Unique Roles for ISE2 in Chloroplast RNA Metabolism and Regulation of Plasmodesmata-mediated Intercellular Trafficking

INCREASED SIZE EXCLUSION LIMIT 2 (ISE2) is a nuclear gene encoding a chloroplast-localized RNA helicase that is essential for Arabidopsis thaliana embryogenesis, chloroplast RNA metabolic events and the regulation of plasmodesmal permeability. Here I report that ISE2 is essential for the editing of several chloroplast transcripts.

Emb175/PPR103 is a nuclear gene encoding a pentatricopeptide repeat (PPR) protein that was previously reported to be required for embryogenesis in Arabidopsis thaliana and for seedling survival in Zea mays. EMB175/PPR103 was previously identified in our lab in a yeast-two-hybrid interaction screen with ISE2 and subsequently named ISE2 PROTEIN INTERACTOR (IPI)1. Confocal fluorescence microscopy illustrates that IPI1-YFP, similar to ISE2-YFP, localizes to chloroplasts, consistent with its predicted chloroplast N-terminal targeting sequence. In Nicotiana benthamiana, silencing of emb175/PPR103/IPI1 in mature leaf tissue produces a chlorotic phenotype coupled to defective chloroplast structural integrity. Interestingly, virus induced gene silencing (VIGS) of N. benthamiana emb175/PPR103/IPI1 or N. benthamiana ISE2 revealed defects in the RNA editing of N. benthamiana chloroplast transcripts. However, ISE2-silenced plants displayed increased plasmodesmata-mediated intercellular trafficking, whereas no intercellular trafficking defect was observed in N. benthamiana plants silenced for emb175/PPR103/IPI1. These results indicate that ISE2 performs unique functions in the regulation of PD permeability.

Collectively, our results identify IPI1 as an ISE2 interacting protein that localizes to the chloroplast and that participates in the proper RNA editing of select N. benthamiana chloroplast transcripts. These observations add to the rapidly growing knowledge base of RNA helicase and PPR protein function in plants.