We research gene regulation by non-coding RNAs, with an emphasis on microRNA (miRNA) . miRNAs are a group of small non-coding RNAs that regulate the expression of other genes at the post-transcriptional level. Therefore, miRNAs are important for controlling human developmental, differentiation, and disease processes. In particular, we focus on miRNAs generated by alternative mechanisms, which surprisingly incorporate fundamental cellular machineries involved in processing other classes of RNAs. Our research, therefore, aims to expand the appreciation of these RNA processing machineries’ impact on small RNA populations and oncogenesis.
Canonically, miRNAs are produced from primary transcripts that are cleaved by the nuclear Microprocessor complex, with the resulting precursor (pre-)miRNA hairpins exported from the nucleus by Exportin-5, and further processed by cytoplasmic Dicer. Accumulating evidence suggests that various miRNAs are aberrantly expressed in human cancer cells, so it is important to elucidate the mechanisms by which miRNAs are expressed and how their biogenesis is regulated.
This graphic presents the canonical miRNA biogenesis pathway alongside two non-canonical pathways that are objects of study in our lab: viral miRNA biogenesis in the Herpesvirus saimiri (HVS), and the m7G-capped pre-miRNA pathway in mammals.
HVS miRNA BIOGENESIS PATHWAY
Infection by Herpesvirus saimiri (HVS) causes fatal T-cell leukemias and lymphomas in New World primates and immortalizes human T cells in vitro, making them an invaluable model for elucidating human T-cell malignancies. In HVS-transformed T cells, viral miRNAs are co-transcribed downstream of viral small nuclear (sn)RNAs. These are the first snRNA-miRNA chimeras identified. Intriguingly, the host Integrator, a 14-subunit complex that is known to process the 3′ ends of snRNAs, recognizes and processes conserved sequence elements flanking the viral pre-miRNAs, therefore replacing Microprocessor cleavage.
The search for snRNA-miRNA chimeric precursors in host cells then led to the discovery of a group of Microprocessor-independent pre-miRNAs capped with 7-methylguanosine (m7G). The 5′ ends of these m7G-capped pre-miRNAs coincide with RNA polymerase II (Pol II) transcription start sites. Due to the presence of the cap, these pre-miRNAs are exported via the PHAX-Exportin-1 pathway, instead of the canonical Exportin-5 pathway. After Dicer cleavage, only the 3p-miRNA is efficiently loaded into the Argonaute protein to form a functional microRNP (ribonucleoprotein) that guides gene repression. This finding enables the development of special shRNA expression constructs that produce a single 3p-siRNA.
Our future research will address whether such pathways are conserved in other organisms and what the advantage of using alternative pathways over the canonical pathway for generating specific miRNAs is. We will also continue to explore the mechanisms of m7G-capped pre-miRNA 3′-end formation and Integrator-mediated RNA processing.
We are grateful for the financial support our research has received:
- UF Health Cancer Center Startup fund 9/1/2016 –
- University of Florida, Office of Research Research Opportunity Seed Fund 9/1/2018 – 8/31/2021 (NCE)
Title: Processing of transcription start site (TSS-) microRNAs by Dicer proteins
- NIH/NIGMS R35 GM 128753 8/13/2018 – 7/31/2023
Title: RNA metabolism mediated by the Integrator complex
- Brown Foundation Seed Fund 2020 – 2021
Title: Novel microRNA processing pathways
- Florida Department of Health Live Like Bella Pediatric Initiative 6/8/2021 – 4/30/2024
Title: Target RNAs induce microRNA degradation in apoptotic T-cell acute lymphoblastic leukemia cells
- Thomas H. Maren Foundation F013372 4/1/2018 – 3/31/2020
Title: Junior Investigators Fund
- NIH/NCI R00 CA 190886 9/19/2014 – 2/28/2021 (NCE)
Title: Noncanonical microRNA biogenesis and function in a gamma herpesvirus and mammals