Investigators from the University of Illinois have created a whole-genome map that charts the occupancies of RNA polymerase enzymes in humans, discovering unexpected patterns in the process. Their findings appear in Molecular Cell.
DNA transcription — the process of copying sequences of DNA into RNA molecules — involves enzymes called RNA polymerase (Pol). In humans, three distinct RNA polymerase enzymes are conventionally thought to transcribe specific categories of genes; for example, Pol II generally transcribes genes encoding proteins, while Pol I and Pol III produce specific classes of functional, noncoding RNAs. Illinois investigators wanted to quantify the patterns of these polymerase enzymes, uncovering unexpected nuances in the process.
“We are asking big questions that center on basic transcriptional activities within the cell,” said Kevin Van Bortle, an Assistant Professor of Cell & Developmental Biology and senior author on the paper. “Our textbooks often describe patterns and behaviors as being cut and dry and as simple as possible, but our work opens an unexpected nuance and complexity to the canonical definitions of where these enzymes function. The rules are not as black and white as we thought.”
Historically, scientists believed that protein-coding genes were exclusively transcribed by Pol II. Unexpectedly, researchers from the Van Bortle Lab discovered that Pol III also occupies the promoter regions of many protein-coding genes, which is considered a noncanonical or unconventional pattern. Further, researchers were surprised to learn that Pol III likely initiates transcription at these sites.
“We know that it’s not transcribing the entire gene, but it appears to take a handful of steps before falling off,” said Rajendra K C, a graduate student and lead author of the paper. “This behavior is significant because that short walk appears to promote the activity of Pol II to continue building the full-length mRNA. I think we are one of the first groups to find this evidence of crosstalk between Pol II and Pol III at these sites.”
Van Bortle and K C had initially observed instances of Pol III localization at unexpected sites. This observation led the group to develop an innovative framework to globally map all three polymerases and their activity patterns. The comprehensive map was a result of significant quantitative legwork.
“It’s easy to say, ‘we saw something unexpected,’” Van Bortle said. “The hard part is proving a meaningful signal or pattern. Rajendra had to use all the current but disparate annotations of coding and non-coding genomic elements to score RNA polymerase patterns across the entire genome. The value of our work here is that it relies on a data-driven statistical approach. This whole story is truly driven by the data.”
The investigators’ findings also point to a link between noncanonical Pol III activity with genes that generally encode factors involved in biosynthesis and cell growth, many of which are implicated in cancer.
The result of the researchers’ newfound answers?
More questions.
Knowing that Pol III is active beyond its previously defined set of genes introduces queries like, how does Pol III mechanistically help Pol II?
The group’s next steps include using data to predict protein-protein and macromolecular complex-complex interactions, which could ultimately shed light on the recruitment and regulation of Pol III and offer insight into other associated proteins. Even with these advancements, they anticipate more questions.
“It’s surprising that we’re the first to see these patterns of activity,” Van Bortle said. “This is an example of not discovering things until you look for them, and there are so many things we haven’t looked for. Most of our questions don’t have answers, which means there is so much more to learn.”
