DNA-Binding with One Finger (Dof) Transcription Factor Gene Family Study Reveals Differential Stress-Responsive Transcription Factors in Contrasting Drought Tolerance Potato Species.

DNA-binding with one finger (Dof) proteins comprise a large family that play central roles in stress tolerance by regulating the expression of stress-responsive genes via the DOFCORE element or by interacting with other regulatory proteins. Although the Dof TF has been identified in a variety of species, its systemic analysis in potato (Solanum tuberosum L.) is lacking and its potential role in abiotic stress responses remains unclear. A total of 36 potential Dof genes in potato were examined at the genomic and transcriptomic levels in this work. Five phylogenetic groups can be formed from these 36 Dof proteins. An analysis of cis-acting elements revealed the potential roles of Dofs in potato development, including under numerous abiotic stress conditions. The cycling Dof factors (CDFs) might be the initial step in the abiotic stress response signaling cascade. In potato, five CDFs (StCDF1/StDof19, StCDF2/StDof4, StCDF3/StDof11, StCDF4/StDof24, and StCDF5/StDof15) were identified, which are homologs of Arabidopsis CDFs. The results revealed that these genes were engaged in a variety of abiotic reactions. Moreover, an expression analysis of StDof genes in two potato cultivars ('Long10' (drought tolerant) and 'DXY' (drought susceptible)) of contrasting tolerances under drought stress was carried out. Further, a regulatory network mediated by lncRNA and its target Dofs was established. The present study provides fundamental knowledge for further investigation of the roles of Dofs in the adaptation of potato to drought stress, aiming to provide insights into a viable strategy for crop improvement and stress-resistance breeding.

Potato Stu-miR398b-3p Negatively Regulates Cu/Zn-SOD Response to Drought Tolerance.

One of the main impacts of drought stress on plants is an excessive buildup of reactive oxygen species (ROS). A large number of ·OH, highly toxic to cells, will be produced if too much ROS is not quickly cleared. At the heart of antioxidant enzymes is superoxide dismutase (SOD), which is the first antioxidant enzyme to function in the active oxygen scavenging system. To shield cells from oxidative injury, SOD dismutation superoxide anion free radicals generate hydrogen peroxide and molecule oxygen. Cu/Zn SOD is a kind of SOD antioxidant enzyme that is mostly found in higher plants' cytoplasm and chloroplasts. Other studies have demonstrated the significance of the miR398s family of miRNAs in the response of plants to environmental stress. The cleavage location of potato stu-miR398b-3p on Cu/Zn SOD mRNA was verified using RLM-5'RACE. Using the potato variety 'Desiree', the stu-miR398b-3p overexpression mutant was created, and transgenic lines were raised. SOD activity in transgenic lines was discovered to be decreased during drought stress, although other antioxidant enzyme activities were mostly unaltered. Transgenic plants will wilt more quickly than wild-type plants without irrigation. Additionally, this demonstrates that the response of Cu/Zn SOD to drought stress is adversely regulated by potato stu-miR398b-3p.

Stu-miR827-Targeted StWRKY48 Transcription Factor Negatively Regulates Drought Tolerance of Potato by Increasing Leaf Stomatal Density.

Stomata are specialized portals in plant leaves to modulate water loss from plants to the atmosphere by control of the transpiration, thereby determining the water-use efficiency and drought resistance of plants. Despite that the stomata developmental progression is well-understood at the molecular level, the experimental evidence that miRNA regulates stomata development is still lacking, and the underlying mechanism remains elusive. This study demonstrates the involvement of stu-miR827 in regulating the drought tolerance of potato due to its control over the leaf stomatal density. The expression analysis showed that stu-miR827 was obviously repressed by drought stresses and then rapidly increased after rewatering. Suppressing the expression of stu-miR827 transgenic potato lines showed an increase in stomatal density, correlating with a weaker drought resistance compared with wildtype potato lines. In addition, StWRKY48 was identified as the target gene of stu-miR827, and the expression of StWRKY48 was obviously induced by drought stresses and was greatly upregulated in stu-miR827 knockdown transgenic potato lines, suggesting its involvement in the drought stress response. Importantly, the expression of genes associated with stomata development, such as SDD (stomatal density and distribution) and TMM (too many mouths), was seriously suppressed in transgenic lines. Altogether, these observations demonstrated that suppression of stu-miR827 might lead to overexpression of StWRKY48, which may contribute to negatively regulating the drought adaptation of potato by increasing the stomatal density. The results may facilitate functional studies of miRNAs in the process of drought tolerance in plants.

Calcium-Dependent Protein Kinase 28 Maintains Potato Photosynthesis and Its Tolerance under Water Deficiency and Osmotic Stress.

Calcium-dependent protein kinases (CDPK) are implicated in signaling transduction in eukaryotic organisms. It is largely unknown whether StCDPK28 plays a role in the response to water deficiency and osmotic stress in potato plants (Solanum tuberosum L.). Potato cv. Zihuabai was cultivated under natural, moderate, and severe water deficiency conditions; to induce osmotic stress, potato plants were treated with 10% or 20% PEG. StCDPK28-overexpression and StCDPK28-knockdown plants were constructed. StCDPKs were evaluated by qRT-PCR. The subcellular location of the StCDPK28 protein was observed with confocal scanning laser microscopy. Phenotypic changes were indicated by photosynthetic activity, the contents of H2O2, MDA and proline, and the activities of CAT, SOD and POD. Results showed water deficiency and osmotic stress altered StCDPK expression patterns. StCDPK28 exhibited a membrane, cytosolic and nuclear localization. Water deficiency and osmotic stress induced StCDPK28 upregulation. Photosynthetic activity was enhanced by StCDPK28 overexpression, while decreased by StCDPK2 knockdown under water deficiency and osmotic stress. StCDPK28 overexpression decreased H2O2 and MDA, and increased proline, while StCDPK28 knockdown showed reverse results, compared with the wild type, in response to water deficiency and osmotic stress. StCDPK28 overexpression increased the activities of CAT, SOD and POD, while StCDPK28-knockdown plants indicated the reverse trend under water deficiency and osmotic stress conditions. Regulation of StCDPK28 expression could be a promising approach to improve the tolerance ability of potato plants in response to drought or high salt media.

Overexpression of StCDPK23 promotes wound healing of potato tubers by regulating StRbohs.

Calcium-dependent protein kinase (CDPK) is a Ca2+ sensor that can phosphorylate and regulate respiratory burst oxidase homolog (Rboh), inducing the production of O2-. However, little is known about how StCDPK23 affects ROS production in the deposition of suberin at potato tuber wounds by regulating StRbohs. In this study, we found that StCDPK23 was induced significantly by the wound in potato tubers, which contains a typical CDPK structure, and was highly homologous to AtCDPK13 in Arabidopsis. Subcellular localization of results showed that StCDPK23 was located in the nucleus and plasma membrane of N. benthamiana epidermis cells. StCDPK23-overexpressing plants and tubers were obtained via Agrobacterium transformation. The expression of StCDPK23 was significantly upregulated in the overexpressing tubers during healing and increased 2.3-fold at 5 d. The expression levels of StRbohs (A-E) were also upregulated in the overexpressing tubers. Among them, StrbohA showed significant expression in the early stage of healing, which was 16.3-fold higher than that of the wild-type tubers at 8 h of healing. Moreover, the overexpressing tubers produced more O2- and H2O2, which are 1.1-fold and 3.5-fold higher than that of the wild-type at 8 h, respectively. More SPP deposition was observed at the wounds of the overexpressing tubers. The thickness of SPP cell layers was 53.2% higher than that of the wild-type after 3 d of the wound. It is suggested that StCDPK23 may participate in the wound healing of potato tubers by regulating Strbohs, which mainly contributes to H2O2 production during healing.

Genome-Wide Identification, Characterization and Expression Analysis of the CIPK Gene Family in Potato (Solanum tuberosum L.) and the Role of StCIPK10 in Response to Drought and Osmotic Stress.

The potato (Solanum tuberosum L.), one of the most important food crops worldwide, is sensitive to environmental stresses. Sensor-responder complexes comprising calcineurin B-like (CBL) proteins and CBL-interacting protein kinases (CIPKs) not only modulate plant growth and development but also mediate numerous stress responses. Here, using a Hidden Markov Model and BLAST searches, 27 CIPK genes were identified in potato and divided into five groups by phylogenetic analysis and into two clades (intron-poor and intron-rich) by gene structure analysis. Quantitative reverse-transcription PCR (qRT-PCR) assays revealed that StCIPK genes play important roles in plant growth, development and abiotic stress tolerance. Up-regulated expression of StCIPK10 was significantly induced by drought, PEG6000 and ABA. StCIPK10 enhances both the ability of potato to scavenge reactive oxygen species and the content of corresponding osmoregulation substances, thereby strengthening tolerance to drought and osmotic stress. StCIPK10 is located at the intersection between the abscisic acid and abiotic stress signaling pathways, which control both root growth and stomatal closure in potato. In addition, StCIPK10 interacts with StCBL1, StCBL4, StCBL6, StCBL7, StCBL8, StCBL11 and StCBL12, and is specifically recruited to the plasma membrane by StCBL11.

Knockdown of MicroRNA160a/b by STTM leads to root architecture changes via auxin signaling in Solanum tuberosum.

The root phenotype is an important aspect of plant architecture and plays a critical role in plant facilitation of the extraction of water and nutrition from the soil. MicroRNAs (miRNAs) are classes of small RNAs with important roles in regulating endogenous gene expression at the post-transcriptional level that function in a range of plant development processes and in the response to abiotic stresses. However, little is known concerning the molecular mechanism of miRNAs in regulating the generation and development of plant root architecture. Herein, we demonstrated that potato miR160a/b acted as a critical regulator and affected plant root architecture by targeting the mRNA of StARF10 and StARF16 for cleavage. The miR160a/b precursor was cloned from potato. Quantitative PCR assays showed that the expression levels of miR160 and its targets were down- or up-regulated with the development of potato roots, respectively. Moreover, transgenic lines with suppressed stu-miR160 expression were established with the short tandem targets mimic (STTM), and the results showed that the ectopic expression of miR160a/b altered the levels of auxin and the expression of auxin signaling-related genes and caused drastic change in root architecture compared with that in control plants. Suppressing the expression of miR160 led to a severe reduction in root length, an increase in the number of lateral roots, and a decrease in fresh root weight in potato. Collectively, our data established a key role of miR160 in modulating plant root architecture in potato.

Jianpi Jieyu Decoction, An Empirical Herbal Formula, Exerts Psychotropic Effects in Association With Modulation of Gut Microbial Diversity and GABA Activity.

Background: Recent studies suggest that gut microbiota was associated with the bidirectional gut-brain axis which could modulate neuropsychological functions of the central nervous system. Gut microbiota could produce gamma aminobutyric acid (GABA) that could modulate the gut-brain axis response. Jianpi Jieyu (JPJY) decoction, a traditional Chinese formula, is mainly composed of Astragalus membranaxeus and Radix Pseudostellariae. Although the JPJY decoction has been used to treat the depression in China, the potential action of its antidepressant has not been well understood. Thus this study was aim to investigate the role of JPJY improve gut microbiota homeostasis in the chronic stress induced depressive mice. Methods: The antidepressant effect of JPJY on chronic unpredictable mild stress (CUMS) mice was evaluated by using sucrose preference test, tail suspension test and forced swim test. Fatigue-like behaviors were evaluated using degree of redness, grip strength test, and exhaustive swimming test. The new object recognition test was used to evaluate cognition performance. Fecal samples were collected and taxonomical analysis of intestinal microbial distribution was conducted with 16S rDNA. Serum level of GABA was measured using high performance liquid chromatography (HPLC). The expression of GluR1 and p-Tau protein in the hippocampus was determined using Western blotting. Results: The dose of 9.2 g/kg JPJY produced antidepressant-like effects. JPJY and its major components also modulated gut microbiota diversity in the CUMS mice. Serum level of GABA and the expressions of hippocampal GluR1 and p-Tau were reversed after the administration of JPJY in CUMS mice. Conclusion: JPJY regulates gut microbiota to produce antidepressant-like effect and improve cognition deficit in depressive mice while its molecular mechanism possibly be enhanced NR1 and Tau expression in hippocampus and increased GABA in serum.

Mitogen-activated protein kinase 11 (MAPK11) maintains growth and photosynthesis of potato plant under drought condition.

KEY MESSAGE: StMAPK11 overexpression promotes potato growth, physiological activities and photosynthesis under drought conditions. Mitogen-activated protein kinases (MAPKs) are import regulators of MAPK pathway in plants under drought condition. However, the critical role in potato (Solanum tuberosum L.) drought resistance is not fully understood. In this study, we aimed to explore the role of StMAPK11 under drought stress. The result of RT-qPCR for assay of StMAPKs expression demonstrated that 15 StMAPKs were differentially expressed in leaves, flowers, petioles, stamens, pistils, stems, stolons, roots, tubers and tuber peels of potato. StMAPKs was dynamically modulated by abiotic stresses and plant hormone treatments, and StMAPK11 was apparently up-regulated under drought conditions. Therefore, the vectors pCPB-StMAPK11 and pCPBI121-miRmapk11 for over-expression and down-regulation of StMAPK11 were constructed, respectively, and introduced into potato cultivar Atlantic. The result showed that StMAPK11 promoted potato growth under drought conditions, as well as the physiological activities evidenced by changes in SOD, CAT and POD activity and H2O2, proline and MDA content. StMAPK11 up-regulation intensified drought resistance of potato plant by elevating antioxidant activities and photosynthesis. Moreover, we consolidated the protective role of StMAPK11 in tobacco and Arabidopsis against drought stress. The result could provide new insights into the function of StMAPK11 in drought response and its possible mechanisms.

Protection against peripheral artery disease injury by Ruan Jian Qing Mai formula via metabolic programming.

Ruan Jian Qing Mai formula (RJQM), a multicomponent herbal formula, has been widely used to treat peripheral arterial disease (PAD) in China. However, its active compounds and mechanisms of action are still unknown. First, RNA sequencing analysis of 15 healthy and 16 PAD samples showed that 524 PAD differential genes were significantly enriched in Go Ontology (ribonucleotide metabolic process, oxidoreductase complex, and electron transfer activity), Kyoto Encyclopedia of Genes and Genomes (KEGG) and GSEA pathways (OXPHOS and TCA cycle), miRNA (MIR183), and kinase (PAK6). Fifty-three active ingredients in RJQM had similar structures to the seven drug molecules in CLUE. Then, network topology analysis of the 53 components-target-pathway-disease network yielded 10 active ingredients. Finally, computational toxicity estimations showed that the median lethal dose (LD50) of the 10 active ingredients was above 1000 mg/kg, and eight of them did not cause hepatotoxicity, mutagenicity, carcinogenicity, cytotoxicity, and immunotoxicity nor activate 12 toxic pathways. In conclusion, RJQM has a protection effect on PAD by regulating a complex molecular network. Part of the mechanism is associated with the regulation of OXPHOS by 10 active components, which may alleviate mitochondrial dysfunction and pathological metabolic programming.

StMAPK3 controls oxidase activity, photosynthesis and stomatal aperture under salinity and osmosis stress in potato.

Mitogen-activated protein kinase 3 (MAPK3) is involved in plant growth and development, as well as response to adverse stress. Here we aimed to explore the role of StMAPK3 in response to salt and osmosis stress. Polyethylene glycol (PEG) (5% and 10%) and mannitol (40 mM and 80 mM) were used to induce osmosis stress. To induce salinity stress, potato plant was cultured with NaCl (40 mM and 80 mM). StMAPK3 overexpression and RNA interference-mediated StMAPK3 knockdown were constructed to explore the role of StMAPK3 in potato growth, stomatal aperture size, activity of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), and contents of H2O2, proline and malonaldehyde (MDA). Meanwhile, we detected transpiration, net photosynthesis, stomatal conductance, and water use efficiency. Subcellular location of StMAPK3 protein was also detected. PEG, mannitol and NaCl treatments induced the accumulation of StMAPK3 mRNA in potato plants. StMAPK3 protein was located on the membrane and nucleus. Abnormal expression of StMAPK3 changed potato phenotypes, enzyme activity of SOD, CAT and POD, as well as H2O2, proline and MDA contents under osmosis and salinity stress. Photosynthesis and stomatal aperture were regulated by StMAPK3 in potato treated by PEG, mannitol and NaCl. Modulation of potato phenotypes and physiological activity indicates StMAPK3 as a regulator of osmosis and salinity tolerance.

Protection against COVID-19 injury by qingfei paidu decoction via anti-viral, anti-inflammatory activity and metabolic programming.

Qingfei Paidu decoction (QFPD), a multi-component herbal formula, has been widely used to treat COVID-19 in China. However, its active compounds and mechanisms of action are still unknown. Firstly, we divided QFPD into five functional units (FUs) according to the compatibility theory of traditional Chinese medicine. The corresponding common targets of the five FUs were all significantly enriched in Go Ontology (oxidoreductase activity, lipid metabolic process, homeostatic process, etc.), KEGG pathways (steroid biosynthesis, PPAR signaling pathway, adipocytokine signaling pathway, etc.), TTD diseases (chronic inflammatory diseases, asthma, chronic obstructive pulmonary Disease, etc.), miRNA (MIR183), kinase (CDK7) and TF (LXR). QFPD contained 257 specific targets in addition to HCoV, pneumonia and ACE2 co-expression proteins. Then, network topology analysis of the five components-target-pathway-disease networks yielded 67 active ingredients. In addition, ADMET estimations showed that 20 compounds passed the stringent lead-like criteria and in silico drug-likeness test with high gastrointestinal absorption and the median lethal dose (LD50 > 1600 mg/kg). Moreover, 4 specific ingredients (M3, S1, X2 and O2) and 5 common ingredients (MS1, MX16, SX1, WO1 and XO1) of QFPD presented good molecular docking score for 2019-nCov structure and non-structure proteins. Finally, drug perturbation of COVID-19 network robustness showed that all five FUs may protect COVID-19 independently, and target 8 specifically expressed drug-attacked nodes which were related to the bacterial and viral responses, immune system, signaling transduction, etc. In conclusion, our new FUNP analysis showed that QFPD had a protection effect on COVID-19 by regulating a complex molecular network with safety and efficacy. Part of the mechanism was associated with the regulation of anti-viral, anti-inflammatory activity and metabolic programming.

Improving salt tolerance in potato through overexpression of AtHKT1 gene.

BACKGROUND: Survival of plants in response to salinity stress is typically related to Na+ toxicity, but little is known about how heterologous high-affinity potassium transporter (HKT) may help alleviate salt-induced damages in potato (Solanum tuberosum L.). RESULTS: In this study, we used the Arabidopsis thaliana high-affinity potassium transporter gene (AtHKT1) to enhance the capacity of potato plants to tolerate salinity stress by decreasing Na+ content and improving K+/Na+ ratio in plant leaves, while maintaining osmotic balance. Seven AtHKT1 transformed potato lines (namely T1, T2, T3, T5, T11, T13 and T15) were compared with non-transgenic control plant at molecule and whole-plant levels. The lines T3 and T13 had the highest AtHKT1 expression with the tolerance index (an quantitative assessment) being 6.8 times that of the control. At 30 days under 100 and 150 mmol L- 1 NaCl stress treatments, the T3 and T13 lines had least reductions in net photosynthetic rate, stomatal conductance and transpiration rate among the seven lines, leading to the increased water use efficiency and decreased yield loss. CONCLUSIONS: We conclude that the constitutive overexpression of AtHKT1 reduces Na+ accumulation in potato leaves and promotes the K+/Na+ homeostasis that minimizes osmotic imbalance, maintains photosynthesis and stomatal conductance, and increases plant productivity.

Identification and expression analysis of StGRAS gene family in potato (Solanum tuberosum L.).

The GRAS gene family is a class of plant-specific transcription factors which play pivotal roles in the regulation of plant growth and development. At present, the GRAS gene family has been completely identified in Arabidopsis thaliana, however, there are no systematic research reports in potato. In the present study, we obtained an overview of the GRAS gene family including gene structure, gene expression, chromosome mapping and phylogenetic analysis, and 52 StGRASs were identified in the potato by bioinformatics analysis, which could be divided into eight subfamilies based on phylogeny. More than 90% of genes do not contain introns and the StGRAS family major function is protein binding according to gene ontology analysis (GO).The tissue specific expression analysis showed that StGRAS3, StGRAS35 and StGRAS50 gene had the higher expression in roots, stems and leaves compared with other StGRAS, StGRAS9 and StGRAS28 genes were responded to plant hormones IAA, ABA and GA3 treatment. The result could provide a basis for further studying the function of GRAS genes and GRAS-mediated signal transduction pathways in potato.

Genome-wide identification and expression analysis of StTCP transcription factors of potato (Solanum tuberosum L.).

The plant-specific TCP transcription factors, which play critical roles in diverse aspects of biological processes, have been identified and analyzed in various plant species. However, no systematical study of TCP family genes in potato (Solanum tuberosum L.) has been undertaken. In this study, a total of 31 non-redundant TCP transcription factors of potato were identified and divided into two subfamilies including three distinct subclades. The various orthologous TCP genes in Arabidopsis, rice, potato and tomato were identified using synteny and phylogenetic analysis. Protein motif analysis demonstrated that StTCPs in the same subclade shared similar conserved motif structures. Gene structure analysis showed that almost all StTCPs displayed highly conserved exon-intron organization. The analysis of StTCP gene promoter regions revealed that multiple cis-acting elements were involved in plant growth, development, hormone responses as well as stress responses. The result of StTCP gene expression profiles showed they had tissue-specific expression patterns which implied their differentiated functions. According to the results of quantitative RT-PCR (qRT-PCR), 7 StTCP genes were dramatically up-regulated during the release of tuber dormancy and some specific StTCP genes were strongly responding to different abiotic stresses and multiple hormones, which suggested they had important roles in potato growth and development processes. The results of our findings could provide comprehensive insights in StTCP family genes of potato for further functional investigations.

Functional analysis of StDWF4 gene in response to salt stress in potato.

The DWARF4 (DWF4) gene encodes a C-22 hydroxylase which is pivotal for brassinosteroids (BRs) biosynthesis. In this research, aimed to understand the molecular mechanism of DWF4 on regulation of potatoes tolerance to salt stress, DWF4 was cloned from potato, named as StDWF4. Its 1476 bp open reading frame encodes a protein of 491 amino acids. The StDWF4-overexpressing (OE) and interference-expressing (RNAi) transgenic potato plants were acquired using Agrobacterium-mediated transformation, respectively. Tissue specific analysis using Quantitative real-time polymerase chain reaction (qRT-PCR) demonstrated that the StDWF4 gene expressed in the leaves, stems and roots of the transgenic and un-transgenic (NT) plants, with specially increased (StDWF4-OE)/reduced (StDWF4-RNAi) expression in the roots. The content of malondialdehyde (MDA) in StDWF4-OE potato plants was lower than that of NT, and proline content was higher than that of NT. MDA and proline content in StDWF4-OE and NT under salt-stress was significantly higher than that of the control and was increased at different sampling times. The content of soluble protein, soluble sugar and the activities of superoxide dismutase (SOD), peroxidase (POD) and ascorbate peroxidase (APX) was higher in the StDWF4-OE plantlets at varied salt treatment time than in the NT potatoes. Reduction of H2O2 content in the StDWF4-OE plants was observed. All above plant physiology indicators in the StDWF4-RNAi potatoes showed opposite variation trends. The results proved that the overexpressing of StDWF4 in potato plantlets can enhance the salt resistance by alleviating the negative effects of salt-stress. However, its interference expression in potato plants depresses the salt resistance. The results lay the groundwork for intensive study of BRs regulation in potato growth and development, and will help us to reveal the molecular mechanisms of how the BRs signaling regulate potato salt tolerance.

Expression of the Galanthus nivalis agglutinin (GNA) gene in transgenic potato plants confers resistance to aphids.

Aphids, the largest group of sap-sucking pests, cause significant yield losses in agricultural crops worldwide every year. The massive use of pesticides to combat this pest causes severe damage to the environment, putting in risk the human health. In this study, transgenic potato plants expressing Galanthus nivalis agglutinin (GNA) gene were developed using CaMV 35S and ST-LS1 promoters generating six transgenic lines (35S1-35S3 and ST1-ST3 corresponding to the first and second promoter, respectively). Quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that the GNA gene was expressed in leaves, stems and roots of transgenic plants under the control of the CaMV 35S promoter, while it was only expressed in leaves and stems under the control of the ST-LS1 promoter. The levels of aphid mortality after 5 days of the inoculation in the assessed transgenic lines ranged from 20 to 53.3%. The range of the aphid population in transgenic plants 15 days after inoculation was between 17.0±1.43 (ST2) and 36.6±0.99 (35S3) aphids per plant, which corresponds to 24.9-53.5% of the aphid population in non-transformed plants. The results of our study suggest that GNA expressed in transgenic potato plants confers a potential tolerance to aphid attack, which appears to be an alternative against the use of pesticides in the future.

Transgenic potato plants expressing cry3A gene confer resistance to Colorado potato beetle.

The Colorado potato beetle (Leptinotarsa decemlineata Say, CPB) is a fatal pest, which is a quarantine pest in China. The CPB has now invaded the Xinjiang Uygur Autonomous Region and is constantly spreading eastward in China. In this study, we developed transgenic potato plants expressing cry3A gene. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that the cry3A gene expressed in leaves, stems and roots of the transgenic plants under the control of CaMV 35S promoter, while they expressed only in leaves and stems under the control of potato leaf and stem-specific promoter ST-LS1. The mortality of the larvae was higher (28% and 36%) on the transgenic plant line 35S1 on the 3rd and 4th days, and on ST3 (48%) on the 5th day after inoculation with instar larvae. Insect biomass accumulation on the foliage of the transgenic plant lines 35S1, 35S2 and ST3 was significantly lower (0.42%, 0.43% and 0.42%). Foliage consumption was lowest on transgenic lines 35S8 and ST2 among all plant foliage (7.47 mg/larvae/day and 12.46 mg/larvae/day). The different transgenic plant foliages had varied inhibition to larval growth. The survivors on the transgenic lines obviously were smaller than their original size and extremely weak. The transgenic potato plants with CPB resistance could be used to develop germplasms or varieties for controlling CPB damage and halting its spread in China.

Transcriptomic changes during tuber dormancy release process revealed by RNA sequencing in potato.

Potato tuber dormancy release is a critical development process that allows potato to produce new plant. The first Illumina RNA sequencing to generate the expressed mRNAs at dormancy tuber (DT), dormancy release tuber (DRT) and sprouting tuber (ST) was performed. We identified 26,639 genes including 5,912 (3,450 up-regulated while 2,462 down-regulated) and 3,885 (2,141 up-regulated while 1,744 down-regulated) genes were differentially expressed from DT vs DRT and DRT vs ST. The RNA-Seq results were further verified using qRT-PCR. We found reserve mobilization events were activated before the bud emergence (DT vs DRT) and highlighted after dormancy release (DRT vs ST). Overexpressed genes related to metabolism of auxin, gibberellic acid, cytokinin and barssinosteriod were dominated in DT vs DRT, whereas overexpressed genes involved in metabolism of ethylene, jasmonate and salicylate were prominent in DRT vs ST. Various histone and cyclin isoforms associated genes involved in cell division/cycle were mainly up-regulated in DT vs DRT. Dormancy release process was also companied by stress response and redox regulation, those genes related to biotic stress, cell wall and second metabolism was preferentially overexpressed in DRT vs ST, which might accelerate dormancy breaking and sprout outgrowth. The metabolic processes activated during tuber dormancy release were also supported by plant seed models. These results represented the first comprehensive picture of a large number of genes involved in tuber dormancy release process.

Identification of novel and conserved microRNAs related to drought stress in potato by deep sequencing.

MicroRNAs (miRNAs) are a group of small, non-coding RNAs that play important roles in plant growth, development and stress response. There have been an increasing number of investigations aimed at discovering miRNAs and analyzing their functions in model plants (such as Arabidopsis thaliana and rice). In this research, we constructed small RNA libraries from both polyethylene glycol (PEG 6,000) treated and control potato samples, and a large number of known and novel miRNAs were identified. Differential expression analysis showed that 100 of the known miRNAs were down-regulated and 99 were up-regulated as a result of PEG stress, while 119 of the novel miRNAs were up-regulated and 151 were down-regulated. Based on target prediction, annotation and expression analysis of the miRNAs and their putative target genes, 4 miRNAs were identified as regulating drought-related genes (miR811, miR814, miR835, miR4398). Their target genes were MYB transcription factor (CV431094), hydroxyproline-rich glycoprotein (TC225721), quaporin (TC223412) and WRKY transcription factor (TC199112), respectively. Relative expression trends of those miRNAs were the same as that predicted by Solexa sequencing and they showed a negative correlation with the expression of the target genes. The results provide molecular evidence for the possible involvement of miRNAs in the process of drought response and/or tolerance in the potato plant.