Control of Jasmonate Biosynthesis and Senescence by miR319 Targets

In plants, microRNAs silence genes by guiding the cleavage or translational repression of complementary messenger RNAs. Because plant microRNA targets require high sequence complementarity, those targets are almost always transcription factors, and that makes the downstream effects relatively easy to trace. Most of the time, one transcription factor regulates another, which creates a cascade. The story that Schommer and colleagues tell breaks that pattern in a way that reframes how we think about leaf development. The microRNA in question is miR319, and its targets are a specific subset of class II TCP transcription factors, a plant-specific family whose name comes from its founding members: Teosinte Branched1, CYCLOIDEA, and the PCF proteins. In Arabidopsis, miR319 targets five of these: TCP2, TCP3, TCP4, TCP10, and TCP24. The key experimental system is a mutant called jaw-D, the first plant microRNA mutant ever described, which overexpresses miR319a and therefore has strongly reduced levels of all five TCP transcripts. Jaw-D plants have striking leaves — crinkled, negatively curved, and asymmetric. These shapes arise because TCP proteins normally suppress peripheral leaf growth, and without them, the leaf margin overgrows and buckles. The same phenotype appears in Antirrhinum when the TCP homolog CIN is knocked out and in tomato when miR319 control of TCPs is impaired. The module is conserved, and its primary role in leaf development seemed clear: miR319 downregulates TCPs, TCPs suppress growth, and the balance between them shapes the leaf. What Schommer and colleagues wanted to know was what, exactly, the TCPs were transcribing. Which immediate downstream genes carry out their effects? To find out, they took a convergent approach. They ran microarray comparisons across three genetic backgrounds: jaw-D plants with low TCP activity, rTCP4:GFP plants with elevated TCP activity, and tcp2 tcp4 tcp10 triple loss-of-function mutants. They looked for genes that changed consistently across at least two of those three comparisons. With relaxed thresholds — more than twofold change and per-gene variance with a p-value below 0.05 — they identified one hundred seventeen genes. With stringent criteria, only one gene survived: LIPOXYGENASE2, or LOX2. That result is worth sitting with. Out of the entire Arabidopsis genome, one gene responded so consistently across every TCP perturbation that it cleared the most demanding statistical bar. And LOX2 is not another transcription factor. It's an enzyme. Specifically, it's a chloroplast-localized lipoxygenase that converts alpha-linolenic acid into the first committed intermediate in jasmonic acid biosynthesis. The TCPs aren't feeding into another regulatory cascade. They're directly controlling a hormone factory. The team confirmed this with biochemistry. They expressed the DNA-binding domain of TCP4 in bacteria, purified it, and ran an in vitro selection: presenting the protein with a random pool of DNA sequences to see which ones it retains. Of twenty-seven recovered clones, twenty-five contained the same core motif — GGACCA. That hexamer turned up in the promoters of eight of the nineteen known jasmonic acid biosynthesis genes, while chance alone would predict only about two. Electrophoretic mobility shift assays, where you run protein and DNA together on a gel and watch the band slow down if binding occurs, showed TCP4 binding strongly to at least two of the four matching sites in the LOX2 promoter. The in-plant evidence was just as clear. The team built two LOX2 reporter constructs — one with the wild-type promoter driving a GUS marker gene, and one with all four TCP binding sites mutated. They created twenty independent transgenic lines per construct. In untreated plants, the wild-type promoter drove strong GUS activity throughout the leaf. The mutated promoter had almost none. And in tcp2 tcp4 tcp10 triple mutants, even the wild-type reporter went quiet, confirming that TCP proteins are required for developmental LOX2 expression in vivo. Crucially, wounding or methyl jasmonate treatment activated both reporters equally within forty-five minutes, meaning the TCP sites control developmental expression, but wound-triggered induction runs through a separate pathway entirely. The pathway-level effects were just as clear as the single-gene ones. The average expression of the nineteen known jasmonic acid biosynthetic genes was about twofold lower in jaw-D plants and about fourfold higher in rTCP4:GFP plants compared to wild type. When the team wounded leaves and measured actual jasmonic acid levels by gas chromatography mass spectrometry, jaw-D plants accumulated roughly four times less jasmonic acid at the ninety-minute peak than wild type. TCPs don't just regulate LOX2; they set the gain on the entire jasmonate biosynthesis system. Now comes the second half of the story, and this is where the findings connect into something genuinely surprising. Jasmonic acid has long been proposed to regulate leaf senescence, the programmed aging and dismantling of a leaf that allows the plant to reclaim its nutrients. If TCPs drive jasmonate production, and jasmonate drives senescence, then plants with low TCP activity should age more slowly. That's exactly what Schommer and colleagues found. In jaw-D plants, senescence of the fifth rosette leaf was delayed by about a week compared to wild type. The team confirmed this in detached leaves floated on water in darkness, a standard induced-senescence assay. Jaw-D leaves stayed greener longer and maintained higher Fv/Fm, which is the maximum efficiency of photosystem II photochemistry, and drops as chloroplasts are dismantled. Wild-type leaves declined, while jaw-D leaves held on. The rescue experiment clinched the causal chain. When the team floated detached jaw-D leaves on solutions with increasing concentrations of methyl jasmonate, the senescence delay disappeared. The hormone was sufficient to put the aging program back on schedule, even in a background with crippled TCP activity. This indicates that the block in jaw-D is upstream of jasmonate signaling — the receptors and downstream machinery are intact. The deficit is in making the hormone in the first place. TCP leads to jasmonate, and jasmonate leads to senescence. Schommer and colleagues also checked whether another hormone pathway might be responsible. Salicylic acid can antagonize jasmonate signaling, so if salicylic acid signaling were elevated in jaw-D, it could independently explain the delayed senescence. It wasn't. Induction of the salicylic acid marker gene PR1 appeared normal in jaw-D plants. The delayed aging is about jasmonic acid biosynthesis, not a salicylic acid confound. Step back, and the developmental logic comes into focus. Early in leaf development, miR319 keeps TCP activity low, allowing the leaf to grow and expand. As the leaf matures, that balance shifts: TCP activity rises, LOX2 and related biosynthesis genes are transcribed, jasmonate accumulates, and the senescence program is engaged. Two opposite life-history phases — expansion and programmed death — are coordinated by the same regulatory axis. Schommer and colleagues note that many genes induced by elevated TCP activity are also progressively induced during leaf development, including the senescence regulator WRKY53, which fits the picture of TCPs functioning continuously across the leaf's lifespan rather than switching abruptly at one developmental moment. The precise dynamics of how the miR319-to-TCP ratio changes over an individual leaf's life, and whether this same timing mechanism operates across plant species, remain open questions. The paper documents TCP function in Antirrhinum and growth effects in tomato, but whether the full axis — miR319 decline, TCP rise, developmental activation of jasmonate biosynthesis, and then senescence — plays out with the same timing elsewhere has not been tested. Additionally, because jasmonate also mediates pathogen responses and mechanical stress, TCP control of jasmonic acid biosynthesis connects this developmental module to defense and environmental signaling as well. What Schommer and colleagues delivered is a precise molecular account of how a leaf decides when to stop growing and start dying. One microRNA, five transcription factors, a direct binding site in the promoter of a biosynthetic enzyme, and a hormone that sets the clock. That's a remarkably short chain for a decision as consequential as when a leaf calls it a life. This lecture was created by ennepō. Go to https://ennepo.ai to Discover, Create and Follow the latest research in your field. Read when you can. Listen when you want to.