The National Science Foundation has awarded a grant to MBL Senior Scientist Joshua Rosenthal to investigate how octopus, squid and cuttlefish use RNA editing to acclimate to temperature changes in their external environment.
Most organisms are confronted by a variable thermal environment. Temperatures can fluctuate over hours, days or seasons and the changes are particularly acute for aquatic ectotherms (cold-blooded animals, including fish, amphibians, reptiles and invertebrates) due to the high thermal conductivity of water.
On a molecular level, all enzymatic reactions are governed by temperature, but the degree of sensitivity varies. Coordinating multiple reactions across temperatures is a fundamental challenge for organisms; however, the precise mechanisms used for temperature acclimation are not well understood.
DNA is a permanent molecule, and this imposes constraints on how it can be used to encode acclimation. Having proteins optimized to different temperatures would require multiple genes that encode nearly redundant protein varients (isoforms). Messenger RNA, on the other hand, is transient, making it an excellent template for acclimation. Changing information within mRNA is comparatively economical, because multiple protein isoforms can be generated from a single gene.
All multicellular animals use RNA editing to change genetic information as it flows through RNA. Catalyzed by the ADAR enzymes, select adenosines (A) are converted to inosine (I), a biological mimic for guanosine. When this process occurs within mRNAs, it can recode genetic information, creating multiple protein products from a single gene.
Current data suggest that different species recode by A-to-I editing to different extents, but the process has been explored in very few species whose body temperature depends on the temperature of the environment (poikilotherms).
Work by Rosenthal’s team has shown that coleoid cephalopods (octopus, squid and cuttlefish) recode their brain mRNAs at rates that are orders of magnitude higher than other animals. Transcriptome-wide screens have uncovered tens of thousands of recoding sites, and these large data sets provide a unique opportunity to examine how RNA editing is used to fine-tune physiology, and how it evolves.
Preliminary data show that ~20-30% of all cephalopod edits are temperature sensitive, being edited to a higher extent in the cold. Rosenthal hypothesizes that 1) temperature sensitivity comes from the RNA structures that ADARs recognize, 2) editing sites create protein isoforms that operate more effectively in the cold, and 3) species that live in variable thermal environments have evolved editing sites to help acclimate.
To test these ideas, the team will recapitulate temperature sensitive editing in vitro, investigate how temperature sensitive editing sites affect the function of two proteins that play key roles in neurophysiology, and examine how temperature-sensitive editing sites are selected in closely related species from different thermal environments.
The research will be carried out in collaboration with Eli Eisenberg, professor of physics and astronomy at Tel Aviv University. Rosenthal and Eisenberg have had a productive collaboration since 2013, publishing several papers together including three focused on cephalopod RNA editing.
The project will enhance STEM education for underrepresented minorities, generate resources for cephalopod biology, and provide diverse community outreach. Workshops on model organism development and genome editing will be given to students from the Puerto Rico Center for Environmental Neuroscience of the University of Puerto Rico (UPR). Two UPR students will come to the MBL for one week of hands-on training in CRISPR-Cas9 gene editing, microinjection, and bioinformatics. Rosenthal and Eisenberg will also create public RNA editing databases for use by the community of cephalopod biologists.