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Molecular and Genetic Analysis of Plastid-to-Nucleus Signaling Pathways

We study signal transduction and gene regulation in plants, focusing on signals from plastids such as the chloroplast that regulate the expression of nuclear genes encoding proteins active in photosynthesis.

Plastids are small organelles that perform photosynthesis and a number of other essential functions in plant cells. Although plastids contain genomes that encode a small percentage of plastid-localized proteins, the overwhelming majority of plastid-localized proteins are encoded by nuclear genes. Therefore, regulation of nuclear gene expression in response to developmental and environmental cues plays an important role in plastid development and function. Extraplastidic signaling pathways that regulate nuclear gene expression do not have absolute control over plastid development and function. Plastids send signals to the nucleus that are essential for the proper expression of nuclear genes.   For example, particular chloroplast signals fine tune the expression of nuclear genes that encode components of the photosynthetic machinery to particular light environments; other chloroplast signals induce cytoplasmic antioxidant defense proteins as reactive oxygen species accumulate within the chloroplast.  Plastid signals couple the expression of nuclear genes that encode proteins active in photosynthesis to the functional state of the chloroplast and help coordinate the expression of the nuclear and chloroplast genomes.  Plastid signals are some of the most important and poorly understood signals that regulate photosynthesis-associated nuclear genes (PhANGs).  A more thorough understanding of the interactions between plastid signals and extraplastidic signaling pathways will contribute to our understanding of signal transduction networks and the regulation of photosynthesis.

We have two projects in our laboratory.  The first project aims to understand the regulation of photosynthesis-associated genes by plastid signals.  The regulation of these genes is complex, involving pathways triggered by light, the circadian clock, tissue-specific signals, carbohydrates, hormones, and plastids. The mechanisms plants use to integrate all of these signals are poorly understood.  Our recent findings indicate that plastid signals control the activity of particular light signaling pathways that regulate PhANGs and can change the response PhANGs to light quality and quantity. Plastid signals rewire light signaling pathways at least in part by converting HY5, which is a transcription factor that acts downstream of photoreceptors in light signaling pathways, from a positive to a negative regulator of PhANGs (Figure 1).  These findings indicate that a signaling network made up of plastid and light signaling pathways allows plants to integrate signals that describe particular light environments and signals that describe the functional and developmental state of the chloroplast when regulating PhANG expression.  Integrating these environmental and endogenous signals is expected to help plants more effectively regulate chloroplast biogenesis and photosynthesis.

Figure 1.  The rewiring of a blue light signaling pathway by plastid signals. (A) The regulation of PhANGs by blue light when chloroplast biogenesis and photosynthesis are optimal.  Blue light is perceived by the photoreceptor cryptochrome 1 (cry1) which in turn inhibits COP1, which is an E3 ubiquitin ligase that targets HY5 for degradation by the proteasome.  HY5 is basic-leucine zipper type transcription factor and a positive regulator of particular PhANGs when chloroplast biogenesis and photosynthesis are optimal.

Indeed, mutants with defects in this light-plastid signaling network regulate chloroplast biogenesis and function less effectively than wild type.  For example, the cotyledons of mutants with defects in plastid-to-nucleus signaling are chlorophyll deficient and become more chlorophyll deficient than wild type when light intensities are increased (Figure 2).

Figure 2. Sensitivity of plastid-to-nucleus signaling mutants to light intensity.  Wild type or the indicated mutants performed chloroplast biogenesis in 100, 500, 1000, and 1500 mmol m-2 s-1 white light.  Representative cotyledons are shown.  Representative chlorophyll deficient cotyledons that form at a low frequency in 100 mmol m-2 s-1 white light are also shown for gun1 and gun1 cry1.

Our second project is focused on chlorophyll biosynthesis.  Because accumulation of the chlorophyll precursor Mg-protoporphyrin IX within plastids is a plastid signal that regulates PhANG expression, our work on chlorophyll metabolism is related to our studies on PhANG expression.  We are studying a novel regulator of chlorophyll biosynthesis named GUN4. GUN4 participates in a plastid-to-nucleus signaling pathway that is triggered by Mg-protoporphyrin IX and exhibits no sequence similarity to proteins with known functions.  In addition to affecting plastid-to-nucleus signaling pathways, GUN4 is a major regulator of chlorophyll biosynthesis. GUN4 regulates chlorophyll metabolism by stimulating Mg-chelatase, the enzyme that commits protoporphyrin IX to chlorophyll biosynthesis.  GUN4 helps Mg-chelatase utilize its protoporphyrin IX substrate more effectively and facilitates the release of Mg-protoporphyrin IX from Mg-chelatase using a mechanism that involves binding to GUN5 and to the substrate and product of Mg-chelatase (i.e., protoporphyrin IX and Mg-protoporphyrin IX) (Figure 3).

Figure 3. The novel GUN4 Core domain fold is a porphyrin-binding domain that resembles an all helical hand in a cupped posture. Our studies indicate that GUN4 stimulates Mg-chelatase and binds porphyrins before and after metal insertion by Mg-chelatase. The figure shows the GUN4 Core domain from a 1.78-Å crystal structure of a GUN4 relative from Synechocystis and a porphyrin modeled into the palm region of the GUN4 Core domain. The porphyrin-binding site shown in the model is supported by our structure-function studies. (Figure drawn by Mark Verdecia.)

Selected Publications

Ruckle ME, Demarco SM, Larkin RM (2007) Plastid signals remodel light signaling networks and are essential for efficient chloroplast biogenesis in Arabidopsis. Plant Cell 19: 3944-3960 Abstract

Verdecia MA, Larkin RM, Ferrer JL, Riek R, Chory J, Noel JP (2005) Structure of the Mg-chelatase cofactor GUN4 reveals a novel hand-shaped fold for porphyrin binding. PLoS Biol. 3(5):e151 DOI: 10.1371/journal.pbio.0030151. Abstract

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