Characterization of the chloroplast retrograde signal that regulates alternative splicing

MICAELA A. GODOY HERZ; MA. GUILLERMINA KUBACZKA; EZEQUIEL PETRILLO; GASTÓN SILESS; GABRIELA CABRERA; JORGE PALERMO; ALBERTO KORNBLIHTT

Congreso; SAIB, 51th Annual Meeting, Argentine Society for Biochemistry and Molecular Biology; 2015

We have previously shown that light/dark conditions affect several alternative splicing events in Arabidopsis thaliana. Using drugs that inhibit the photosynthetic electron transport chain, we found that DCMU mimics the effect of darkness, whereas DBMIB mimics the effect of light on alternative splicing. This led us to propose the redox state of the plastoquinone pool as the switch that triggers a retrograde signal that modifies alternative splicing choices in the nucleus. Using high performance liquid chromatography we demonstrated that the redox state of the plastoquinone pool changes with our light/dark protocol. In order to investigate the nature of the retrograde signal, we have established the experimental conditions to obtain Arabidopsis mesophyll protoplasts. Similarly to Arabidopsis seedlings, protoplasts display a light/dark effect on alternative splicing. We incubated protoplast preparations in light or dark for a certain time, and then centrifuged them to collect the conditioned medium. The conditioned medium obtained from light-exposed protoplasts causes changes in alternative splicing in protoplasts kept in the dark, similar to those observed in protoplasts exposed to light. This suggests that, upon light incubation, protoplasts produce a signaling molecule that accumulates in the conditioned medium and then modulates alternative splicing.