RNA transcripts are critical subjects of regulation since they are the informational intermediaries between DNA and protein. Mutations in DNA can be repaired at the RNA level, transcripts can be stabilized by RNA binding proteins, and transcripts can be destroyed altering the expression of proteins. Discovering or engineering new tools of RNA regulation can help treat disease or improve agriculture.
The world's human population is projected to grow from 7.5 billion to over 11.2 billion by 2100. This will put pressure on a food production system reliant on fossil fuels for a majority of its nitrogen. At current levels of production over-fertilization problems have led to eutrophic waters incapable of life downstream of major agricultural fields. Improvement of crops guided by knowledge of photosynthesis might allow increased crop production with reduced environmental damage.
We are investigating RNA processing mechanisms to provide new tools to treat diseases and improve agriculture. In particularly we are interested in RNA editing. RNA editing mechanisms are critical for photosynthesis, aerobic respiration, neural transmission, lipid metabolism, and viral defense in humans and plants. RNA editing mechanisms might be rationally engineered to make specific nucleotide changes in organisms to fix problematic mutations causing disease or lead to improved functions.
C-to-U RNA editing in plants is required for photosynthetic function and is performed by a large complex of over 7 distinct nuclear encoded proteins. A highly speculative model is emerging though the functional relationships between the factors is largely unresolved. One large class of nuclear encoded proteins involved in controlling organelle expression present in most eukaryotes is the pentatricopeptide repeat (PPR) family of proteins. PPR proteins bind ssRNA through a sequence specific mechanism. PPR proteins have been linked to C-to-U RNA editing, RNA maturation, splicing, RNA turnover, and translation. Since PPR proteins have such diverse functions, we are investigating how they alter transcripts. This could lead to new tools useful for the manipulation of RNAs.
1) The editing mechanism involves a bifunctional domain called the DYW deaminase. The DYW domain binds zinc ions and has structural features common to cytidine deaminases. We are investigating the biochemical mechanisms governing shifts in activity.
2) We are currently investigating the biochemical mechanism responsible for C-to-U RNA editing in organelles. A model of the C-to-U RNA editing complex is shown below. Proteins from at least 6 different protein families have been linked to RNA editing. The biochemical roles for OZ, RIP, ISE2, and RRM proteins are unresolved and under investigation. As part of this work we are investigating the role of proteins necessary for RNA editing. Plants with reduced chloroplastic RNA editing capabilities often exhibit albino/virescent phenotypes due to photosynthetic dysfunctions. A loss of editing function mutant is shown below. This illustrates the importance of RNA editing for plant growth. The albino plants shown below have a disruption in one critcial RNA editing factor. We are exploring the biochemical function of the RNA editing factor.
3) PPRs are malleable domains that might be able to be rationally designed to target desired RNAs and perform desired modifications. We are investigating if they can be used to manipulate transcripts in vivo. Genome modifying tools like CRISPR/Cas9 genome editing have found varied applications and specifically modifying transcripts has the potential to improving health and agriculture.