Circular RNAs

Covalently closed circular RNAs (circRNAs) are producing precursor mRNA back-splicing; the exons of thousands of genes in eukaryotes. They are generally expressing at low levels and often exhibit cell-type-specific and tissue-specific patterns. The study of circRNAs, especially their function, is challenging due to their circular conformation and sequence-overlap with linear mRNA counterparts.

In a study published in Cell, a team led by Dr. Chen Lingling at the Shanghai Institute of Biochemistry and Cell Biology of the Chinese Academy of Sciences (CAS) and Dr. Yang Li at the CAS-MPG Partner Institute for Computational Biology discovered that most examined circRNAs; tend to form 16-26 bp intramolecularly imperfect RNA duplexes.

In addition, circRNAs preferentially bind to dsRNA-activated protein kinase (PKR); which is related to innate immunity and act as PKR inhibitors. In collaboration with Dr. Shen Nan at Shanghai Renji Hospital, the study also showed that dysregulation of circRNA expression; and PKR activation is linking to the autoimmune disease systemic lupus erythematosus (SLE).

Global circRNA degradation

As circRNAs are stable in examined cells and tissues under normal conditions; the scientists sought to determine whether they could be degraded under certain types of cellular stress. They found that both poly(I:C) and encephalomyocarditis (EMCV) dsRNAs can trigger global circRNA degradation by activated RNase L upon viral infection.

Intriguingly, although both circRNAs and linear RNAs are degraded by RNase L; earlier studies by Dr. Chen’s lab showed that nascent circRNA production is rare. As a result, the amount of new circRNAs produced from pre-mRNAs cannot overcome rapid degradation by RNase L; within a short period after early innate immune responses. This results in global loss of circRNAs and the subsequent release of circRNA-associated PKR, thus allowing PKR activation.

Cytoplasmic response to dsRNA

PKR is directly activated by dsRNA and plays a central role in the cytoplasmic response to dsRNA. In order to learn how circRNAs bind to PKR and regulate its activation; the researchers optimized selective 2′-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) assays; and discovered that most examined circRNAs tend to form 16-26 bp imperfect RNA duplexes which can act as PKR inhibitors in cells.

Further examination of SLE patient-derived cells showed reduced circRNA expression; spontaneous RNase L activation and increased PKR. It also showed that over-expression of circRNAs robustly reduced the aberrant PKR activation cascade; in SLE patient-derived cells. This shows that massive and rapid degradation of circRNAs by RNase L; is required for PKR activation in the early stage of the innate immune response.