2Konstantinov St.-Petersburg Nuclear Physics Institute of National Research Center “Kurchatov Institute”, 188300 Gatchina, Leningrad Region, Russia
3Department of Plant Physiology, Faculty of Biology, Lomonosov Moscow State University, 119234 Moscow, Russia
4NanoTemper Technologies Rus, 191167 Saint Petersburg, Russia
5Peter the Great Saint Petersburg Polytechnic University, 195251 Saint Petersburg, Russia
6National Research Center “Kurchatov Institute”, 123182 Moscow, Russia
7Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
* To whom correspondence should be addressed.
Received April 12, 2021; Revised July 9, 2021; Accepted July 9, 2021
Serpins constitute a large family of protease inhibitors with regulatory functions found in all living organisms. Most plant serpins have not been functionally characterized, with the exception of Arabidopsis thaliana AtSerpin1, an inhibitor of pro-apoptotic proteases, which is involved in the regulation of the programmed cell death induction, and Cucurbita maxima CmPS1, a phloem protein, which presumably inhibits insect digestive proteases and binds RNA. CmPS1 interacts most efficiently with highly structured RNA; in particular, it forms a specific complex with tRNA. Here, we demonstrated that AtSerpin1 also forms a complex with tRNA. Analysis of tRNA species bound by AtSerpin1 and CmPS1 in the presence of tRNA excess revealed that both proteins have no strict selectivity for individual tRNAs, suggesting specific interaction of AtSerpin1 and CmPS1 proteins with elements of the secondary/tertiary structure universal for all tRNAs. Analysis of CmPS1 binding of the microRNA precursor pre-miR390 and its mutants demonstrated that the pre-miR390 mutant with a perfect duplex in the hairpin stem lost the ability to form a discrete complex with CmPS1, whereas another variant of pre-miR390 with the native unpaired nucleotide residues in the stem retained this ability. These data indicate that specific interactions of plant serpins with structured RNA are based on the recognition of structurally unique spatial motifs formed with the participation of unpaired nucleotide residues in the RNA duplexes.
KEY WORDS: serpin, RNA binding, RNA-binding protein, tRNA, microRNA, RNA structure, phloem