U of T researchers discover DNA-binding C2H2 zinc finger proteins also regulate RNA processing

Researchers at the University of Toronto have shown that an important class of DNA-binding factors can also bind to RNA, regulating gene expression through various mechanisms. The study significantly expands our understanding of these proteins’ functions and establishes them as a new class of RNA-binding proteins.

C2H2 zinc finger proteins are the largest group of DNA-binding factors for transcription — the process of copying genetic information from DNA to messenger RNA (mRNA). As there are more than 700 proteins in this group, most are not well understood. The few that have benefitted from focused research have been studied for their role in transcription.

“We looked at 150 members of the C2H2 zinc finger protein family,” said Syed Nabeel-Shah, first author on the study and recent PhD graduate of molecular genetics at U of T’s Donnelly Centre for Cellular and Biomolecular Research. “Although this family of proteins has been studied in the context of transcription factors, a handful of them were found to also bind to RNA around the time I began my doctoral studies. I was curious if RNA binding was a general property of these proteins — which turned out to be the case.”

The study was published recently in the journal Molecular Cell.

“We know the DNA-binding sites for around 120 of the C2H2 zinc finger proteins we studied,” said Nabeel-Shah, who received the 2024 Donnelly Centre Research Thesis Prize for his doctoral research on the RNA-binding functions of zinc finger proteins. “We suspected that, if the proteins also bind to RNA, the binding sites for DNA and RNA might be related. Generally, they were not, as the proteins targeted promoter regions of DNA and the three prime untranslated regions of mRNA.”

The researcher team found that C2H2 zinc finger proteins regulate a variety of post-transcriptional processes that modify mRNA, which serves as a template for protein production. These processes include splicing precursor mRNA to remove non-coding regions, controlling the length of mRNA through mechanisms called cleavage and polyadenylation and altering mRNA after transcription via enzymes through a process called m6A modification.

In a companion study published in Genome Biology, the team investigated how a specific zinc finger protein regulates m6A modification levels, building on their recent findings in Molecular Cell. The researchers previously found that one of the most extensively studied C2H2 zinc finger transcription factors, called SP1, regulates alternative cleavage and polyadenylation of mRNAs.

“Our team has identified a gap in knowledge around the RNA-binding capabilities of C2H2 zinc finger proteins,” said Jack Greenblatt, principal investigator on the study and professor of molecular genetics at the Donnelly Centre and the Temerty Faculty of Medicine. “Researchers have focused on DNA-binding because of its connection to disease, but there’s a lot unknown about RNA-binding by these proteins and its implications for gene expression. While we’ve mapped the RNA-binding sites for a number of C2H2 zinc finger proteins, we’re just starting to scratch the surface of investigating their functions.”

The study was the only one from Canadian researchers featured in the recent RNA-focused issue of Molecular Cell. It presents RNA-binding maps for many proteins – a valuable resource for researchers investigating the principles of post-transcriptional regulation.

This research was supported by the Canadian Institutes of Health Research.