Jigang Li

Plant hormone jasmonate prioritizes defense over growth by interfering with gibberellin signaling cascade

Plants must effectively defend against biotic and abiotic stresses to survive in nature. However, this defense is costly and is often accompanied by significant growth inhibition. How plants coordinate the fluctuating growth-defense dynamics is not well understood and remains a fundamental question. Jasmonate (JA) and gibberellic acid (GA) are important plant hormones that mediate defense and growth, respectively. Binding of bioactive JA or GA ligands to cognate receptors leads to proteasome-dependent degradation of specific transcriptional repressors (the JAZ or DELLA family of proteins), which, at the resting state, represses cognate transcription factors involved in defense (e.g., MYCs) or growth [e.g. phytochrome interacting factors (PIFs)]. In this study, we found that the coi1 JA receptor mutants of rice (a domesticated monocot crop) and Arabidopsis (a model dicot plant) both exhibit hallmark phenotypes of GA-hypersensitive mutants. JA delays GA-mediated DELLA protein degradation, and the della mutant is less sensitive to JA for growth inhibition. Overexpression of a selected group of JAZ repressors in Arabidopsis plants partially phenocopies GA-associated phenotypes of the coi1 mutant, and JAZ9 inhibits RGA (a DELLA protein) interaction with transcription factor PIF3. Importantly, the pif quadruple (pifq) mutant no longer responds to JA-induced growth inhibition, and overexpression of PIF3 could partially overcome JA-induced growth inhibition. Thus, a molecular cascade involving the COI1–JAZ–DELLA–PIF signaling module, by which angiosperm plants prioritize JA-mediated defense over growth, has been elucidated.

Phytochrome Signaling Mechanisms

Phytochromes are red (R)/far-red (FR) light photoreceptors that play fundamental roles in photoperception of the light environment and the subsequent adaptation of plant growth and development. There are five distinct phytochromes in Arabidopsis thaliana, designated phytochrome A (phyA) to phyE. phyA is light-labile and is the primary photoreceptor responsible for mediating photomorphogenic responses in FR light, whereas phyB-phyE are light stable, and phyB is the predominant phytochrome regulating de-etiolation responses in R light. Phytochromes are synthesized in the cytosol in their inactive Pr form. Upon light irradiation, phytochromes are converted to the biologically active Pfr form, and translocate into the nucleus. phyB can enter the nucleus by itself in response to R light, whereas phyA nuclear import depends on two small plant-specific proteins FAR-RED ELONGATED HYPOCOTYL 1 (FHY1) and FHY1-LIKE (FHL). Phytochromes may function as light-regulated serine/threonine kinases, and can phosphorylate several substrates, including themselves in vitro. Phytochromes are phosphoproteins, and can be dephosphorylated by a few protein phosphatases. Photoactivated phytochromes rapidly change the expression of light-responsive genes by repressing the activity of CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), an E3 ubiquitin ligase targeting several photomorphogenesis-promoting transcription factors for degradation, and by inducing rapid phosphorylation and degradation of Phytochrome-Interacting Factors (PIFs), a group of bHLH transcription factors repressing photomorphogenesis. Phytochromes are targeted by COP1 for degradation via the ubiquitin/26S proteasome pathway.

Genome-Wide Analysis of DNA Methylation and Gene Expression Changes in Two Arabidopsis Ecotypes and Their Reciprocal Hybrids

This work analyzed DNA methylation and gene expression changes in two Arabidopsis ecotypes and their reciprocal hybrids and found that increased DNA methylation in both hybrid genomes, possibly directed by the RdDM pathway, may play a role in heterosis. Heterosis is a fundamental biological phenomenon characterized by the superior performance of a hybrid over its parents in many traits, but the underlying molecular basis remains elusive. To investigate whether DNA methylation plays a role in heterosis, we compared at single-base-pair resolution the DNA methylomes of Arabidopsis thaliana Landsberg erecta and C24 parental lines and their reciprocal F1 hybrids that exhibited heterosis. Both hybrids displayed increased DNA methylation across their entire genomes, especially in transposable elements. Interestingly, increased methylation of the hybrid genomes predominantly occurred in regions that were differentially methylated in the two parents and covered by small RNAs, implying that the RNA-directed DNA methylation (RdDM) pathway may direct DNA methylation in hybrids. In addition, we found that 77 genes sensitive to methylome remodeling were transcriptionally repressed in both reciprocal hybrids, including genes involved in flavonoid biosynthesis and two circadian oscillator genes CIRCADIAN CLOCK ASSOCIATED1 and LATE ELONGATED HYPOCOTYL. Moreover, growth vigor of F1 hybrids was compromised by treatment with an agent that demethylates DNA and by abolishing production of functional small RNAs due to mutations in Arabidopsis RNA methyltransferase HUA ENHANCER1. Together, our data suggest that genome-wide remodeling of DNA methylation directed by the RdDM pathway may play a role in heterosis.

DWA1 and DWA2, two Arabidopsis DWD protein components of CUL4-based E3 ligases, act together as negative regulators in ABA signal transduction.

This article reports that DWA1 and DWA2 may be the substrate receptors for a CULLIN E3 ligase and that they interact with themselves and each other. Heterodimeric cooperation between DWAs is a previously unknown regulatory mechanism for the action of CULLIN E3 ligases. To elucidate potential roles of CUL4-DDB1-DWD (for Cullin 4-Damaged DNA Binding1-DDB1 binding WD40) E3 ligases in abscisic acid (ABA) signaling, we examined ABA sensitivities of T-DNA mutants of a number of Arabidopsis thaliana DWD genes, which encode substrate receptors for CUL4 E3 ligases. Mutants in two DWD genes, DWA1 and DWA2 (DWD hypersensitive to ABA1 and 2), had ABA-hypersensitive phenotypes. Both proteins interacted with DDB1 in yeast two-hybrid assays and associated with DDB1 and CUL4 in vivo, implying they could form CUL4-based complexes. Several ABA-responsive genes were hyperinduced in both mutants, and the ABA-responsive transcription factors ABA INSENSITIVE 5 (ABI5) and MYC2 accumulated to high levels in the mutants after ABA treatment. Moreover, ABI5 interacted with DWA1 and DWA2 in vivo. Cell-free degradation assays showed ABI5 was degraded more slowly in dwa1 and dwa2 than in wild-type cell extracts. Therefore, DWA1 and/or DWA2 may be the substrate receptors for a CUL4 E3 ligase that targets ABI5 for degradation. Our data indicate that DWA1 and DWA2 can directly interact with each other, and their double mutants exhibited enhanced ABA and NaCl hypersensitivities, implying they can act together. This report thus describes a previously unknown heterodimeric cooperation between two independent substrate receptors for CUL4-based E3 ligases.

Arabidopsis CULLIN4-Damaged DNA Binding Protein 1 Interacts with CONSTITUTIVELY PHOTOMORPHOGENIC1-SUPPRESSOR OF PHYA Complexes to Regulate Photomorphogenesis and Flowering Time

CUL4-DDB1 associates with COP1-SPA complexes via its linker protein DDB1 to regulate photomorphogenesis and possibly also flowering time under short-day conditions. The CUL4-DDB1-COP1-SPA supercomplex may represent a novel group of E3 ligases that functions independently of the CDD complex. CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1) possesses E3 ligase activity and promotes degradation of key factors involved in the light regulation of plant development. The finding that CULLIN4 (CUL4)-Damaged DNA Binding Protein1 (DDB1) interacts with DDB1 binding WD40 (DWD) proteins to act as E3 ligases implied that CUL4-DDB1 may associate with COP1-SUPPRESSOR OF PHYA (SPA) protein complexes, since COP1 and SPAs are DWD proteins. Here, we demonstrate that CUL4-DDB1 physically associates with COP1-SPA complexes in vitro and in vivo, likely via direct interaction of DDB1 with COP1 and SPAs. The interactions between DDB1 and COP1, SPA1, and SPA3 were disrupted by mutations in the WDXR motifs of MBP-COP1, His-SPA1, and His-SPA3. CUL4 cosuppression mutants enhanced weak cop1 photomorphogenesis and flowered early under short days. Early flowering of short day–grown cul4 mutants correlated with increased FLOWERING LOCUS T transcript levels, whereas CONSTANS transcript levels were not altered. De-etiolated1 and COP1 can bind DDB1 and may work with CUL4-DDB1 in distinct complexes, but they mediate photomorphogenesis in concert. Thus, a series of CUL4-DDB1-COP1-SPA E3 ligase complexes may mediate the repression of photomorphogenesis and, possibly, of flowering time.

Coordinated transcriptional regulation underlying the circadian clock in Arabidopsis

The circadian clock controls many metabolic, developmental and physiological processes in a time-of-day-specific manner in both plants and animals. The photoreceptors involved in the perception of light and entrainment of the circadian clock have been well characterized in plants. However, how light signals are transduced from the photoreceptors to the central circadian oscillator, and how the rhythmic expression pattern of a clock gene is generated and maintained by diurnal light signals remain unclear. Here, we show that in Arabidopsis thaliana, FHY3, FAR1 and HY5, three positive regulators of the phytochrome A signalling pathway, directly bind to the promoter of ELF4, a proposed component of the central oscillator, and activate its expression during the day, whereas the circadian-controlled CCA1 and LHY proteins directly suppress ELF4 expression periodically at dawn through physical interactions with these transcription-promoting factors. Our findings provide evidence that a set of light- and circadian-regulated transcription factors act directly and coordinately at the ELF4 promoter to regulate its cyclic expression, and establish a potential molecular link connecting the environmental light–dark cycle to the central oscillator.

A Molecular Framework of Light-Controlled Phytohormone Action in Arabidopsis

Environmental changes strongly affect plant growth and development. Phytohormones, endogenous plant-made small molecules such as ethylene, regulate a wide range of processes throughout the lifetime of plants. The ability of plants to integrate external signals with endogenous regulatory pathways is vital for their survival. Ethylene has been found to suppress hypocotyl elongation in darkness while promoting it in light. How ethylene regulates hypocotyl elongation in such opposite ways is largely unknown. In particular, how light modulates and even reverses the function of ethylene has yet to be characterized. Here we show that the basic-helix-loop-helix transcription factor phytochrome-interacting factor 3 (PIF3) is directly activated by ETHYLENE-INSENSITIVE 3 (EIN3) and is indispensible for ethylene-induced hypocotyl elongation in light. Ethylene via EIN3 concomitantly activates two contrasting pathways: the PIF3-dependent growth-promoting pathway and an ethylene response factor 1 (ERF1)-mediated growth-inhibiting pathway. In the light, growth-promoting PIFs are limiting due to light-dependent destabilization, and thus ethylene stimulates growth under these conditions. In contrast, ERF1 is destabilized, and thus limiting, under dark conditions, explaining why ethylene inhibits growth in the dark. Our findings provide a mechanistic insight into how light modulates internal hormone-regulated plant growth.

Arabidopsis COP1/SPA1 complex and FHY1/FHY3 associate with distinct phosphorylated forms of phytochrome A in balancing light signaling.

Fine tuning of light signaling is crucial to plant development. Following light-triggered nuclear translocation, the photoreceptor phytochrome A (phyA) regulates gene expression under continuous far-red light and is rapidly destabilized upon red light irradiation by E3 ubiquitin ligases, including COP1. Here we provide evidence that the light signaling repressors SPA proteins contribute to COP1-mediated phyA degradation and that a COP1/SPA1 protein complex is tightly associated with phyA ubiquitination activity. Furthermore, a phosphorylated phyA form accumulates in the nucleus and preferentially associates with the COP1/SPA1 complex. In contrast, underphosphorylated phyA predominantly associates with the phyA-signaling intermediates FHY3 and FHY1. However, COP1 associates with underphosphorylated phyA in the absence of FHY3 or FHY1, suggesting that phyA associations with FHY3 and FHY1 protect underphosphorylated phyA from being recognized by the COP1/SPA complex. We propose that light-induced phyA phosphorylation acts as a switch controlling differential interactions of the photoreceptor with signal propagation or attenuation machineries.

Genome-Wide Binding Site Analysis of FAR-RED ELONGATED HYPOCOTYL3 Reveals Its Novel Function in Arabidopsis Development

This work analyzed genome-wide binding sites of a transposase-derived transcription factor, FHY3, using a chromatin immunoprecipitation–based sequencing approach and identified a role for FHY3 in controlling chloroplast development by directly activating the expression of ARC5, a key gene involved in chloroplast division. FAR-RED ELONGATED HYPOCOTYL3 (FHY3) and its homolog FAR-RED IMPAIRED RESPONSE1 (FAR1), two transposase-derived transcription factors, are key components in phytochrome A signaling and the circadian clock. Here, we use chromatin immunoprecipitation–based sequencing (ChIP-seq) to identify 1559 and 1009 FHY3 direct target genes in darkness (D) and far-red (FR) light conditions, respectively, in the Arabidopsis thaliana genome. FHY3 preferentially binds to promoters through the FHY3/FAR1 binding motif (CACGCGC). Interestingly, FHY3 also binds to two motifs in the 178-bp Arabidopsis centromeric repeats. Comparison between the ChIP-seq and microarray data indicates that FHY3 quickly regulates the expression of 197 and 86 genes in D and FR, respectively. FHY3 also coregulates a number of common target genes with PHYTOCHROME INTERACTING FACTOR 3-LIKE5 and ELONGATED HYPOCOTYL5. Moreover, we uncover a role for FHY3 in controlling chloroplast development by directly activating the expression of ACCUMULATION AND REPLICATION OF CHLOROPLASTS5, whose product is a structural component of the latter stages of chloroplast division in Arabidopsis. Taken together, our data suggest that FHY3 regulates multiple facets of plant development, thus providing insights into its functions beyond light and circadian pathways.

Global identification of miRNAs and targets in Populus euphratica under salt stress

Populus euphratica, a typical hydro-halophyte, is ideal for studying salt stress responses in woody plants. MicroRNAs (miRNAs) are endogenous non-coding small RNAs that fulfilled an important post-transcriptional regulatory function. MiRNA may regulate tolerance to salt stress but this has not been widely studied in P. euphratica. In this investigation, the small RNAome, degradome and transcriptome were studied in salt stress treated P. euphratica by deep sequencing. Two hundred and eleven conserved miRNAs between Populus trichocarpa and P. euphratica have been found. In addition, 162 new miRNAs, belonging to 93 families, were identified in P. euphratica. Degradome sequencing experimentally verified 112 targets that belonged to 51 identified miRNAs, few of which were known previously in P. euphratica. Transcriptome profiling showed that expression of 15 miRNA-target pairs displayed reverse changing pattern under salt stress. Together, these results indicate that, in P. euphratica under salt stress, a large number of new miRNAs could be discovered, and both known and new miRNA were functionally cleaving to their target mRNA. Expression of miRNA and target were correspondingly induced by salt stress but that it was a complex process in P. euphratica.