A recent study led by Charles Danko, an associate professor at Cornell University, has shed light on a significant yet previously overlooked aspect of gene regulation known as promoter-proximal pausing. This process, which involves a brief halt in gene activity, may have played a crucial role in the evolution of complex life forms from simpler cells. The findings were published in a paper co-authored by Danko and several colleagues, including John Lis, an esteemed figure in molecular biology.
Promoter-proximal pausing occurs shortly after a cell’s RNA polymerase II—the mechanism responsible for synthesizing RNA—initiates its function. The polymerase pauses after transcribing approximately 20 to 60 nucleotides, effectively waiting for further signals. According to Danko, the duration of this pause can range from one to ten minutes, which is substantial in the context of cellular processes.
The concept of promoter-proximal pausing has been known since the 1980s when John Lis first described it. Initially, research focused heavily on yeast, which did not exhibit this pausing behavior, leading many scientists to underestimate its significance. However, Lis’s postdoctoral trainee, Karen Adelman, demonstrated that such pauses exist in both fruit flies and human cells. This revelation prompted Danko and his team to explore when and how this pausing mechanism evolved without disrupting overall transcription processes across various species.
Utilizing a technique known as PRO-seq, developed in Lis’s laboratory, the research team mapped promoter-proximal pausing across a wide range of organisms, from bacteria to complex animals. Their analysis revealed that a primitive form of this pause existed in single-celled organisms. Over time, as organisms evolved, the pause became more pronounced and better regulated, primarily due to the emergence of new protein complexes such as the negative elongation factor (NELF).
Danko explained that NELF comprises four subunits that appeared at different evolutionary stages. Two core subunits were present in many eukaryotes, while the remaining two emerged later, enhancing the ability of RNA polymerase to pause and allowing for finer control over gene expression. He expressed enthusiasm about the implications of this discovery, stating, “This gives a lot of context on when these particular pausing systems evolved.”
To further investigate NELF’s role, the Cornell researchers collaborated with the Memorial Sloan Kettering Cancer Center to deplete two of its subunits in mouse cells. The results indicated that, without these regulators, RNA polymerase advanced along genes too rapidly. Consequently, many genes failed to respond adequately to heat stress, which typically triggers the transcription of key heat shock genes. Danko noted that the absence of NELF led to a significant reduction in gene up-regulation compared to when NELF was intact.
Danko likened NELF’s function to adding knobs on a stereo, allowing cells to finely tune the “volume” of gene expression. “This control is vital for the development of multicellular organisms, enabling precise regulation of gene activity,” he said. The evolution of NELF proteins in common ancestors leading to multicellular animals supports this perspective.
Understanding promoter-proximal pausing is crucial, as disruptions in this process can be linked to various diseases, including cancer. “It’s essential to grasp what drives transcription to understand its connections with these diseases,” Danko emphasized. Without such understanding, researchers risk only identifying altered gene expressions without comprehending their underlying significance.
The research team also included faculty members from Cornell, such as Anna-Katerina Hadjantonakis, Ilana L. Brito, and John Lis. Their work received funding from the National Institutes of Health. This study not only advances knowledge in the field of gene regulation but also highlights the intricate molecular mechanisms that have evolved over billions of years, shaping the complexity of life as we know it.
