UC Berkeley

Plant cells transport molecules through small channels called plasmodesmata (PD); these nanoscopic channels are ~30-50 nm in diameter and cross through cellulosic walls to connect neighboring cells. PD regulate the movement of key developmental factors (e.g. transcription factors, small RNAs, and hormones) from cell to cell, which influence the formation of new organs, cell types, embryos, and the meristem. Mutants that affect PD transport were identified indirectly, in screens for defective developmental processes (e.g. stomata formation or root development), and in screens directed at finding PD transport mutants.

The genes identified in mutant screens for altered PD transport encode proteins that do not localize to PD. These genes that affect PD transport include INCREASED SIZE EXCLUSION LIMIT (ISE) 1 and ISE2, which encode RNA helicase proteins that localize to mitochondria and plastids, respectively; GFP ARRESTED TRAFFICKING 1 (GAT1), which encodes a plastid-localized thioredoxin; CHAPERONIN CONTAINING TCP1 8 (CCT8), which encodes a cytoplasmic chaperonin subunit; and DECREASED SIZE EXCLUSION LIMIT 1 (DSE1), which encodes a WD40-repeat protein found in the cytoplasm and nucleus.

Loss of DSE1 causes decreased PD transport both in Arabidopsis thaliana embryos and in the leaves of a distantly related eudicot species Nicotiana benthamiana. Through transcriptome analysis, we demonstrate here that DSE1 affects the expression of seed maturation and abscisic acid (ABA) response genes and that ABA INSENSITIVE 5 (ABI5), a key ABA transcription factor, reduces cell-cell transport. Further, dse1 embryos have reduced PD frequency, suggesting that the PD transport phenotype observed in this mutant is due to fewer PD in the cell wall.

We characterize a new PD mutant ise3 that was identified in a screen for mutants with increased PD transport in A. thaliana embryos. ISE3 encodes a SEL1-like repeat mitochondrial protein and we show that this protein interacts with an EMBRYO DEFECTIVE-mitochondrial-PPR-domain-containing protein in Bimolecular Fluorescence Complementation. Loss of the N. benthamiana orthologs of ISE3 or its interactor, ISE3 INTERACTING PROTEIN 1 (IPR1), cause increased PD transport in leaves. We also show that ISE3, like ISE1 affects the expression of genes involved in reactive oxygen species (ROS) regulation and show that loss of ISE3 or IPR1 cause increased H2O2 production. These results support findings that ROS can regulate PD transport, and further suggest that the mitochondria and the plastids signal to the nucleus (through ‘retrograde signaling’) to coordinate nuclear gene expression and PD transport, in a pathway called ‘organelle-nucleus-PD signaling’, or ONPS. This thesis supports the growing body of evidence that intercellular transport is driven by critical intracellular signals and processes.

Retrograde signalling pathways require signal exchange among the organelles and the nucleus, and a putative pathway for this exchange is through “stromules,” which are stroma-filled tubular extensions of plastids. Herein we describe the first identified signal transduction pathways initiated by the plastid that are linked to stromule formation. We show that stromule frequency is affected by light cues as well as by ROS and the redox status of the chloroplast. Finally, we demonstrate that isolated chloroplasts can make stromules, indicating that the plastid drives the formation of these tubule structures, which may be involved in intracellular signalling.