens more often near certain gene types, and has functions well beyond DNA repair.
RNA polymerase II translates DNA code into a related material called RNA, which then directs the building of the proteins. Past studies have shown that as RNA polymerase II backtracks, it forces out (extrudes) from its interior channel tip of the RNA chain it has been building based on the DNA code. As prolonged backtracking is prone to causing detrimental collisions, transcription is thought to be quickly restored by the transcription factor IIS (TFIIS), which promotes the cutting off (cleavage) of the extruded, “backtracked” RNA. This clears the way for RNA polymerase II to resume its forward code reading.
Other studies have shown that when polymerase backtracks beyond a certain distance (e.g. 20 nucleobase DNA building blocks), the backtracked RNA can attach to the channel through which it is extruded, holding it in place longer. Locked, backtracked complexes are less likely to be rescued by TFIIS-driven cleavage and more likely to delay transcription of the gene involved. This led to theory that backtracking, in addition to playing a key role in DNA repair pathways, may dial up or down the action of genes as a major regulatory mechanism.
The researchers found that the genes that control histones are highly prone to persistent backtracking, which may control the timing of large-scale histone accumulation needed during cell division to rebuild chromatin. They also suggest that persistent backtracking may influence the timely transcription of genes vital to tissue development.
The study authors suggest that the measurement of backtracking in the context of aging or cancer may help understand malfunctions that occur in the cell stress response and cell replication. This may lead to new treatment approaches.
The researchers involved in the study from the Department of Biochemistry and Molecular Pharmacology at NYU Langone Health included Aviram Rasouly, Vitaly Epshtein, Criseyda Martinez, Thao Nguyen, and Ilya Shamovsky. The study is funded by the Blavatnik Family Foundation, Howard Hughes Medical Institute, and National Institutes of Health grants R01GM126891 and T32 AI007180.