Integrated Analysis of miRNAs Associated With Sugarcane Responses to Low-Potassium Stress

Sugarcane is among the most important global crops and a key bioenergy source. Sugarcane production is restricted by limited levels of available soil potassium (K+). The ability of plants to respond to stressors can be regulated by a range of microRNAs (miRNAs). However, there have been few studies regarding the roles of miRNAs in the regulation of sugarcane responses to K+-deficiency. To understand how these non-coding RNAs may influence sugarcane responses to low-K+ stress, we conducted expression profiling of miRNAs in sugarcane roots under low-K+ conditions via high-throughput sequencing. This approach led to the identification of 324 and 42 known and novel miRNAs, respectively, of which 36 were found to be differentially expressed miRNAs (DEMs) under low-K+ conditions. These results also suggested that miR156-x/z and miR171-x are involved in these responses as potential regulators of lateral root formation and the ethylene signaling pathway, respectively. A total of 705 putative targets of these DEMs were further identified through bioinformatics predictions and degradome analyses, and GO and KEGG enrichment analyses revealed these target mRNAs to be enriched for catalytic activity, binding functions, metabolic processes, plant hormone signal transduction, and mitogen-activated protein kinase (MAPK) signaling. In summary, these data provide an overview of the roles of miRNAs in the regulation of sugarcane response to low-K+ conditions.

Arabidopsis CPR5 Plays a Role in Regulating Nucleocytoplasmic Transport of mRNAs in Ethylene Signaling Pathway

The ETR1 receptor plays a predominant role in ethylene signaling in Arabidopsis thaliana. Previous studies showed that both RTE1 and CPR5 can directly bind to the ETR1 receptor and regulate ethylene signaling. RTE1 was suggested to promote the ETR1 receptor signaling by influencing its conformation, but little is known about the regulatory mechanism of CPR5 in ethylene signaling. In this study, we presented data showing that both RTE1 and CPR5 bound to the N-terminal domains of ETR1, and regulated ethylene signaling via the ethylene receptor. On the other hand, the research provided evidence indicating that CPR5 could act as a nucleoporin to regulate the ethylene-related mRNAs export out of the nucleus, while RTE1 or its homolog (RTH) had no effect on the nucleocytoplasmic transport of mRNAs. Nuclear qRT-PCR analysis and poly(A)-mRNA in situ hybridization showed that defect of CPR5 restricted nucleocytoplasmic transport of mRNAs. These results advance our understanding of the regulatory mechanism of CPR5 in ethylene signaling.

Overexpression of Galactinol Synthase 1 From Solanum Commersonii (ScGolS1) Confers Freezing Tolerance In Transgenic Potato

Potato (Solanum tuberosum L.) is the fourth largest food crop in the world. Low temperature causes serious damage to potato plants every year, and freezing tolerance has become a hot spot in potato research. Galactinol synthase (GolS) is a key enzyme in the synthesis of raffinose family oligosaccharides (RFOs), and plays an important role in the response of plants to abiotic stress. In this study, the ScGolS1 gene from S. commersonii was cloned and introduced into the S. tuberosum cultivars ‘Atlantic’ and ‘Desiree’ via Agrobacterium-mediated transformation. Phenotyping assay showed that overexpression of the ScGolS1 could significantly improve freezing tolerance in transgenic potato plants. Further physiological and biochemical results showed that the relative conductivity, malondialdehyde (MDA) content, and 3,3'-Diaminobenzidine (DAB) staining of the transgenic lines decreased, and the plant survival rate increased compared with wild type (WT). Moreover, CBF1, CBF2, CBF3, CBF downstream cold responsive genes COR413, COR47 and ERF transcription factor genes ERF3, ERF4, ERF6 in the ethylene signaling pathway were all induced by freezing treatment, while higher levels were observed in ScGolS1 overexpression lines compared with WT. In addition, other genes such as MIPS, STS and RS genes from RFO metabolic pathway and some sugars content were altered in response to freezing treatment. This indicates that overexpression of the ScGolS1 gene induced both the regulation of the ethylene signaling pathway and the metabolism of raffinose series oligosaccharides, regulating the balance of sugar composition and improved anti-peroxidation capacity, and thereby improved freezing tolerance in potato. These results provide theoretical support and genetic resources for freezing tolerance breeding in potato.

A Novel Effector Protein SsERP1 Inhibits Plant Ethylene Signaling to Promote Sclerotinia sclerotiorum Infection

Sclerotinia sclerotiorum is one of the most devastating pathogens in Brassica napus and causes huge economic loss worldwide. Though around one hundred putative effectors have been predicted in Sclerotinia sclerotiorum genome, their functions are largely unknown. In this study, we cloned and characterized a novel effector, SsERP1 (ethylene pathway repressor protein 1), in Sclerotinia sclerotiorum. SsERP1 is a secretory protein highly expressed at the early stages of Sclerotinia sclerotiorum infection. Ectopic overexpression of SsERP1 in plant leaves promoted Sclerotinia sclerotiorum infection, and the knockout mutants of SsERP1 showed reduced pathogenicity but retained normal mycelial growth and sclerotium formation, suggesting that SsERP1 specifically contributes to the pathogenesis of Sclerotinia sclerotiorum. Transcriptome analysis indicated that SsERP1 promotes Sclerotinia sclerotiorum infection by inhibiting plant ethylene signaling pathway. Moreover, we showed that knocking down SsERP1 by in vitro synthesized double-strand RNAs was able to effectively inhibit Sclerotinia sclerotiorum infection, which verifies the function of SsERP1 in Sclerotinia sclerotiorum pathogenesis and further suggests a potential strategy for Sclerotinia disease control.

The Diverse Salt-Stress Response of Arabidopsis ctr1-1 and ein2-1Ethylene Signaling Mutants Is Linked to Altered Root Auxin Homeostasis

We explored the interplay between ethylene signals and the auxin pool in roots exposed to high salinity using Arabidopsisthaliana wild-type plants (Col-0), and the ethylene-signaling mutants ctr1-1 (constitutive) and ein2-1 (insensitive). The negative effect of salt stress was less pronounced in ctr1-1 individuals, which was concomitant with augmented auxin signaling both in the ctr1-1 controls and after 100 mM NaCl treatment. The R2D2 auxin sensorallowed mapping this active auxin increase to the root epidermal cells in the late Cell Division (CDZ) and Transition Zone (TZ). In contrast, the ethylene-insensitive ein2-1 plants appeared depleted in active auxins. The involvement of ethylene/auxin crosstalk in the salt stress response was evaluated by introducing auxin reporters for local biosynthesis (pTAR2::GUS) and polar transport (pLAX3::GUS, pAUX1::AUX1-YFP, pPIN1::PIN1-GFP, pPIN2::PIN2-GFP, pPIN3::GUS) in the mutants. The constantly operating ethylene-signaling pathway in ctr1-1 was linked to increased auxin biosynthesis. This was accompanied by a steady expression of the auxin transporters evaluated by qRT-PCR and crosses with the auxin transport reporters. The results imply that the ability of ctr1-1 mutant to tolerate high salinity could be related to the altered ethylene/auxin regulatory loop manifested by a stabilized local auxin biosynthesis and transport.

Ethylene-Cytokinin Interaction Determines Early Defense Response of Wheat against Stagonospora nodorum Berk.

Ethylene, salicylic acid (SA), and jasmonic acid are the key phytohormones involved in plant immunity, and other plant hormones have been demonstrated to interact with them. The classic phytohormone cytokinins are important participants of plant defense signaling. Crosstalk between ethylene and cytokinins has not been sufficiently studied as an aspect of plant immunity and is addressed in the present research. We compared expression of the genes responsible for hormonal metabolism and signaling in wheat cultivars differing in resistance to Stagonospora nodorum in response to their infection with fungal isolates, whose virulence depends on the presence of the necrotrophic effector SnTox3. Furthermore, we studied the action of the exogenous cytokinins, ethephon (2-chloroethylphosphonic acid, ethylene-releasing agent) and 1-methylcyclopropene (1-MCP, inhibitor of ethylene action) on infected plants. Wheat susceptibility was shown to develop due to suppression of reactive oxygen species production and decreased content of active cytokinins brought about by SnTox3-mediated activation of the ethylene signaling pathway. SnTox3 decreased cytokinin content most quickly by its activated glucosylation in an ethylene-dependent manner and, furthermore, by oxidative degradation and inhibition of biosynthesis in ethylene-dependent and ethylene-independent manners. Exogenous zeatin application enhanced wheat resistance against S. nodorum through inhibition of the ethylene signaling pathway and upregulation of SA-dependent genes. Thus, ethylene inhibited triggering of SA-dependent resistance mechanism, at least in part, by suppression of the cytokinin signaling pathway.

Combined Linkage Analysis and Genome-wide Association Study Reveal QTLs and Candidate Genes Conferring Genetic Control of Prolificacy Trait in Maize

For the different harvest targets, the requirement for the prolificacy trait of maize was also different, so prolificacy is of great significance for modern production. Although some QTLs and genes associated with prolificacy in teosinte have been reported, the genetic mechanism of prolificacy in maize has not been fully elucidated. In this study, two RIL populations and GWAS population were used to genetic research of prolificacy trait in maize, with multi-environment. Combine linkage analysis and Genome-wide association study has identified a total of 13 QTLs and 8 significant SNPs. There were two genes related to tissue differentiation in the stable QTL qP9-2, and two significant SNPs corresponding to three genes were in QTL qP5-1 and QTL qP7-1, respectively. Four candidate genes GRMZM2G317262, GRMZM2G317584, GRMZM5G882364 and GRMZM2G141679 were finally screened out by qRT-PCR analysis. Based on the function of candidate genes, ethylene signaling pathway plays an important role in the formation of prolificacy in maize. It has deepened our understanding of the formation mechanism of prolificacy and laid a foundation for breeding new varieties with various prolificacy in maize.

A nuclear-localized cysteine desulfhydrase plays a role in fruit ripening in tomato

Hydrogen sulfide (H2S) is a gaseous signaling molecule that plays multiple roles in plant development. However, whether endogenous H2S plays a role in fruit ripening in tomato is still unknown. In this study, we show that the H2S-producing enzyme l-cysteine desulfhydrase SlLCD1 localizes to the nucleus. By constructing mutated forms of SlLCD1, we show that the amino acid residue K24 of SlLCD1 is the key amino acid that determines nuclear localization. Silencing of SlLCD1 by TRV-SlLCD1 accelerated fruit ripening and reduced H2S production compared with the control. A SlLCD1 gene-edited mutant obtained through CRISPR/Cas9 modification displayed a slightly dwarfed phenotype and accelerated fruit ripening. This mutant also showed increased cysteine content and produced less H2S, suggesting a role of SlLCD1 in H2S generation. Chlorophyll degradation and carotenoid accumulation were enhanced in the SlLCD1 mutant. Other ripening-related genes that play roles in chlorophyll degradation, carotenoid biosynthesis, cell wall degradation, ethylene biosynthesis, and the ethylene signaling pathway were enhanced at the transcriptional level in the lcd1 mutant. Total RNA was sequenced from unripe tomato fruit treated with exogenous H2S, and transcriptome analysis showed that ripening-related gene expression was suppressed. Based on the results for a SlLCD1 gene-edited mutant and exogenous H2S application, we propose that the nuclear-localized cysteine desulfhydrase SlLCD1 is required for endogenous H2S generation and participates in the regulation of tomato fruit ripening.

Phytol, a constituent of chlorophyll, induces root-knot nematode resistance in Arabidopsis via the ethylene signaling pathway

Root-knot nematodes (RKNs; Meloidogyne spp.) parasitize the roots and/or stems of a wide range of plant species, resulting in severe damage to the parasitized plant. The phytohormone ethylene (ET) plays an important role in signal transduction pathways leading to resistance against RKNs. However, little is currently known about the induction mechanisms of ET-dependent RKN resistance. Inoculation of Arabidopsis (Arabidopsis thaliana) roots with RKNs decreased chlorophyll contents in aerial parts of the plant. We observed accumulation of phytol, a constituent of chlorophyll and a precursor of tocopherols, in RKN-parasitized roots. Application of sclareol, a diterpene that has been shown to induce ET-dependent RKN resistance, to the roots of Arabidopsis plants increased phytol contents in roots accompanied by a decrease in chlorophyll in aerial parts. Exogenously applied phytol inhibited RKN penetration of roots without exhibiting nematicidal activity. This phytol-induced inhibition of RKN penetration was attenuated in the ET-insensitive Arabidopsis mutant ein2-1. Exogenously applied phytol enhanced the production of α-tocopherol and expression of VTE5, a gene involved in tocopherol production, in Arabidopsis roots. α-Tocopherol exerted similar induction of RKN resistance as phytol and showed increased accumulation in roots inoculated with RKNs. Furthermore, the Arabidopsis vte5 mutant displayed no inhibition of RKN penetration in response to phytol. These results suggest that exogenously applied phytol induces EIN2-dependent RKN resistance, possibly via tocopherol production.