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Background Starch is the principle constituent of potato tubers and is of considerable importance for food and non-food applications. Its metabolism has been subject of extensive research over the past decades. Despite its importance, a description of the complete inventory of genes involved in starch metabolism and their genome organization in potato plants is still missing. Moreover, mechanisms regulating the expression of starch genes in leaves and tubers remain elusive with regard to differences between transitory and storage starch metabolism, respectively. This study aimed at identifying and mapping the complete set of potato starch genes, and to study their expression pattern in leaves and tubers using different sets of transcriptome data. Moreover, we wanted to uncover transcription factors co-regulated with starch accumulation in tubers in order to get insight into the regulation of starch metabolism. Results We identified 77 genomic loci encoding enzymes involved in starch metabolism. Novel isoforms of many enzymes were found. Their analysis will help to elucidate mechanisms of starch biosynthesis and degradation. Expression analysis of starch genes led to the identification of tissue-specific isoenzymes suggesting differences in the transcriptional regulation of starch metabolism between potato leaf and tuber tissues. Selection of genes predominantly expressed in developing potato tubers and exhibiting an expression pattern indicative for a role in starch biosynthesis enabled the identification of possible transcriptional regulators of tuber starch biosynthesis by co-expression analysis. Conclusions This study provides the annotation of the complete set of starch metabolic genes in potato plants and their genomic localizations. Novel, so far undescribed, enzyme isoforms were revealed. Comparative transcriptome analysis enabled the identification of tuber- and leaf-specific isoforms of starch genes. This finding suggests distinct regulatory mechanisms in transitory and storage starch metabolism. Putative regulatory proteins of starch biosynthesis in potato tubers have been identified by co-expression and their expression was verified by quantitative RT-PCR.

Reactivation of dormant meristems is of central importance for plant fitness and survival. Due to their large meristem size, potato (Solanum tuberosum) tubers serve as a model system to study the underlying molecular processes. The phytohormones cytokinins (CK) and gibberellins (GA) play important roles in releasing potato tuber dormancy and promoting sprouting, but their mode of action in these processes is still obscure. Here, we established an in vitro assay using excised tuber buds to study the dormancy-releasing capacity of GA and CK and show that application of gibberellic acid (GA₃) is sufficient to induce sprouting. In contrast, treatment with 6-benzylaminopurine induced bud break but did not support further sprout growth unless GA₃ was administered additionally. Transgenic potato plants expressing Arabidopsis (Arabidopsis thaliana) GA 20-oxidase or GA 2-oxidase to modify endogenous GA levels showed the expected phenotypical changes as well as slight effects on tuber sprouting. The isopentenyltransferase (IPT) from Agrobacterium tumefaciens and the Arabidopsis cytokinin oxidase/dehydrogenase1 (CKX) were exploited to modify the amounts of CK in transgenic potato plants. IPT expression promoted earlier sprouting in vitro. Strikingly, CKX-expressing tubers exhibited a prolonged dormancy period and did not respond to GA₃. This supports an essential role of CK in terminating tuber dormancy and indicates that GA is not sufficient to break dormancy in the absence of CK. GA₃-treated wild-type and CKX-expressing tuber buds were subjected to a transcriptome analysis that revealed transcriptional changes in several functional groups, including cell wall metabolism, cell cycle, and auxin and ethylene signaling, denoting events associated with the reactivation of dormant meristems.

The lipid biopolymer suberin plays a major role as a barrier both at plant-environment interfaces and in internal tissues, restricting water and nutrient transport. In potato (Solanum tuberosum), tuber integrity is dependent on suberized periderm. Using microarray analyses, we identified ABCG1, encoding an ABC transporter, as a gene responsive to the pathogenassociated molecular pattern Pep-13. Further analyses revealed that ABCG1 is expressed in roots and tuber periderm, as well as in wounded leaves. Transgenic ABCG1-RNAI potato plants with downregulated expression of ABCG1 display major alterations in both root and tuber morphology, whereas the aerial part of the ABCG1-RHAI plants appear normal. The tuber periderm and root exodermis show reduced suberin staining and disorganized cell layers. Metabolite analyses revealed reduction of esterified suberin components and hyperaccumulation of putative suberin precursors in the tuber periderm of RNA interference plants, suggesting that ABCG1 is required for the export of suberin components.

Isoamylases hydrolyse (1-6)-alpha-D-glucosidic linkages in starch and are involved in both starch granule formation and starch degradation. In plants, three isoamylase isoforms with distinct functions in starch synthesis (ISA1 and ISA2) and degradation (ISA3) have been described. Here, we created transgenic potato plants with simultaneously decreased expression of all three isoamylases using a chimeric RNAi construct targeting all three isoforms. Constitutive expression of the hairpin RNA using the 35S CaMV promoter resulted in efficient silencing of all three isoforms in leaves, growing tubers, and sprouting tubers. Neither plant growth nor tuber yield was effected in isoamylase-deficient potato lines. Interestingly, starch metabolism was found to be impaired in a tissue-specific manner. While leaf starch content was unaffected, tuber starch was significantly reduced. The reduction in tuber starch content in the transgenic plants was accompanied by a decrease in starch granules size, an increased sucrose content and decreased hexose levels. Despite the effects on granule size, only little changes in chain length composition of soluble and insoluble glucose polymers were detected. The transgenic tubers displayed an early sprouting phenotype that was accompanied by an increased level of sucrose in parenchyma cells below the outgrowing bud. Since high sucrose levels promote sprouting, we propose that the increased number of small starch granules may cause an accelerated turnover of glucan chains and hence a more rapid synthesis of sucrose. This observation links alterations in starch structure/degradation with developmental processes like meristem activation and sprout outgrowth in potato tubers.

Background Even though the process of potato tuber starch biosynthesis is well understood, mechanisms regulating biosynthesis are still unclear. Transcriptome analysis provides valuable information as to how genes are regulated. Therefore, this work aimed at investigating transcriptional regulation of starch biosynthetic genes in leaves and tubers of potato plants under various conditions. More specifically we looked at gene expression diurnally in leaves and tubers, during tuber induction and in tubers growing at different velocities. To determine velocity of potato tuber growth a new method based on X-ray Computed Tomography (X-ray CT) was established. Results Comparative transcriptome analysis between leaves and tubers revealed striking similarities with the same genes being differentially expressed in both tissues. In tubers, oscillation of granule bound starch synthase (GBSS) expression) was observed which could be linked to sucrose supply from source leaves. X-ray CT was used to determine time-dependent changes in tuber volume and the growth velocity was calculated. Although there is not a linear correlation between growth velocity and expression of starch biosynthetic genes, there are significant differences between growing and non-growing tubers. Co-expression analysis was used to identify transcription factors positively correlating with starch biosynthetic genes possibly regulating starch biosynthesis. Conclusion Most starch biosynthetic enzymes are encoded by gene families. Co-expression analysis revealed that the same members of these gene families are co-regulated in leaves and tubers. This suggests that regulation of transitory and storage starch biosynthesis in leaves and tubers, respectively, is surprisingly similar. X-ray CT can be used to monitor growth and development of belowground organs and allows to link tuber growth to changes in gene expression. Comparative transcriptome analysis provides a useful tool to identify transcription factors possibly involved in the regulation of starch biosynthesis.

Identification of molecular markers defining the end of tuber dormancy prior to visible sprouting is of agronomic interest for potato growers and the potato processing industry. In potato tubers, breakage of dormancy is associated with the reactivation of meristem function. In dormant meristems, cells are arrested in the G₁/G₀ phase of the cell cycle and re-entry into the G₁ phase followed by DNA replication during the S phase enables bud outgrowth. Deoxyuridine triphosphatase (dUTPase) is essential for DNA replication and was therefore tested as a potential marker for meristem reactivation in tuber buds. The corresponding cDNA clone was isolated from potato by PCR. The deduced amino acid sequence showed 94% similarity to the tomato homologue. By employing different potato cultivars, a positive correlation between dUTPase expression and onset of tuber sprouting could be confirmed. Moreover, gene expression analysis of tuber buds during storage time revealed an up-regulation of the dUTPase 1 week before visible sprouting occurred. Further analysis using an in vitro sprout assay supported the assumption that dUTPase is a good molecular marker to define the transition from dormant to active potato tuber meristems.

Reactivation of dormant meristems is of central importance for plant fitness and survival. Due to their large meristem size, potato (Solanum tuberosum) tubers serve as a model system to study the underlying molecular processes. The phytohormones cytokinins (CK) and gibberellins (GA) play important roles in releasing potato tuber dormancy and promoting sprouting, but their mode of action in these processes is still obscure. Here, we established an in vitro assay using excised tuber buds to study the dormancy-releasing capacity of GA and CK and show that application of gibberellic acid (GA(3)) is sufficient to induce sprouting. In contrast, treatment with 6-benzylaminopurine induced bud break but did not support further sprout growth unless GA(3) was administered additionally. Transgenic potato plants expressing Arabidopsis (Arabidopsis thaliana) GA 20-oxidase or GA 2-oxidase to modify endogenous GA levels showed the expected phenotypical changes as well as slight effects on tuber sprouting. The isopentenyltransferase (IPT) from Agrobacterium tumefaciens and the Arabidopsis cytokinin oxidase/dehydrogenase1 (CKX) were exploited to modify the amounts of CK in transgenic potato plants. IPT expression promoted earlier sprouting in vitro. Strikingly, CKX-expressing tubers exhibited a prolonged dormancy period and did not respond to GA(3). This supports an essential role of CK in terminating tuber dormancy and indicates that GA is not sufficient to break dormancy in the absence of CK. GA(3)-treated wild-type and CKX-expressing tuber buds were subjected to a transcriptome analysis that revealed transcriptional changes in several functional groups, including cell wall metabolism, cell cycle, and auxin and ethylene signaling, denoting events associated with the reactivation of dormant meristems.

Da Kartoffelknollen vor allem frisch verarbeitet bzw. verzehrt werden besteht das ganze Jahr über ein hoher Bedarf an frischem Erntegut. Aufgrund der klimatischen Bedingungen können Kartoffelknollen jedoch nicht das ganze Jahr über angebaut werden, so dass eine Langzeitlagerung der Knollen erforderlich ist. Nachdem das Auskeimen von Kartoffelknollen mit einer Remobilisierung von Speicherstoffen und Wasserverlust einhergeht, ist dies ein Hauptfaktor für eine Verringerung der Erntequalität. Daher ist es von großem Interesse die molekulare Regulation der Dormanz von Kartoffelknollen besser zu verstehen. Durch die Identifizierung von Kandidatengenen für potentielle Regulatoren der Knollendormanz bzw. Knollenmeristem-spezifischer Promotoren könnte es möglich werden, die Länge der Keimruhe von Kartoffelknollen gezielt zu beeinflussen.Wie zuvor bekannt kann die Keimruhe von Kartoffelknollen durch Zugabe von Gibberellin (GA) gebrochen werden. Daher sollte die Rolle dieses Phytohormons bei der Knollenkeimung näher untersucht werden. Mithilfe transgener Pflanzen mit erhöhtem bzw. verringertem endogenen GA-Gehalt konnte gezeigt werden, dass die Erhöhung des endogenen GA-Gehalts unter anderem zu einer leicht verfrühten Keimung und stark verlängerten Keimen führt. Pflanzen mit verringertem GA-Gehalt wiesen dagegen eine verlängerte Dormanzphase und stark verkürzte Keime auf. GA ist demnach sowohl an der Brechung der Keimruhe als auch am Elongationswachstum des Knollenkeimes beteiligt.Da über die molekularen Mechanismen der Knollenkeimung noch wenig bekannt ist, sollten Mikroarrayexperimente zu einem besseren Verständnis der Keimung von Kartoffelknollen beitragen. Zudem sollten Kandidatengene für potentielle Regulatoren der Knollendormanz ausgewählt und mithilfe transgener Pflanzen näher charakterisiert werden. Die Analyse globaler transkriptioneller Unterschiede in dormanten und keimenden Knollenaugen führte unter anderem zur Identifizierung von GARP (\"GA-regulated protein“) als potentiellem Regulator der Knollenkeimung. Knollen transgener Pflanzen mit erhöhter bzw. verringerter GARP-Genexpression zeigten jedoch kein verändertes Keimverhalten, was zu dem Schluss führte, dass GARP keinen Hauptregulator der Knollenkeimung darstellt. Zusätzlich führte die Transkriptomanalyse dormanter und keimender Knollenaugen zur Identifizierung der dUTPase als molekularem Marker für die Reaktivierung des Knollenmeristems.Die Analyse transkriptioneller Veränderungen während eines \"sprout release assay“-Experiments konnte zu einem besseren Verständnis der molekularen Veränderungen während der Knollenkeimung beitragen. Es zeigte sich, dass die Zellwandbiosynthese bzw. modifikation zu den sehr frühen Prozessen der Knollenkeimung zählt. Wie erwartet war auch der Wiedereintritt in den Zellzyklus entsprechend der Mikroarraydaten ein sehr früher Prozess der Initiation der Knollenkeimung. Eine frühe Induktion von Zellzyklusregulatoren des G1/ S-Phasenübergangs wurde mittels \"Real-Time“ PCR-Analysen verifiziert.Durch den Vergleich von Mikroarraydaten aus GA3- bzw. BA-induzierter Knollenkeimung und dormanten bzw. keimenden Knollenaugen konnten früh, generell bzw. spät während der Keimung regulierte Gene identifiziert werden. Aus diesen Ergebnissen wurde ein Modell über die molekularen Veränderungen während der Knollenkeimung abgeleitet. Zusätzlich wurde das KNOX-Gen StKn2 als möglicher Regulator der Knollenkeimung identifiziert und mithilfe transgener Pflanzen weiter charakterisiert. StKn2 ist sehr wahrscheinlich an der Reaktivierung des Knollenmeristems bei der Initiation der Keimung beteiligt.Durch vergleichende Analyse von Genexpressionsprofilen induzierter Stolone und aktiver Knollenmeristeme konnte die Hypothese bestätigt werden, dass die Knolleninduktion und die Knollenkeimung auf transkriptioneller Ebene eher antagonistisch reguliert werden. Mittels vergleichender Analysen von Transkriptionsprofilen aktiver Knollenmeristeme mit Mikroarraydaten aus verschiedenen Meristem-haltigen Kartoffelgeweben sollten gemeinsam und vor allem auch spezifisch exprimierte Gene identifiziert werden. Zuletzt wurden durch einen Vergleich von Mikroarraydaten von verschiedenen Zeitpunkten der Knollenkeimung und einer Reihe verschiedenster Kartoffelgewebe eine sehr wahrscheinlich spezifisch bei der Knollenkeimung exprimierte putative Pektinesterase (pPE) identifiziert.