Experimental and Computational Tools for Mapping Circular RNA Modifications

Posttranscriptional RNA modifications and their impact on the structure and function have been extensively studied on linear RNAs. As RNA modifications affect RNA structure, stability, localization, and translation, several methods have been developed to study modifications on linear RNAs. These modifications on RNAs ultimately regulate gene expression. One of the poorly characterized classes of RNA critical for gene expression is circular RNAs (circRNAs), which are ubiquitously expressed and are covalently closed RNA molecules. Although advances in sequencing technologies have greatly enhanced the identification of RNA modifications and their importance in gene regulation, the effects of these modifications on circRNA structure and function are only now emerging. Only a few modifications, like N⁶-methyladenosine (m⁶A), 5-methylcytosine (m⁵C), adenosine-to-inosine (A-to-I), and to a lesser extent N1-methyladenosine (m¹A), have been reported so far for circRNAs. In this article, we discuss the importance of these modifications and their implications in circRNA function. Moreover, we will discuss molecular and computational methods for detecting circRNA modifications, to help researchers studying circRNAs select the most effective methods for investigating modifications in this novel class of circRNAs.