Arabidopsis SIB1: its role in chloroplast-nucleus communication during the immune response and regulation of its expression by its NAD-capped RNAs

A growing body of evidence indicates that chloroplasts play critical roles in sensing pathogen infection and other stresses and in relaying the information to the nucleus for activation of an appropriate immune response. SIGMA FACTORs (SIGs) are proteins that mediate initiation of transcription in chloroplasts. SIB1 was initially identified by its interaction with SIG1 in Arabidopsis chloroplasts. We previously found that SIB1 functions as a positive regulator of the plant immune response. Another group later found that SIB1 is also localized in the nucleus, where it acts as a co-activator of the transcription factor WRKY33 to enhance expression of defense-related genes. These findings indicate that SIB1 might link chloroplast-nucleus communication with the defense response by coordinating expression of both chloroplast and nuclear genes, but the underlying mechanism remains to be defined.

Intriguingly, the SIB1 gene is also among the limited number of Arabidopsis genes that are subjected to a non-canonical capping of its transcripts. Eukaryotic mRNAs generally contain a methyl guanosine cap (m7G cap) at their 5 ́ ends. In recent years, some RNAs, in both prokaryotic and eukaryotic cells, were found to contain a non-canonical 5’ cap - the NAD cap. We developed a method termed NADtag-Seq, which is based on a click chemistry reaction to tag NAD-capped RNAs (NAD-RNAs) followed by direct RNA sequencing using the Oxford Nanopore sequencing platform, for genome-wide identification of NAD-RNAs. We found that NAD-RNAs are produced selectively from several hundred genes in Arabidopsis. Most NAD-RNAs in plants are produced from nuclear-encoded genes that produce proteins that function in the chloroplast. SIB1 is one Arabidopsis gene that produces a high proportion of RNAs capped with NAD amongst its transcript pool. Our recent findings suggest that NAD-RNAs in plants might be involved in regulating gene expression through chloroplast-nucleus cross-talk. We will use SIB1 as an example to analyze the mode of action of NAD-RNAs in controlling gene expression and study how SIB1 regulates chloroplast-nucleus communication during the immune response. We will determine whether SIB1’s NAD- RNAs can be translated and how changes in SIB1’s NAD-RNA levels might affect SIB1 expression and the immune response. This study will not only define the role of SIB1 in plant immunity but also further the general understanding of the molecular function of NAD-RNAs