The Retinoblastoma-related gene RBL901 can trigger drought response actions in potato.

KEY MESSAGE: This study provided new insights into response of potato to drought stress.

Glycoalkaloid Composition and Flavonoid Content as Driving Forces of Phytotoxicity in Diploid Potato.

Despite their advantages, biotechnological and omic techniques have not been applied often to characterize phytotoxicity in depth. Here, we show the distribution of phytotoxicity and glycoalkaloid content in a diploid potato population and try to clarify the source of variability of phytotoxicity among plants whose leaf extracts have a high glycoalkaloid content against the test plant species, mustard. Six glycoalkaloids were recognized in the potato leaf extracts: solasonine, solamargine, α-solanine, α-chaconine, leptinine I, and leptine II. The glycoalkaloid profiles of the progeny of the group with high phytotoxicity differed from those of the progeny of the group with low phytotoxicity, which stimulated mustard growth. RNA sequencing analysis revealed that the upregulated flavonol synthase/flavonone 3-hydroxylase-like gene was expressed in the progeny of the low phytotoxicity group, stimulating plant growth. We concluded that the metabolic shift among potato progeny may be a source of different physiological responses in mustard. The composition of glycoalkaloids, rather than the total glycoalkaloid content itself, in potato leaf extracts, may be a driving force of phytotoxicity. We suggest that, in addition to glycoalkaloids, other metabolites may shape phytotoxicity, and we assume that these metabolites may be flavonoids.

The MYB33, MYB65, and MYB101 transcription factors affect Arabidopsis and potato responses to drought by regulating the ABA signaling pathway.

Drought is one of the main climate threats limiting crop production. Potato is one of the four most important food crop species worldwide and is sensitive to water shortage. The CBP80 gene was shown to affect Arabidopsis and potato responses to drought by regulating the level of microRNA159 and, consequently, the levels of the MYB33 and MYB101 transcription factors (TFs). Here, we show that three MYB TFs, MYB33, MYB65, and MYB101, are involved in plant responses to water shortage. Their downregulation in Arabidopsis causes stomatal hyposensitivity to abscisic acid (ABA), leading to reduced tolerance to drought. Transgenic Arabidopsis and potato plants overexpressing these genes, with a mutated recognition site in miR159, show hypersensitivity to ABA and relatively high tolerance to drought conditions. Thus, the MYB33, MYB65, and MYB101 genes may be potential targets for innovative breeding to obtain crops with relatively high tolerance to drought.

Transcriptomic and proteomic data provide new insights into cold-treated potato tubers with T- and D-type cytoplasm.

MAIN CONCLUSION: Tuber-omics in potato with the T- and D-types of cytoplasm showed different sets of differentially expressed genes and proteins in response to cold storage. For the first time, we report differences in gene and protein expression in potato (Solanum tuberosum L.) tubers possessing the T- or D-type cytoplasm. Two F1 diploid reciprocal populations, referred to as T and D, were used. The pooling strategy was applied for detection of differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) in tubers consisting of extreme chip colour after cold storage. RNA and protein bulks were constructed from contrasting phenotypes. We recognized 48 and 15 DEGs for the T and D progenies, respectively. DEPs were identified in the amyloplast and mitochondrial fractions. In the T-type cytoplasm, only 2 amyloplast-associated and 5 mitochondria-associated DEPs were detected. Of 37 mitochondria-associated DEPs in the D-type cytoplasm, there were 36 downregulated DEPs in the dark chip colour bulks. These findings suggest that T- and D-type of cytoplasm might influence sugar accumulation in cold-stored potato tubers in different ways. We showed that the mt/nucDNA ratio was higher in D-possessing tubers after cold storage than in T progeny. For the D-type cytoplasm, the pt/nucDNA ratio was higher for tubers characterized by dark chip colour than for those with light chip colour. Our findings suggest that T- and D-type cytoplasm might influence sugar accumulation in cold-stored potato tubers in different ways.

Transcriptional and proteomic insights into phytotoxic activity of interspecific potato hybrids with low glycoalkaloid contents.

BACKGROUND: Glycoalkaloids are bioactive compounds that contribute to the defence response of plants against herbivore attack and during pathogenesis. Solanaceous plants, including cultivated and wild potato species, are sources of steroidal glycoalkaloids. Solanum plants differ in the content and composition of glycoalkaloids in organs. In wild and cultivated potato species, more than 50 steroidal glycoalkaloids were recognized. Steroidal glycoalkaloids are recognized as potential allelopathic/phytotoxic compounds that may modify the growth of target plants. There are limited data on the impact of the composition of glycoalkaloids on their phytotoxic potential. RESULTS: The presence of α-solasonine and α-solamargine in potato leaf extracts corresponded to the high phytotoxic potential of the extracts. Among the differentially expressed genes between potato leaf bulks with high and low phytotoxic potential, the most upregulated transcripts in sample of high phytotoxic potential were anthocyanin 5-aromatic acyltransferase-like and subtilisin-like protease SBT1.7-transcript variant X2. The most downregulated genes were carbonic anhydrase chloroplastic-like and miraculin-like. An analysis of differentially expressed proteins revealed that the most abundant group of proteins were those related to stress and defence, including glucan endo-1,3-beta-glucosidase acidic isoform, whose expression level was 47.96× higher in potato leaf extract with low phytotoxic. CONCLUSIONS: The phytotoxic potential of potato leaf extract possessing low glycoalkaloid content is determined by the specific composition of these compounds in leaf extract, where α-solasonine and α-solamargine may play significant roles. Differentially expressed gene and protein profiles did not correspond to the glycoalkaloid biosynthesis pathway in the expression of phytotoxic potential. We cannot exclude the possibility that the phytotoxic potential is influenced by other compounds that act antagonistically or may diminish the glycoalkaloids effect.

eQTL mapping of the 12S globulin cruciferin gene PGCRURSE5 as a novel candidate associated with starch content in potato tubers.

Tuber starch content (TSC) is a very important trait in potato (Solanum tuberosum L.). This study is the first to use expression quantitative trait loci (eQTL) mapping of transcript-derived markers for TSC in potato. Thirty-four differentially expressed genes were selected by comparing the RNA-seq data of contrasting bulked segregants. For the 11 candidate genes, we determined their relative expression levels across the segregating diploid potato population using RT-qPCR. We detected 36 eQTL as candidate genes distributed on all twelve potato chromosomes, and nine of them overlapped with QTL for TSC. Peaks for two eQTL, eAGPaseS-a and ePGRCRURSE5, were close to the corresponding loci of the large subunit of ADP-glucose pyrophosphorylase (AGPaseS-a) and the 12S globulin cruciferin gene (PGCRURSE5), respectively. The eQTL peaks for AGPaseS-a and PGRCRURSE5 explained 41.0 and 28.3% of the phenotypic variation at the transcript level. We showed the association of the DNA markers for AGPaseS-a and PGRCRURSE5 with QTL for TSC, and significant correlation between the expression level of PGRCRURSE5 and TSC. We did not observe a significant correlation between the expression level of AGPaseS-a and TSC. We concluded that the cruciferin gene PGRCRURSE5 is a novel candidate involved in the regulation of starch content in potato tubers.

Extreme resistance to Potato virus Y in potato carrying the Rysto gene is mediated by a TIR-NLR immune receptor.

Potato virus Y (PVY) is a major potato (Solanum tuberosum L.) pathogen that causes severe annual crop losses worth billions of dollars worldwide. PVY is transmitted by aphids, and successful control of virus transmission requires the extensive use of environmentally damaging insecticides to reduce vector populations. Rysto , from the wild relative S. stoloniferum, confers extreme resistance (ER) to PVY and related viruses and is a valuable trait that is widely employed in potato resistance breeding programmes. Rysto was previously mapped to a region of potato chromosome XII, but the specific gene has not been identified to date. In this study, we isolated Rysto using resistance gene enrichment sequencing (RenSeq) and PacBio SMRT (Pacific Biosciences single-molecule real-time sequencing). Rysto was found to encode a nucleotide-binding leucine-rich repeat (NLR) protein with an N-terminal TIR domain and was sufficient for PVY perception and ER in transgenic potato plants. Rysto -dependent extreme resistance was temperature-independent and requires EDS1 and NRG1 proteins. Rysto may prove valuable for creating PVY-resistant cultivars of potato and other Solanaceae crops.

Quantitative trait loci for starch-corrected chip color after harvest, cold storage and after reconditioning mapped in diploid potato.

The objective of this study was to map the quantitative trait loci (QTLs) for chip color after harvest (AH), cold storage (CS) and after reconditioning (RC) in diploid potato and compare them with QTLs for starch-corrected chip color. Chip color traits AH, CS, and RC significantly correlated with tuber starch content (TSC). To limit the effect of starch content, the chip color was corrected for TSC. The QTLs for chip color (AH, CS, and RC) and the starch-corrected chip color determined with the starch content after harvest (SCAH), after cold storage (SCCS) and after reconditioning (SCRC) were compared to assess the extent of the effect of starch and the location of genetic factors underlying this effect on chip color. We detected QTLs for the AH, CS, RC and starch-corrected traits on ten potato chromosomes, confirming the polygenic nature of the traits. The QTLs with the strongest effects were detected on chromosomes I (AH, 0 cM, 11.5% of variance explained), IV (CS, 43.9 cM, 12.7%) and I (RC, 49.7 cM, 14.1%). When starch correction was applied, the QTLs with the strongest effects were revealed on chromosomes VIII (SCAH, 39.3 cM, 10.8% of variance explained), XI (SCCS, 79.5 cM, 10.9%) and IV (SCRC, 43.9 cM, 10.8%). Applying the starch correction changed the landscape of QTLs for chip color, as some QTLs became statistically insignificant, shifted or were refined, and new QTLs were detected for SCAH. The QTLs on chromosomes I and IV were significant for all traits with and without starch correction.

Comparative proteomic analysis of resistant and susceptible potato cultivars during Synchytrium endobioticum infestation.

MAIN CONCLUSION: We report the first comparative study of protein expression profiles in tuber sprouts between Katahdin-derived potato cultivars resistant and susceptible to Synchytrium endobioticum. Synchytrium endobioticum causes wart disease in potato (Solanum tuberosum L.) and is considered as the most important quarantine pathogen in almost all countries where potatoes are grown. We performed a comparative analysis of differentially expressed proteins in the tuber sprouts of potato cultivars differing in resistance to pathotype 1(D1) of S. endobioticum using two-dimensional electrophoresis (2-DE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) approaches. Bulks prepared from two resistant (Calrose and Humalda) and three susceptible (Sebago, Seneca and Wauseon) potato cultivars were studied. When protein profiles were compared between mock- and S. endobioticum-inoculated sprouts, 35 and 63 protein spots, indicating qualitative or quantitative differences, were detected in the resistant and susceptible cultivars, respectively. In turn, 24 proteins associated with resistance to S. endobioticum were revealed by comparison of the resistant and susceptible bulks. These proteins were changed in a constitutive or induced manner and were grouped into four categories: stress and defence, cell structure, protein turnover, and metabolism. Among the 13 proteins classified into the stress and defence group, seven proteins were related to heat-shock proteins (HSPs)/chaperone factors. In addition, four proteins, S-adenosyl-L-homocysteine hydrolase-like, superoxide dismutase [Mn], inactive patatin-3-Kuras 1 and patatin-15, were induced in the resistant bulk; whereas two proteins, patatin-01 and nucleoredoxin 1, showed significant differences in expression between the S. endobioticum-inoculated resistant and susceptible bulks. The detection of such a large number of S. endobioticum-mediated proteins representing the HSP70, HSP60 and HSP20 families suggests their significant role in restricting wart disease in potato tubers.

The effect of drought stress on the leaf relative water content and tuber yield of a half-sib family of 'Katahdin'-derived potato cultivars.

Drought tolerance in plants is a complex trait involving morphological, physiological, and biochemical mechanisms. Hundreds of genes underlie the response of plants to the stress. For crops, selecting cultivars that can produce economically significant yields under drought is a priority. Potato (Solanum tuberosum L.) is considered as drought sensitive crop, although cultivar-dependent differences in tolerance have been described. Cultivar 'Katahdin' possesses many appropriate characteristics and is widely used for breeding purposes worldwide; it also has enhanced tolerance to drought stress. In this study, we evaluated cv. 'Katahdin' and a half-sib family of 17 Katahdin-derived cultivars for leaf relative water content (RWC) and tuber yield under drought stress. The yields of cultivars 'Wauseon', 'Katahdin', 'Magura', 'Calrose', and 'Cayuga' did not significantly decline under drought stress. Among these five, Wauseon exhibited the lowest reduction in both tuber yield and relative water content under water shortage. The data showed that 'Wauseon' is the most attractive cultivar for studies of molecular and physiological processes under drought and for potato breeding due to low yield losses that correspond with high RWC values. This cultivar can serve as a reservoir of potentially useful genes to develop cultivars with enhanced tolerance to this abiotic stress.

Mapping of quantitative trait loci for tuber starch and leaf sucrose contents in diploid potato.

KEY MESSAGE: Most QTL for leaf sucrose content map to positions that are similar to positions of QTL for tuber starch content in diploid potato. In the present study, using a diploid potato mapping population and Diversity Array Technology (DArT) markers, we identified twelve quantitative trait loci (QTL) for tuber starch content on seven potato chromosomes: I, II, III, VIII, X, XI, and XII. The most important QTL spanned a wide region of chromosome I (42.0­104.6 cM) with peaks at 63 and 84 cM which explained 17.6 and 19.2% of the phenotypic variation, respectively. ADP-glucose pyrophosphorylase (AGPase) is the key enzyme for starch biosynthesis. The gene encoding the large subunit of this enzyme, AGPaseS-a, was localized to chromosome I at 102.3 cM and accounted for 15.2% of the variance in tuber starch content. A more than 100-fold higher expression of this gene was observed in RT-qPCR assay in plants with the marker allele AGPaseS-a1334. This study is the first to report QTL for sucrose content in potato leaves. QTL for sucrose content in leaves were located on eight potato chromosomes: I, II, III, V, VIII, IX, X and XII. In 5-week-old plants, only one QTL for leaf sucrose content was detected after 8 h of darkness; four QTL were detected after 8 h of illumination. In 11-week-old plants, 6 and 3 QTL were identified after dark and light phases, respectively. Of fourteen QTL for leaf sucrose content, eleven mapped to positions that were similar to QTL for tuber starch content. These results provide genetic information for further research examining the relationships between metabolic carbon molecule sources and sinks in potato plants.

Novel candidate genes AuxRP and Hsp90 influence the chip color of potato tubers.

Potato (Solanum tuberosum L.) tubers exhibit significant variation in reducing sugar content directly after harvest, cold storage and reconditioning. Here, we performed QTL analysis for chip color, which is strongly influenced by reducing sugar content, in a diploid potato mapping population. Two QTL on chromosomes I and VI were detected for chip color after harvest and reconditioning. Only one region on chromosome VI was linked with cold-induced sweetening. Using the RT-PCR technique, we showed differential expression of the auxin-regulated protein (AuxRP) gene. The AuxRP transcript was presented in light chip color parental clone DG 97-952 and the RNA progeny of the bulk sample consisting of light chip color phenotypes after cold storage. This amplicon was absent in dark chip parental clone DG 08-26/39 and the RNA bulk sample of dark chip progeny. Genetic variation of AuxRP explained up to 16.6 and 15.2 % of the phenotypic variance after harvest and 3 months of storage at 4 °C, respectively. Using an alternative approach, the RDA-cDNA method was used to recognize 25 gene sequences, of which 11 could be assigned to potato chromosome VI. One of these genes, Heat-shock protein 90 (Hsp90), demonstrated higher mRNA and protein expression in RT-qPCR and western blotting assays in the dark chip color progeny bulk sample compared with the light chip color progeny bulk sample. Our study, for the first time, suggests that the AuxRP and Hsp90 genes are novel candidate genes capable of influencing the chip color of potato tubers.

Potato Annexin STANN1 Promotes Drought Tolerance and Mitigates Light Stress in Transgenic Solanum tuberosum L. Plants.

Annexins are a family of calcium- and membrane-binding proteins that are important for plant tolerance to adverse environmental conditions. Annexins function to counteract oxidative stress, maintain cell redox homeostasis, and enhance drought tolerance. In the present study, an endogenous annexin, STANN1, was overexpressed to determine whether crop yields could be improved in potato (Solanum tuberosum L.) during drought. Nine potential potato annexins were identified and their expression characterized in response to drought treatment. STANN1 mRNA was constitutively expressed at a high level and drought treatment strongly increased transcription levels. Therefore, STANN1 was selected for overexpression analysis. Under drought conditions, transgenic potato plants ectopically expressing STANN1 were more tolerant to water deficit in the root zone, preserved more water in green tissues, maintained chloroplast functions, and had higher accumulation of chlorophyll b and xanthophylls (especially zeaxanthin) than wild type (WT). Drought-induced reductions in the maximum efficiency and the electron transport rate of photosystem II (PSII), as well as the quantum yield of photosynthesis, were less pronounced in transgenic plants overexpressing STANN1 than in the WT. This conferred more efficient non-photochemical energy dissipation in the outer antennae of PSII and probably more efficient protection of reaction centers against photooxidative damage in transgenic plants under drought conditions. Consequently, these plants were able to maintain effective photosynthesis during drought, which resulted in greater productivity than WT plants despite water scarcity. Although the mechanisms underlying this stress protection are not yet clear, annexin-mediated photoprotection is probably linked to protection against light-induced oxidative stress.

Hypersensitive response to Potato virus Y in potato cultivar Sárpo Mira is conferred by the Ny-Smira gene located on the long arm of chromosome IX.

Potato virus Y (PVY, Potyvirus) is the fifth most important plant virus worldwide in terms of economic and scientific impact. It infects members of the family Solanaceae and causes losses in potato, tomato, tobacco, pepper and petunia production. In potato and its wild relatives, two types of resistance genes against PVY have been identified. While Ry genes confer symptomless extreme resistance, Ny genes cause a hypersensitive response visible as local necrosis that may also be able to prevent the virus from spreading under certain environmental conditions. The potato cultivar Sárpo Mira originates from Hungary and is highly resistant to PVY, although the source of this resistance remains unknown. We show that cv. Sárpo Mira reacts with a hypersensitive response leading to necrosis after PVYNTN infection in detached leaf, whole plant and grafting assays. The hypersensitivity to PVYNTN segregated amongst 140 individuals of tetraploid progeny of cvs. Sárpo Mira × Maris Piper in a 1:1 ratio, indicating that it was conferred by a single, dominant gene in simplex. Moreover, we identified five DNA markers linked to this trait and located the underlying locus (Ny-Smira) to the long arm of potato chromosome IX. This position corresponds to the location of the Rychc and Ny-1 genes for PVY resistance. A simple PCR marker, located 1 cM from the Ny-Smira gene, can be recommended for selection of PVY-resistant progeny of cv. Sárpo Mira.

Ny-1 and Ny-2 genes conferring hypersensitive response to potato virus Y (PVY) in cultivated potatoes: mapping and marker-assisted selection validation for PVY resistance in potato breeding.

Potato virus Y (PVY) is one of the most important viruses affecting potato (Solanum tuberosum) production. In this study, a novel hypersensitive response (HR) gene, Ny-2, conferring resistance to PVY was mapped on potato chromosome XI in cultivar Romula. In cultivars Albatros and Sekwana, the Ny-1 gene was mapped on chromosome IX. In cv. Romula, the local lesions appeared in leaves inoculated with the PVYN-Wi isolate at 20 and 28 °C; PVY systemic infections were only occasionally observed at the higher temperature. In cvs. Albatros and Sekwana, expression of the necrotic reaction to virus infection was temperature-dependent. PVYN-Wi was localized at 20 °C; at 28 °C, the systemic, symptomless infection was observed. We developed the B11.61600 marker co-segregating with Ny-2 and the S1d11 marker specific for the Ny-1 gene. Fifty potato cultivars were tested with markers B11.6 and S1d11 and marker SC895 linked to the Ny-1 gene in cv. Rywal. These results indicated the utility of these markers for marker-assisted selection of HR-like PVY resistance in potato breeding programs.

Down-regulation of CBP80 gene expression as a strategy to engineer a drought-tolerant potato.

Developing new strategies for crop plants to respond to drought is crucial for their innovative breeding. The down-regulation of nuclear cap-binding proteins in Arabidopsis renders plants drought tolerant. The CBP80 gene in the potato cultivar Desiree was silenced using artificial microRNAs. Transgenic plants displayed a higher tolerance to drought, ABA-hypersensitive stomatal closing, an increase in leaf stomata and trichome density, and compact cuticle structures with a lower number of microchannels. These findings were correlated with a higher tolerance to water stress. The level of miR159 was decreased, and the levels of its target mRNAs MYB33 and MYB101 increased in the transgenic plants subjected to drought. Similar trends were observed in an Arabidopsis cbp80 mutant. The evolutionary conservation of CBP80, a gene that plays a role in the response to drought, suggests that it is a candidate for genetic manipulations that aim to obtain improved water-deficit tolerance of crop plants.

[Starch metabolism in potato tubers]. / Metabolizm skrobi w bulwach ziemniaka.

Sucrose is a substrate for starch biosynthesis, unloaded symplastically into the developing potato tubers. Sucrose is converted to glucose-6-phosphate in the cytosol and exported into the amyloplasts. Starch may be degraded either hydrolytically or phosphorolytically. Glucose and maltose are products of hydrolytic starch breakdown in the sprouting tubers. Glucose phosphates are products of the phosphorolytic activity, metabolized to glucose and fructose in cold-stored tubers. Development of molecular tools for assaying potato gene function provide opportunities to receive genetic progress in the sugar-starch potato breeding.

Endonuclease restriction of SCAR amplicons SC811 is required to identify Ns-false-positive markers in PVS-susceptible potato cultivars.

The Ns gene confers resistance of potato to Potato virus S (PVS). Sixteen German and Dutch potato cultivars, all registered in Poland, were found to be susceptible to PVS infection. However, scoring of the cultivars for the presence of the Ns-linked SCAR marker SC811(454) revealed additional amplicons with a similar electrophoretic migration rate as that of SC811(454), which resulted in ambiguous determination of the genotype at the Ns locus. MboI or FokI treatment of the PCR products allowed to detect their Ns-unspecificity in PVS-susceptible potato cultivars.

Genetic relationship between Polish and Chinese strains of the mushroom Agaricus bisporus (Lange) Sing., determined by the RAPD method.

The genetic relationship between twenty-six strains of Agaricus bisporus were analysed by the RAPD (random amplified polymorphic DNA) method. DNA amplification was performed with the use of twelve arbitrary 10-mer primers. Four primers, which gave polymorphic band patterns were chosen for RAPD analysis. In total, they gave 24 distinguishable bands, of which nine were polymorphic. The conducted research showed that there is a great genetic similarity among the examined strains. Low polymorphism of the strains may be a proof of a limited genetic pool used in the cultivation of those strains.