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Stomata are natural openings through which many pathogenic bacteria enter plants. Successful bacterial pathogens have evolved various virulence factors to promote stomatal opening. Here, we show that the Pseudomonas syringae type III effector protein AvrB induces stomatal opening and enhances bacterial virulence in a manner dependent on RPM1-INTERACTING4 (RIN4), which promotes stomatal opening by positively regulating the Arabidopsis plasma membrane H+-ATPase (AHA1), which is presumed to directly regulate guard cell turgor pressure. In support of a role of AHA1 in AvrB-induced stomatal opening, AvrB enhances ATPase activity in plants. Unexpectedly, AHA1 promotes the interaction between the jasmonate (JA) receptor CORONATINE INSENSITIVE1 (COI1) and JASMONATE ZIM-DOMAIN (JAZ) proteins and enhances JA signaling. JA signaling is required for optimum stomatal infection in AHA1-active plants. Similarly, AvrB also induces the COI1-JAZ9 interaction and the degradation of multiple JAZ proteins. AvrB-induced stomatal opening and virulence require the canonical JA signaling pathway, which involves the COI1 and NAC transcription factors. The findings thus point to a previously unknown pathway exploited by P. syringae that acts upstream of COI1 to regulate JA signaling and stomatal opening. [ABSTRACT FROM AUTHOR]

To restrict pathogen entry, plants close stomata as an integral part of innate immunity. To counteract this defense, Pseudomonas syringae pv tomato produces coronatine (COR), which mimics jasmonic acid (JA), to reopen stomata for bacterial entry. It is believed that abscisic acid (ABA) plays a central role in regulating bacteria-triggered stomatal closure and that stomatal reopening requires the JA /COR pathway, but the downstream signaling events remain unclear. We studied the stomatal immunity of tomato (Solanum lycopersicum) and report here the distinct roles of two homologous NAC (for NAM, ATAF1,2, and CUC2) transcription factors, JA2 (for jasmonic acid2) and JA2L (for JA2-like), in regulating pathogen-triggered stomatal movement. ABA activates JA2 expression, and genetic manipulation of JA2 revealed its positive role in ABA -mediated stomatal closure. We show that JA2 exerts this effect by regulating the expression of an ABA biosynthetic gene. By contrast, JA and COR activate JA2L expression, and genetic manipulation of JA2L revealed its positive role in JA /COR-mediated stomatal reopening. We show that JA2L executes this effect by regulating the expression of genes involved in the metabolism of salicylic acid. Thus, these closely related NAC proteins differentially regulate pathogen-induced stomatal closure and reopening through distinct mechanisms. [ABSTRACT FROM AUTHOR]

Although the control of organ size is a fundamental question in developmental biology, little is known about the genetic and molecular mechanisms that determine the final size of seeds in plants. We previously demonstrated that the ubiquitin receptor DA1 acts synergistically with the E3 ubiquitin ligases DA2 and ENHANCER1 OF DA1 (EOD1)/BIG BROTHER to restrict seed growth in Arabidopsis thaliana. Here, we describe UBIQUITIN-SPECIFIC PROTEASE15 (UBP15), encoded by SUPPRESSOR2 OF DA1 (SOD2), which acts maternally to regulate seed size by promoting cell proliferation in the integuments of ovules and developing seeds. The sod2/ubp15 mutants form small seeds, while overexpression of UBP15 increases seed size of wild-type plants. Genetic analyses indicate that UBP15 functions antagonistically in a common pathway with DA1 to influence seed size, but does so independently of DA2 and EOD1. Further results reveal that DA1 physically associates with UBP15 in vitro and in vivo and modulates the stability of UBP15. Therefore, our findings establish a genetic and molecular framework for the regulation of seed size by four ubiquitin-related proteins DA1, DA2, EOD1, and UBP15 and suggest that they are promising targets for increasing seed size in crops. [ABSTRACT FROM AUTHOR]

Plants have evolved sophisticated mechanisms for integration of endogenous and exogenous signals to adapt to the changing environment. Both the phytohormones jasmonate (JA) and ethylene (ET) regulate plant growth, development, and defense. In addition to synergistic regulation of root hair development and resistance to necrotrophic fungi, JA and ET act antagonistically to regulate gene expression, apical hook curvature, and plant defense against insect attack. However, the molecular mechanism for such antagonism between JA and ET signaling remains unclear. Here, we demonstrate that interaction between the JA -activated transcription factor MYC2 and the ET -stabilized transcription factor ETHYLENE-INSENSITIVE3 (EIN3) modulates JA and ET signaling antagonism in Arabidopsis thaliana. MYC2 interacts with EIN3 to attenuate the transcriptional activity of EIN3 and repress ET -enhanced apical hook curvature. Conversely, EIN3 interacts with and represses MYC2 to inhibit JA -induced expression of wound-responsive genes and herbivory-inducible genes and to attenuate JA -regulated plant defense against generalist herbivores. Coordinated regulation of plant responses in both antagonistic and synergistic manners would help plants adapt to fluctuating environments. [ABSTRACT FROM AUTHOR]

Glutathione is involved in thiol redox signaling and acts as a major redox buffer against reactive oxygen species, helping to maintain a reducing environment in vivo. Glutathione reductase (GR) catalyzes the reduction of glutathione disulfide (GSSG) into reduced glutathione (GSH). The Arabidopsis thaliana genome encodes two GRs : GR1 and GR2. Whereas the cytosolic/peroxisomal GR1 is not crucial for plant development, we show here that the plastid-localized GR2 is essential for root growth and root apical meristem (RAM) maintenance. We identify a GR2 mutant, miao , that displays strong inhibition of root growth and severe defects in the RAM , with GR activity being reduced to ∼50%. miao accumulates high levels of GSSG and exhibits increased glutathione oxidation. The exogenous application of GSH or the thiol-reducing agent DTT can rescue the root phenotype of miao , demonstrating that the RAM defects in miao are triggered by glutathione oxidation. Our in silico analysis of public microarray data shows that auxin and glutathione redox signaling generally act independently at the transcriptional level. We propose that glutathione redox status is essential for RAM maintenance through both auxin/PLETHORA (PLT)-dependent and auxin/PLT-independent redox signaling pathways. [ABSTRACT FROM AUTHOR]

Both blue light (BL) and auxin are essential for phototropism in Arabidopsis thaliana. However, the mechanisms by which light is molecularly linked to auxin during phototropism remain elusive. Here, we report that PHYTOCHROME INTERACTING FACTOR4 (PIF4) and PIF5 act downstream of the BL sensor PHOTOTROPIN1 (PHOT1) to negatively modulate phototropism in Arabidopsis. We also reveal that PIF4 and PIF5 negatively regulate auxin signaling. Furthermore, we demonstrate that PIF4 directly activates the expression of the AUXIN / INDOLE-3-ACETIC ACID (IAA) genes IAA19 and IAA29 by binding to the G-box (CACGTG) motifs in their promoters. Our genetic assays demonstrate that IAA19 and IAA29, which physically interact with AUXIN RESPONSE FACTOR7 (ARF7), are sufficient for PIF4 to negatively regulate auxin signaling and phototropism. This study identifies a key step of phototropic signaling in Arabidopsis by showing that PIF4 and PIF5 link light and auxin. [ABSTRACT FROM AUTHOR]

Plant clathrin-mediated membrane trafficking is involved in many developmental processes as well as in responses to environmental cues. Previous studies have shown that clathrin-mediated endocytosis of the plasma membrane (PM) auxin transporter PIN-FORMED1 is regulated by the extracellular auxin receptor AUXIN BINDING PROTEIN1 (ABP1). However, the mechanisms by which ABP1 and other factors regulate clathrin-mediated trafficking are poorly understood. Here, we applied a genetic strategy and time-resolved imaging to dissect the role of clathrin light chains (CLCs) and ABP1 in auxin regulation of clathrin-mediated trafficking in Arabidopsis thaliana. Auxin was found to differentially regulate the PM and trans -Golgi network/early endosome (TGN / EE) association of CLCs and heavy chains (CHCs) in an ABP1-dependent but TRANSPORT INHIBITOR RESPONSE1/AUXIN-BINDING F-BOX PROTEIN (TIR1/AFB)-independent manner. Loss of CLC2 and CLC3 affected CHC membrane association, decreased both internalization and intracellular trafficking of PM proteins, and impaired auxin-regulated endocytosis. Consistent with these results, basipetal auxin transport, auxin sensitivity and distribution, and root gravitropism were also found to be dramatically altered in clc2 clc3 double mutants, resulting in pleiotropic defects in plant development. These results suggest that CLCs are key regulators in clathrin-mediated trafficking downstream of ABP1-mediated signaling and thus play a critical role in membrane trafficking from the TGN / EE and PM during plant development. [ABSTRACT FROM AUTHOR]

Transcriptional regulation plays a central role in plant hormone signaling. At the core of transcriptional regulation is the Mediator, an evolutionarily conserved, multisubunit complex that serves as a bridge between gene-specific transcription factors and the RNA polymerase machinery to regulate transcription. Here, we report the action mechanisms of the MEDIATOR25 (MED25) subunit of the Arabidopsis thaliana Mediator in regulating jasmonate- and abscisic acid (ABA)–triggered gene transcription. We show that during jasmonate signaling, MED25 physically associates with the basic helix-loop-helix transcription factor MYC2 in promoter regions of MYC2 target genes and exerts a positive effect on MYC2-regulated gene transcription. We also show that MED25 physically associates with the basic Leu zipper transcription factor ABA -INSENSITIVE5 (ABI5) in promoter regions of ABI5 target genes and shows a negative effect on ABI5-regulated gene transcription. Our results reveal that underlying the distinct effects of MED25 on jasmonate and ABA signaling, the interaction mechanisms of MED25 with MYC2 and ABI5 are different. These results highlight that the MED25 subunit of the Arabidopsis Mediator regulates a wide range of signaling pathways through selectively interacting with specific transcription factors. [ABSTRACT FROM AUTHOR]

The root stem cell niche, which in the Arabidopsis thaliana root meristem is an area of four mitotically inactive quiescent cells (QCs) and the surrounding mitotically active stem cells, is critical for root development and growth. We report here that during jasmonate-induced inhibition of primary root growth, jasmonate reduces root meristem activity and leads to irregular QC division and columella stem cell differentiation. Consistently, jasmonate reduces the expression levels of the AP2-domain transcription factors PLETHORA1 (PLT1) and PLT2, which form a developmentally instructive protein gradient and mediate auxin-induced regulation of stem cell niche maintenance. Not surprisingly, the effects of jasmonate on root stem cell niche maintenance and PLT expression require the functioning of MYC2/JASMONATE INSENSITIVE1, a basic helix-loop-helix transcription factor that involves versatile aspects of jasmonate-regulated gene expression. Gel shift and chromatin immunoprecipitation experiments reveal that MYC2 directly binds the promoters of PLT1 and PLT2 and represses their expression. We propose that MYC2-mediated repression of PLT expression integrates jasmonate action into the auxin pathway in regulating root meristem activity and stem cell niche maintenance. This study illustrates a molecular framework for jasmonate-induced inhibition of root growth through interaction with the growth regulator auxin. [ABSTRACT FROM AUTHOR]

The root stem cell niche defines the area that specifies and maintains the stem cells and is essential for the maintenance of root growth. Here, we characterize and examine the functional role of a quiescent center (QC)–expressed RAC/ROP GTPase activator, RopGEF7 , in Arabidopsis thaliana. We show that RopGEF7 interacts with At RAC1 and overexpression of a C-terminally truncated constitutively active RopGEF7 (RopGEF7 ΔC) activates RAC/ROP GTPases. Knockdown of RopGEF7 by RNA interference causes defects in embryo patterning and maintenance of the QC and leads to postembryonic loss of root stem cell population. Gene expression studies indicate that RopGEF7 is required for root meristem maintenance as it regulates the expression of PLETHORA1 (PLT1) and PLT2, which are key transcription factors that mediate the patterning of the root stem cell niche. Genetic analyses show that RopGEF7 interacts with PLT genes to regulate QC maintenance. Moreover, RopGEF7 is induced transcriptionally by auxin while its function is required for the expression of the auxin efflux protein PIN1 and maintenance of normal auxin maxima in embryos and seedling roots. These results suggest that RopGEF7 may integrate auxin-derived positional information in a feed-forward mechanism, regulating PLT transcription factors and thereby controlling the maintenance of root stem cell niches. [ABSTRACT FROM AUTHOR]

Recent identification of the Arabidopsis thaliana tyrosylprotein sulfotransferase (TPST) and a group of Tyr-sulfated peptides known as root meristem growth factors (RGFs) highlights the importance of protein Tyr sulfation in plant growth and development. Here, we report the action mechanism of TPST in maintenance of the root stem cell niche, which in the Arabidopsis root meristem is an area of four mitotically inactive quiescent cells plus the surrounding mitotically active stem cells. Mutation of TPST leads to defective maintenance of the root stem cell niche, decreased meristematic activity, and stunted root growth. We show that TPST expression is positively regulated by auxin and that mutation of this gene affects auxin distribution by reducing local expression levels of several PIN genes and auxin biosynthetic genes in the stem cell niche region. We also show that mutation of TPST impairs basal- and auxin-induced expression of the PLETHORA (PLT) stem cell transcription factor genes and that overexpression of PLT2 rescues the root meristem defects of the loss-of-function mutant of TPST. Together, these results support that TPST acts to maintain root stem cell niche by regulating basal- and auxin-induced expression of PLT1 and PLT2. TPST-dependent sulfation of RGFs provides a link between auxin and PLTs in regulating root stem cell niche maintenance. [ABSTRACT FROM AUTHOR]

The phytohormone abscisic acid (ABA) and the Polycomb group proteins have key roles in regulating plant growth and development; however, their interplay and underlying mechanisms are not fully understood. Here, we identified an Arabidopsis ( Arabidopsis thaliana ) nodulin homeobox (AtNDX) protein as a negative regulator in the ABA signaling pathway. AtNDX mutants are hypersensitive to ABA, as measured by inhibition of seed germination and root growth, and the expression of AtNDX is downregulated by ABA. AtNDX interacts with the Polycomb Repressive Complex1 (PRC1) core components AtRING1A and AtRING1B in vitro and in vivo, and together, they negatively regulate the expression levels of some ABA-responsive genes. We identified ABA - INSENSITIVE ( ABI4 ) as a direct target of AtNDX. AtNDX directly binds the downstream region of ABI4 and deleting this region increases the ABA sensitivity of primary root growth. Furthermore, ABI4 mutations rescue the ABA-hypersensitive phenotypes of ndx mutants and ABI4 -overexpressing plants are hypersensitive to ABA in primary root growth. Thus, our work reveals the critical functions of AtNDX and PRC1 in some ABA-mediated processes and their regulation of ABI4 ., (© 2020 American Society of Plant Biologists. All rights reserved.)

JASMONATE ZIM-DOMAIN (JAZ) transcriptional repressors are key regulators of jasmonate (JA) signaling in plants. At the resting stage, the C-terminal Jas motifs of JAZ proteins bind the transcription factor MYC2 to repress JA signaling. Upon hormone elicitation, the Jas motif binds the hormone receptor CORONATINE INSENSITIVE1, which mediates proteasomal degradation of JAZs and thereby allowing the Mediator subunit MED25 to activate MYC2. Subsequently, plants desensitize JA signaling by feedback generation of dominant JAZ splice variants that repress MYC2. Here we report the mechanistic function of Arabidopsis ( Arabidopsis thaliana ) MED25 in regulating the alternative splicing of JAZ genes through recruiting the splicing factors PRE-mRNA-PROCESSING PROTEIN 39a (PRP39a) and PRP40a. We demonstrate that JA-induced generation of JAZ splice variants depends on MED25 and that MED25 recruits PRP39a and PRP40a to promote the full splicing of JAZ genes. Therefore, MED25 forms a module with PRP39a and PRP40a to prevent excessive desensitization of JA signaling mediated by JAZ splice variants., (© 2020 American Society of Plant Biologists. All rights reserved.)

Groucho/Thymidine uptake 1 (Gro/Tup1) family proteins are evolutionarily conserved transcriptional coregulators in eukaryotic cells. Despite their prominent function in transcriptional repression, little is known about their role in transcriptional activation and the underlying mechanism. Here, we report that the plant Gro/Tup1 family protein LEUNIG_HOMOLOG (LUH) activates MYELOCYTOMATOSIS2 (MYC2)-directed transcription of JAZ2 and LOX2 via the Mediator complex coactivator and the histone acetyltransferase HAC1. We show that the Mediator subunit MED25 physically recruits LUH to MYC2 target promoters that then links MYC2 with HAC1-dependent acetylation of Lys-9 of histone H3 (H3K9ac) to activate JAZ2 and LOX2 Moreover, LUH promotes hormone-dependent enhancement of protein interactions between MYC2 and its coactivators MED25 and HAC1. Our results demonstrate that LUH interacts with MED25 and HAC1 through its distinct domains, thus imposing a selective advantage by acting as a scaffold for MYC2 activation. Therefore, the function of LUH in regulating jasmonate signaling is distinct from the function of TOPLESS, another member of the Gro/Tup1 family that represses MYC2-dependent gene expression in the resting stage., (© 2019 American Society of Plant Biologists. All rights reserved.)

In tomato ( Solanum lycopersicum ), as in other plants, the immunity hormone jasmonate (JA) triggers genome-wide transcriptional changes in response to pathogen and insect attack. These changes are largely regulated by the basic helix-loop-helix (bHLH) transcription factor MYC2. The function of MYC2 depends on its physical interaction with the MED25 subunit of the Mediator transcriptional coactivator complex. Although much has been learned about the MYC2-dependent transcriptional activation of JA-responsive genes, relatively less studied is the termination of JA-mediated transcriptional responses and the underlying mechanisms. Here, we report an unexpected function of MYC2 in regulating the termination of JA signaling through activating a small group of JA-inducible bHLH proteins, termed MYC2-TARGETED BHLH1 (MTB1), MTB2, and MTB3. MTB proteins negatively regulate JA-mediated transcriptional responses via their antagonistic effects on the functionality of the MYC2-MED25 transcriptional activation complex. MTB proteins impair the formation of the MYC2-MED25 complex and compete with MYC2 to bind to its target gene promoters. Therefore, MYC2 and MTB proteins form an autoregulatory negative feedback circuit to terminate JA signaling in a highly organized manner. We provide examples demonstrating that gene editing tools such as CRISPR/Cas9 open up new avenues to exploit MTB genes for crop protection., (© 2019 American Society of Plant Biologists. All rights reserved.)

Although the control of organ size is a fundamental question in developmental biology, little is known about the genetic and molecular mechanisms that determine the final size of seeds in plants. We previously demonstrated that the ubiquitin receptor DA1 acts synergistically with the E3 ubiquitin ligases DA2 and ENHANCER1 OF DA1 (EOD1)/BIG BROTHER to restrict seed growth in Arabidopsis thaliana. Here, we describe UBIQUITIN-SPECIFIC PROTEASE15 (UBP15), encoded by SUPPRESSOR2 OF DA1 (SOD2), which acts maternally to regulate seed size by promoting cell proliferation in the integuments of ovules and developing seeds. The sod2/ubp15 mutants form small seeds, while overexpression of UBP15 increases seed size of wild-type plants. Genetic analyses indicate that UBP15 functions antagonistically in a common pathway with DA1 to influence seed size, but does so independently of DA2 and EOD1. Further results reveal that DA1 physically associates with UBP15 in vitro and in vivo and modulates the stability of UBP15. Therefore, our findings establish a genetic and molecular framework for the regulation of seed size by four ubiquitin-related proteins DA1, DA2, EOD1, and UBP15 and suggest that they are promising targets for increasing seed size in crops.

Transcriptional regulation plays a central role in plant hormone signaling. At the core of transcriptional regulation is the Mediator, an evolutionarily conserved, multisubunit complex that serves as a bridge between gene-specific transcription factors and the RNA polymerase machinery to regulate transcription. Here, we report the action mechanisms of the MEDIATOR25 (MED25) subunit of the Arabidopsis thaliana Mediator in regulating jasmonate- and abscisic acid (ABA)-triggered gene transcription. We show that during jasmonate signaling, MED25 physically associates with the basic helix-loop-helix transcription factor MYC2 in promoter regions of MYC2 target genes and exerts a positive effect on MYC2-regulated gene transcription. We also show that MED25 physically associates with the basic Leu zipper transcription factor ABA-INSENSITIVE5 (ABI5) in promoter regions of ABI5 target genes and shows a negative effect on ABI5-regulated gene transcription. Our results reveal that underlying the distinct effects of MED25 on jasmonate and ABA signaling, the interaction mechanisms of MED25 with MYC2 and ABI5 are different. These results highlight that the MED25 subunit of the Arabidopsis Mediator regulates a wide range of signaling pathways through selectively interacting with specific transcription factors.

The root stem cell niche, which in the Arabidopsis thaliana root meristem is an area of four mitotically inactive quiescent cells (QCs) and the surrounding mitotically active stem cells, is critical for root development and growth. We report here that during jasmonate-induced inhibition of primary root growth, jasmonate reduces root meristem activity and leads to irregular QC division and columella stem cell differentiation. Consistently, jasmonate reduces the expression levels of the AP2-domain transcription factors PLETHORA1 (PLT1) and PLT2, which form a developmentally instructive protein gradient and mediate auxin-induced regulation of stem cell niche maintenance. Not surprisingly, the effects of jasmonate on root stem cell niche maintenance and PLT expression require the functioning of MYC2/JASMONATE INSENSITIVE1, a basic helix-loop-helix transcription factor that involves versatile aspects of jasmonate-regulated gene expression. Gel shift and chromatin immunoprecipitation experiments reveal that MYC2 directly binds the promoters of PLT1 and PLT2 and represses their expression. We propose that MYC2-mediated repression of PLT expression integrates jasmonate action into the auxin pathway in regulating root meristem activity and stem cell niche maintenance. This study illustrates a molecular framework for jasmonate-induced inhibition of root growth through interaction with the growth regulator auxin.

Plant roots show an impressive degree of plasticity in adapting their branching patterns to ever-changing growth conditions. An important mechanism underlying this adaptation ability is the interaction between hormonal and developmental signals. Here, we analyze the interaction of jasmonate with auxin to regulate lateral root (LR) formation through characterization of an Arabidopsis thaliana mutant, jasmonate-induced defective lateral root1 (jdl1/asa1-1). We demonstrate that, whereas exogenous jasmonate promotes LR formation in wild-type plants, it represses LR formation in jdl1/asa1-1. JDL1 encodes the auxin biosynthetic gene ANTHRANILATE SYNTHASE alpha1 (ASA1), which is required for jasmonate-induced auxin biosynthesis. Jasmonate elevates local auxin accumulation in the basal meristem of wild-type roots but reduces local auxin accumulation in the basal meristem of mutant roots, suggesting that, in addition to activating ASA1-dependent auxin biosynthesis, jasmonate also affects auxin transport. Indeed, jasmonate modifies the expression of auxin transport genes in an ASA1-dependent manner. We further provide evidence showing that the action mechanism of jasmonate to regulate LR formation through ASA1 differs from that of ethylene. Our results highlight the importance of ASA1 in jasmonate-induced auxin biosynthesis and reveal a role for jasmonate in the attenuation of auxin transport in the root and the fine-tuning of local auxin distribution in the root basal meristem.

Jasmonic acid (JA) is a lipid-derived signal that regulates plant defense responses to biotic stress. Here, we report the characterization of a JA-deficient mutant of tomato (Lycopersicon esculentum) that lacks local and systemic expression of defensive proteinase inhibitors (PIs) in response to wounding. Map-based cloning studies demonstrated that this phenotype results from loss of function of an acyl-CoA oxidase (ACX1A) that catalyzes the first step in the peroxisomal beta-oxidation stage of JA biosynthesis. Recombinant ACX1A exhibited a preference for C12 and C14 straight-chain acyl-CoAs and also was active in the metabolism of C18 cyclopentanoid-CoA precursors of JA. The overall growth, development, and reproduction of acx1 plants were similar to wild-type plants. However, the mutant was compromised in its defense against tobacco hornworm (Manduca sexta) attack. Grafting experiments showed that loss of ACX1A function disrupts the production of the transmissible signal for wound-induced PI expression but does not affect the recognition of this signal in undamaged responding leaves. We conclude that ACX1A is essential for the beta-oxidation stage of JA biosynthesis and that JA or its derivatives is required both for antiherbivore resistance and the production of the systemic wound signal. These findings support a role for peroxisomes in the production of lipid-based signaling molecules that promote systemic defense responses.

Genetic analysis of the wound response pathway in tomato indicates that systemin and its precursor protein, prosystemin, are upstream components of a defensive signaling cascade that involves the synthesis and subsequent action of the octadecatrienoic acid (18:3)-derived plant hormone jasmonic acid (JA). The suppressor of prosystemin-mediated responses2 (spr2) mutation, which was isolated previously as a suppressor of (pro)systemin-mediated signaling, impairs wound-induced JA biosynthesis and the production of a long-distance signal for the expression of defensive Proteinase inhibitor genes. Using a map-based cloning approach, we demonstrate here that Spr2 encodes a chloroplast fatty acid desaturase involved in JA biosynthesis. Loss of Spr2 function reduced the 18:3 content of leaves to <10% of wild-type levels, abolished the accumulation of hexadecatrienoic acid, and caused a corresponding increase in the level of dienoic fatty acids. The effect of spr2 on the fatty acyl content of various classes of glycerolipids indicated that the Spr2 gene product catalyzes most, if not all, omega3 fatty acid desaturation within the "prokaryotic pathway" for lipid synthesis in tomato leaves. Despite the reduced levels of trienoic fatty acids, spr2 plants exhibited normal growth, development, and reproduction. However, the mutant was compromised in defense against attack by tobacco hornworm larvae. These results indicate that jasmonate synthesis from chloroplast pools of 18:3 is required for wound- and systemin-induced defense responses and support a role for systemin in the production of a transmissible signal that is derived from the octadecanoid pathway.