No description available.

To gain an understanding of the transcriptional regulation of sucrose transport and metabolism in wheat and to examine their relationships with dry matter accumulation in seeds, this study has characterized the expression patterns of two sucrose transporters (TaSUT1 and TaSUT2) and two sucrose synthase (TaSuSy1 and TaSuSy2) genes in five spring wheat cultivars at different seed developmental stages. Cultivar ‘AC Andrew’ with higher TaSUT1 and TaSuSy2 transcript abundance particularly during the early grain filling stage, exhibited higher dry grain weight than other cultivars. The result, overall, suggests the significance of coordinated expression between TaSUT1 and TaSuSy2 for grain growth. This study also demonstrated the seed specificity of high molecular weight glutenin promoter subunit Dy10, which can be used as an important tool to drive the expression of any sucrose and starch related genes specifically in wheat grains to further our understanding of carbon partitioning and/or increase wheat starch yield.

To gain an understanding of the transcriptional regulation of sucrose transport and metabolism in wheat and to examine their relationships with dry matter accumulation in seeds, this study has characterized the expression patterns of two sucrose transporters (TaSUT1 and TaSUT2) and two sucrose synthase (TaSuSy1 and TaSuSy2) genes in five spring wheat cultivars at different seed developmental stages. Cultivar ‘AC Andrew’ with higher TaSUT1 and TaSuSy2 transcript abundance particularly during the early grain filling stage, exhibited higher dry grain weight than other cultivars. The result, overall, suggests the significance of coordinated expression between TaSUT1 and TaSuSy2 for grain growth. This study also demonstrated the seed specificity of high molecular weight glutenin promoter subunit Dy10, which can be used as an important tool to drive the expression of any sucrose and starch related genes specifically in wheat grains to further our understanding of carbon partitioning and/or increase wheat starch yield.

Thesis (MSc (Botany and Zoology))--University of Stellenbosch, 2005. / The main substrate use during shoot establishment from the lateral bud of sugarcane setts and enzymes involved in sucrose metabolism were investigated in the shoots and the internodes acting as source of carbohydrates. Radiolabelling studies were conducted to investigate the metabolism of sucrose and glucose during shoot establishment. The internode’s total dry mass over the 21-day of shoot establishment period was reduced by 25% and 30% in plants incubated in dark/light and total darkness, respectively.

No description available.

Thesis (MSc (Biochemistry))--Stellenbosch University, 2008. / The aim of this study is the theoretical investigation of carbon fixation in sugarcane leaves. Sugarcane has a well known reputation for accumulating sucrose in the stalk to levels as high as 650 mM, almost a fifth of the plant’s fresh weight. Although this is an efficient accumulating mechanism, there is an even more efficient ‘carbon pump’ found in C4 plants. This is a well documented carbon concentrating mechanism and one of the first to be studied. However scientists are still trying to understand the carboxylating mechanism and the regulation thereof. It has been speculated that this mechanism is at its saturation level and elevating carbon dioxide will have little or no effect on further carbon fixation. Futher, studies suggest that the sucrose accumulating sink is able to regulate photosynthesis. Therefore a regulatory mechanism should exist from the sink to carbon fixation in order for such regulation to occur. Thework in this thesis therefore lays the foundation for investigating regulation of photosynthesis. The field of systems biology is the study of cellular networks by assemblingmodels. Pathways are considered as systems and notmerely collections of single components. This allows the interaction of pathway metabolites and the regulation that they have on one another to be studied. The questions asked pertaining to a pathway, will determine the types of model analysis. Structural analysis is useful for studying stoichiometric models, determining characteristics like energy consumption, futile cycles and valid pathways through a system at steady-state. Kinetic analysis on the other hand, gives insight into system dynamics and the control exerted by the system components, predicting time-course and steady states. In this thesis we begin to investigate photosynthesis in sugarcane leaves and the role it has in accumulating sucrose in the plant. Firstly, a structural model was developed incorporating carbon fixation, sucrose production in the leaf and subsequent transport of sucrose to the storage parenchyma and accumulation. The model was analysed using elementary mode analysis, showing that there are twelve routes for producing sucrose with no pathway beingmore energy efficient than any other. Further, it highlighted a futile cycle transporting triose phosphates and phosphoglycerate between the two photosynthetic compartments in the leaf. In the storage parenchyma, manymore futile cycleswere revealed,many of them energetically wasteful. Three other sets of elementary modes describe sucrose’s destination in either the vacuole or use in glycolysis or fibre formation, each with a different amount of required energy equivalents. The fourth set describes how sucrose cannot be converted to fibre precursors without also being used for glycolyis building blocks. Secondly, a kinetic model of carbon fixation in the leaf was assembled with the primary goal of characterising thismoiety-conserved cycle. This included the collation of kinetic data, incorporating volumes of the compartments and the areas of the location of the transporters into the model. This model was then analysed using metabolic control analysis. The model was able to predict metabolite concentration in the pathway at steady-state which were compared to those found experimentally. However, modifications need to be made to the model before further analysis is done so that the model predicted values match the experimental values more accurately. Time course analysis and response coefficients were also calculated for the carbon fixation cycle. Thework in this thesis therefore paves the way for understanding photosynthesis and its regulation in sugarcane leaves. Such work has the potential to pinpoint genetic engineering target points, allowing for better hybrid selection and propagation.

Dissertation (PhD) -- University of Stellenbosch, 2004. / ENGLISH ABSTRACT: In most plants the efficiency of sucrose production and the systemic distribution thereof are the major determinants of growth, development and yield. The factors governing sugar partitioning co-ordinate its distribution in response to intrinsic and environmental signals. These factors include sugar transporters and invertases as well as metabolites, including sucrose and glucose, which function as signalling molecules to modulate gene expression. The genetic transformation of plants and the subsequent development of transgenic lines with disturbed sugar metabolism have made an unprecedented impact on the study of sugar translocation and -partitioning. For instance, the transformation of plants with a yeast-derived invertase targeted to different subcellular compartments has led to the elucidation of several key aspects of sugar metabolism, including phloem loading mechanisms, the regulation of photosynthesis by sugars, the importance of sugar-metabolism compartmentation with regards to sucrose biosynthesis, storage and distribution, as well as the role of cell-wall invertase in phloem unloading and sink strength. In this study, a similar strategy of transgenic plant analysis was employed to expand our insight into the regulation of sugar partitioning. The yeast-invertase Suc2 gene, from Saccharomyces cere visiae , was overexpressed in either the cytosol, vacuole or apoplast of transgenic tobacco plants. These transgenic lines displayed varying increases in invertase activity, altered sugar levels and consequently disturbed sink-source interactions and sugar partitioning. Transgenic lines overproducing the yeast-derived invertase in either the vacuole (Vac-Inv) or apoplast (Apo-Inv) were utilised to analyse the effect of the altered sugar levels in sink and source organs on the expression of sugar transporters, as well as the endogenous cell wall invertase and inhibitors in these plants. Transcript levels of the sucrose transporter NtSUT1 and hexose transporter NtMST1 encoding genes increased significantly in the source leaves and roots of Vac-Inv lines, whereas increased NtMst1 transcript levels were also detected in the roots of Apo-Inv lines. The increased mRNA levels could be correlated to the altered invertase activities and sugar levels in these tissues. It is concluded that NtSUT1 and NtMST1 are differentially regulated by sucrose and/or hexose content on a transcriptional level. Furthermore, the regulatory effect of the altered sugar levels on transporter expression depended on the subcellular compartment in which the yeast invertase was expressed. It would seem that the subcellular compartmentation of sugar metabolism is also fundamental to the regulation of sugar partitioning. The transcription levels of the endogenous cell wall invertase (CWt) and cell wall invertase inhibitor (Cwi-Inh) genes were examined in the various tissues of Apo-Inv and Vac-Inv lines at both the vegetative and flowering growth stages. In comparison with the control lines, the various tissues of the Apo-Inv and Vac-Inv lines displayed altered Cwi and Cwi-Inh expression levels, depending on the sink-source status and growth stage. However, no obvious correlation between the Cwi and Cwi-Inh expression levels and soluble sugar content of these tissues was found. It is suggested that the post-transcriptional and post-translation control of these proteins by sugars might play an important role in their regulation. Analysis of the Cwi:Cwi-lnh mRNA ratio and growth observations of the various tissues of control as well as Apo-Inv and Vac-Inv lines indicated that this transcription ratio could be an accurate indicator of the sink strength of sink organs. In addition, the influence of sink-source interactions on sugar partitioning was investigated. Reciprocal grafting between Apo-Inv and control lines resulted in scions with an altered sucrose metabolism in either the sink or source organs. These scions were subjected to biomass distribution, soluble sugar quantification and C4C]- radiolabelling experiments. The latter revealed an unaltered state of sugar partitioning from the above-ground tissues of the Apo/GUS scions and a significant shift in sugar partitioning towards the roots of the GUS/Apo scions in comparison to the control GUS/GUS scions. Phenotypic changes, opposite to those observed in Apo-Inv lines expressing the heterologous invertase in both sink and source organs, could initially be observed in the GUS/Apo and Apo/GUS scions. However, no significant differences in phenotype or biomass distribution could be observed between the mature GUS/Apo, Apo/GUS and GUS/GUS scions seven weeks postgrafting. This inconsistency between phenotype and sugar partitioning might be explained by an increase in the respiration rate of the tissues as supported by the soluble sugar content. These results highlight the complexity and adaptability of sucrose metabolism and sugar partitioning. In addition, it confirms that sugar partitioning can be modulated by sink-source interactions and emphasise the importance of invertases in the regulation of sugar partitioning through its ability to alter sink strength. This study forms part of the rapidly expanding initiative to unravel the control mechanisms of sugar partitioning. The results obtained in this study confirmed again that the introduction and expression of a single heterologous gene in transgenic plants could provide significant insight into the regulation of this process. It was shown here that the expression of sugar transporters is closely regulated by sugar levels and therefore fulfils a vital function in sugar sensing and consequently the regulation of sugar partitioning. The data presented in this study also demonstrated the intricate and flexible nature of the relationship that exists between sugar metabolism, partitioning and growth phenomena. / AFRIKAANSE OPSOMMING: Die doeltreffendheid van sukroseproduksie, tesame met die sistemiese verspreiding daarvan, is die vernaamste faktore wat die groei, ontwikkeling en opbrengsvermoë van die meeste plante bepaal. Die faktore wat suikerverdeling beheer, funksioneer om suikerverspreiding te koordineer in reaksie op beide inherente- en omgewingsseine. Hierdie faktore sluit suikertransporters en invertases in, asook metaboliete soos sukrose en glukose wat funksioneer as seinmolekule in die modulering van geenuitdrukking. Die genetiese transformasie van plante en die gevolglike daarstelling van transgeniese lyne met veranderde suikermetabolismes het 'n beduidende inwerking op die bestudering van suikervervoer en -verdeling gehad. Byvoorbeeld, die transformasie van plante met 'n gis-invertase geteiken na verskillende sub-sellulêre kompartemente, het tot die toeligting van verskeie aspekte van suikermetabolisme gelei, insluitende dié van floëemladingsmeganismes, die regulering van fotosintese deur suikers, die belang van kompartementalisering ten opsigte van sukrosebiosintese, -opberging en -verspreiding, en die rol van selwand-invertases in floëemontlaaiing en swelgpuntkrag. In hierdie studie is van soortgelyke transgeniese plantontledings gebruik gemaak om 'n dieper insig tot die regulering van suikerverdeling te verkry. Die gis-invertase Suc2 geen, afkomstig van Saccharomyces cerevisiae, is ooruitgedruk in óf die sitosol, vakuool óf apoplastiese ruimte van transgeniese tabakplante. Hierdie transgeniese lyne het wisselende toenames in invertase-aktiwiteite en veranderde suikervlakke getoon, asook gevolglike versteurde bron-swelgpunt interaksies en suikerverdeling. Transgeniese lyne met ooruitdrukking van die gis-invertase in óf die vakuool (Vac-Inv) óf die apoplast (Apo-Inv) is gebruik om die gevolg van die veranderde suikervlakke in bron- en swelgpuntorgane op die uitdrukking van suikertransporters, asook die endogene selwand-invertase en invertase-inhibitor in hierdie plante te bepaal. Transkripsievlakke van die sukrosetransporter NtSut1 en die heksosetransporter, NtMst1, het beduidend toegeneem in die bron-blare en wortels van die Vac-Inv lyne; 'n toename in NtMst1 transkripsievlakke is ook in die wortels van Apo-Inv lyne bevestig. Die toenames in boodskapper RNA kon gekorreleer word met die veranderde invertase-aktiwiteite en suikervlakke in hierdie weefsels. Die gevolgtrekking word gemaak dat NtSUT1 en NtMST1 differensieël gereguleer word op transkripsionele vlak deur die sukrose en/of heksose inhoud van weefsels. Meer nog, die regulerende effek van die veranderde suikervlakke op transporteruitdrukking het afgehang van die subsellulêre kompartement waarin die gis-invertase uitgedruk is. Dit wil dus voorkom dat die subsellulêre kompartementalisering van suikermetabolisme fundamenteel tot die deurgee en waarneming van suikerseine is, met In gevolglike eweneens belangrike rol in die regulering van suikerverdeling. Die transkripsievlakke van beide die endogene selwand-invertase (CWI) en die selwand-invertase-inhibitor (CWI-Inh) enkoderende gene is in verskeie weefsels van die Apo-Inv en Vac-Inv lyne, tydens beide die vegetatiewe- en blomstadia, bestudeer. Die onderskeie weefsels van die Apo-Inv en Vac-Inv lyne het, in vergelyking met die kontrole lyne, veranderde Cwi en Cwi-inh transkripsievlakke getoon wat bepaal is deur bron-swelgpunt status en groeistadium. Geen duidelike korrelasie kon tussen beide Cwi en Cwi-inh uitdrukkingsvlakke en oplosbare suiker inhoud gevind word nie. Daar word voorgestel dat post-transkripsionele en posttranslasionele beheer deur suikers 'n belangrike rol in die regulering van hierdie proteïne speel. Bestudering van die Cwi:Cwi-lnh mRNA verhouding, asook groei verskynsels van die onderskeie weefsels van kontrole en Apo-Inv en Vac-Inv lyne, dui daarop dat hierdie transkripsievlak-verhouding moontlik 'n akkurate aanwyser van die swelgpuntkrag van 'n swelgpuntorgaan kan wees. Voorts is die invloed van bron-swelgpuntorgaan interaksies op suikerverdeling ondersoek. Omgekeerde enting tussen Apo-Inv en kontrole lyne het entlote met gemodifiseerde suikermetabolisme in óf hul bron- óf hul swelgpuntorgane tot gevolg gehad. Hierdie entlote is aan biomassaverspreidings-, oplosbare suiker kwantifisering en C4C]-radiomerking eksperimente onderwerp. Hierdie resultate het gewys dat, in vergelyking met die kontrole (GUS/GUS) ente, daar geen verandering in die status van suikerverdeling vanaf die bogrondse plantdele in die Apo/GUS ente is nie, maar wel 'n beduidende verskuiwing in suikerverdeling na die wortels van die GUS/Apo ente. Fenotipiese veranderinge, wat teenoorgesteld van dié teenwoordig in die Apo- Inv lyne waar die heteroloë invertase in beide bron en swelgpuntorgane uitgedruk word, is aanvanklik in die GUS/Apo en Apo/GUS ente waargeneem. Geen verskille in fenotipe of biomassa-verspreiding kon egter sewe weke na die entings prosedures tussen die GUS/Apo, Apo/GUS and GUS/GUS ente gevind word nie. Dit mag verduidelik word deur 'n moontlike toename in respirasietempo in die betrokke weefsels; die oplosbare suikervlakke wat in die verskillende ente aangeteken is ondersteun dié moontlikheid. Hierdie resultate as geheelonderstreep die kompleksiteit en aanpasbaarheid van suikermetabolisme en -verdeling. Verder bevestig dit dat suikerverdeling beïnvloed kan word deur bron-swelgpunt interaksies, asook die belang van invertases in die regulering van suikerverdeling gegewe die vermoë om swelgpuntkrag te verander. Hierdie studie vorm deel van 'n vinnig groeiende inisiatief om die beheermeganismes van suikerverdeling te ontrafel. Die resultate verkry in hierdie studie bekragtig die belang van rekombinante DNA tegnologie in die bestudering van fundamentele plantprosesse. Die invoeging en uitdrukking van 'n geteikende gisinvertase in transgeniese plante het gelei tot veranderde suikervlakke en bronswelgpunt interaksies in hierdie lyne met die gevolglike ontginning van waardevolle inligting ten opsigte van die regulering van suikerverdeling in reaksie tot interne seine. Daar is aangetoon dat suikertransporters onlosmaakbaar gekoppel is aan die deurgee en waarneming van suikerseine, spesifiek op die vlak van transkripsionele regulering, en dus ook die regulering van suikerverdeling. Voorts wys die resultate op die komplekse en aanpasbare aard van die verhouding wat bestaan tussen suikermetabolisme, -verdeling en groeiverskynsels.

Thesis (PhD (Genetics. Plant Biotechnology))--University of Stellenbosch, 2005. / The current understanding of the regulation of sucrose accumulation is still incomplete even though many scientists have investigated this subject. Components of trehalose metabolism have been implicated in the regulation of carbon flux in bacteria, yeast and more recently in plants. With a view to placing trehalose metabolism in the context of cytosolic sugarcane sucrose metabolism and carbon partitioning we have investigated the metabolites, transcripts and enzymes involved in this branch of carbohydrate metabolism in sugarcane internodal tissues. Sugarcane internodal trehalose levels varied between 0.31 ± 0.09 and 3.91 ± 0.99 nmol.g-1 fresh weight (FW). From statistical analysis of the metabolite profile it would appear that trehalose does not directly affect sucrose accumulation, although this does not preclude involvement of trehalose- 6-phosphate in the regulation of carbon partitioning. The metabolite data generated in this study demanded further investigation into the enzymes (and their transcripts) responsible for trehalose metabolism. Trehalose is synthesised in a two step process by the enzymes trehalose-6-phosphate synthase (EC 2.4.1.15, TPS) and trehalose-6-phosphate phosphatase (EC 3.1.3.12, TPP), and degraded by trehalase (EC 3.2.1.28). Two novel sugarcane partial cDNAs that coded for trehalase (tre) and actin (required for normalisation in profiling experiments) were isolated and used along with partial transcripts for TPS and TPP to determine transcript levels in different tissue- and genotypes. A putative full-length SugTPS cDNA was isolated and characterised. Enzyme activities for TPS, TPP and trehalase were measured at levels of 2.7 nmol.min-1.mg-1protein, 8.5 nmol.min-1.mg-1protein and 6.2 nmol.min-1.mg-1protein respectively, from young internodal protein extracts of sugarcane, variety N19. TPP enzyme activity and transcript levels were higher in S. spontaneum than Saccharum interspecific hybrids. Kinetic analysis of TPP and trehalase activities were performed with the purpose of providing parameters for an in silico kinetic model of trehalose and sucrose metabolism. Three isoforms of TPP were identified and desingated TPPAI, TPPAII and TPPB. Both TPPA isoforms had pH optima of 6.0, and TPPB of pH 6.5. Apparent Km values were determined as 0.447 ± 0.007 mM for TPPAI, 13.82 ± 1.98 mM for TPPAII and 1.387 ± 0.18 mM for TPPB. Partial purification and characterisation of trehalase demonstrated dual pH optima of 3.5 and 6.0, with Km values between 0.345 and 0.375 mM. These data were used as the basis for a kinetic model of trehalose metabolism. A previously described kinetic model of cytosolic sucrose metabolism has been expanded to include the trehalose pathway (TPS, TPP and trehalase). The aim was to supplement the available information on cytosolic metabolism in sugarcane storage parenchyma, identify points of control between sucrose and trehalose metabolism, and provide a platform from which further experimental and in silico modelling can be launched. The model predicted trehalose in the same order of magnitude as those determined in the metabolite profiling experiments. The majority of control of flux over the trehalose pathway resided in the TPS step, with flux control coefficients > 70% of the total pathway. Incorporation of the trehalose branch into the original sucrose model showed that reactions from the original model significantly affected the steady-state attributes of the trehalose pathway. Due to the relatively low flux through the trehalose branch of the expanded model, complete recycling of trehalose, and the lack of allosteric regulation by trehalose-6-phosphate or trehalose on any of the reactions from the original sucrose model, incorporation of the trehalose branch had no significant effect on either steady-state cytosolic sucrose concentration or flux of sucrose into the vacuole. The expanded model affords a basis from which to further investigate trehalose metabolism in the context of plant sucrose accumulation.

Thesis (PhD (Biochemistry))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: The sugarcane stalk, besides being the main structural component of the plant, is also the major storage organ for carbohydrates. Sucrose forms the bulk of stored carbohydrates. Previous studies have modelled the sucrose accumulation pathway in the internodal storage parenchyma of sugarcane using kinetic models cast as systems of ordinary differential equations. Typically, results were analysed with methods such as metabolic control analysis. The present study extends those original models within an advection-diffusion-reaction framework, requiring the use of partial differential equations to model sucrose metabolism coupled to phloem translocation. Let N be a stoichiometric matrix, v a vector of reaction rates, s a vector of species concentrations and r the gradient operator. Consider a coupled network of chemical reactions where the species may be advected with velocities, U, or diffuse with coefficients, D, or both. We propose the use of the dynamic system, s + r (Us) + r (Drs) = Nv; for a kinetic model where species can exist in different compartments and can be transported over long distances in a fluid medium, or involved in chemical reactions, or both. Darcy’s law is used to model fluid flow and allows a simplified, phenomenological approach to be applied to translocation in the phloem. Similarly, generic reversible Hill equations are used to model biochemical reaction rates. These are also phenomenological equations, where all the parameters have operationally defined interpretations. Numerical solutions to this formulation are demonstrated with time-courses of two toy models. The first model uses a simple “linear” pathway definition to study the impact of the system geometry on the solutions. Although this is an elementary model, it is able to demonstrate the up-regulation of photosynthesis in response to a change in sink demand. The second model elaborates on the reaction pathway while keeping the same geometry definition as the first. This pathway is designed to be an abstracted model of sucrose metabolism. Finally, a realistic model of sucrose translocation, metabolism and accumulation is presented, spanning eight internodes and four compartments. Most of the parameters and species concentrations used as initial values were obtained from experimental measurements. To analyse the models, a method of sensitivity analysis called the Fourier Amplitude Sensitivity Test (FAST) is employed. FAST calculates the contribution of the possible variation in a parameter to the total variation in the output from the model, i.e. the species concentrations and reaction rates. The model predicted that the most important factors affecting sucrose accumulation are the synthesis and breakdown of sucrose in futile cycles and the rate of cross-membrane transport of sucrose. The models also showed that sucrose moves down a concentration gradient from the leaves to the symplast, where it is transported against a concentration gradient into the vacuole. There was a net gain in carbohydrate accumulation in the realistic model, despite an increase in futile cycling with internode maturity. The model presented provides a very comprehensive description of sucrose accumulation and is a rigorous, quantitative framework for future modelling and experimental design. / AFRIKAANSE OPSOMMING: Benewens sy strukturele belang, is die suikerrietstingel ook die primêre bergingsorgaan vir koolhidrate. Die oorgrote meerderheid van hierdie koolhidrate word as sukrose opgeberg. Studies tot dusver het die metabolisme rondom sukroseberging in die parenchiem van die onderskeie stingellitte as stelsels gewone differensiaalvergelykings gemodelleer. Die resultate is ondermeer met metaboliese kontrole-analise geanaliseer. Hierdie studie brei uit op die oorspronklike modelle, deur gebruik te maak van ’n stromings-diffusie-reaksie-raamwerk. Parsiële differensiaalvergelykings is geformuleer om die metabolisme van sukrose te koppel aan die vloei in die floëem. Gestel N is ’n stoichiometriese matriks, v ’n vektor van reaksiesnelhede, s ’n vektor van spesie-konsentrasies en r die differensiaalvektoroperator. Beskou ’n netwerk van gekoppelde reaksies waar die onderskeie spesies stroom met snelhede U, of diffundeer met koëffisiënte D, of onderhewig is aan beide prosesse. Dit word voorgestel dat die dinamiese stelsel, _s + r (Us) + r (Drs) = Nv; gebruik kan word vir ’n kinetiese model waar spesies in verskeie kompartemente kan voorkom en vervoer kan word oor lang afstande saam met ’n vloeier, of kan deelneem aan chemiese reaksies, of albei. Darcy se wet word gebruik om die vloeier te modeller en maak dit moontlik om ’n eenvoudige, fenomenologiese benadering toe te pas op floëem-vervoer. Eweneens word generiese, omkeerbare Hill-vergelykings gebruik om biochemiese reaksiesnelhede te modelleer. Hierdie vergelykings is ook fenomenologies van aard en beskik oor parameters met ’n duidelike fisiese betekenis. Hierdie omvattende raamwerk is ondermeer gedemonstreer met behulp van numeriese oplossings van twee vereenvoudigde modelle as voorbeelde. Die eerste model het bestaan uit ’n lineêre reaksienetwerk en is gebruik om die geometrie van die stelsel te bestudeer. Alhoewel hierdie ’n eenvoudige model is, kon dit die toename in fotosintese as gevolg van ’n verandering in metaboliese aanvraag verklaar. Die tweede model het uitgebrei op die reaksieskema van die eerste, terwyl dieselfde stelselgeometrie behou is. Hierdie skema is ontwerp as ’n abstrakte weergawe van sukrosemetabolisme. Ten slotte is ’n realistiese model van sukrosevervoer, metabolisme en berging ontwikkel wat agt stingellitte en vier kompartemente omvat. Die meeste parameters en konsentrasies van biochemiese spesies wat as aanvanklike waardes in die model gebruik is, is direk vanaf eksperimentele metings verkry. Die Fourier Amplitude Sensitiwiteits-Toets (FAST) is gebruik om die modelle te analiseer. FAST maak dit moontlik om die bydrae van parameters tot variasie in modeluitsette soos reaksiesnelhede en die konsentrasies van chemiese spesies te bepaal. Die model het voorspel dat sintese en afbraak van sukrose in ’n futiele siklus, asook transmembraan sukrosevervoer, die belangrikste faktore is wat sukrose-berging beïnvloed. Die model het ook getoon dat sukrose saam met ’n konsentrasiegradiënt beweeg vanaf die blare tot by die stingelparenchiem-sitoplasma, van waar dit teen ’n konsentrasiegradiënt na die vogselholte (vakuool) vervoer word. Volgens die realistiese model was daar ’n netto toename in die totale hoeveelheid koolhidrate, ten spyte van ’n toename in die futile siklus van sukrose in die ouer stingellitte. Die model wat in hierdie proefskrif voorgestel word verskaf ’n uitgebreide, omvattende beskrywing van sukroseberging. Voorts stel dit ’n rigiede kwantitatiewe raamwerk daar vir toekomstige modellering en eksperimentele ontwerp.

The study was aimed at determining the metabolic fate of sucrose in intact sugarcane internodal tissue. Three aspects of the fate of sucrose in storage tissue of whole plants formed the main focus of the work. These were the rate of sucrose accumulation in the developing culm, the characterisation of partitioning of carbon into different cellular organic fractions in the developing culm and the occurrence of sucrose turnover in both immature and mature stem tissues. Specific attention was paid to confirming the occurrence of sucrose turnover in both immature and mature internodal tissue. This sucrose turnover has been described previously in both tissue slices and cell suspension cultures. However, certain results from previous work at the whole plant level have indicated that sucrose turnover does not occur in mature internodal tissue. Radiolabeled carbon dioxide (14CO2) was fed to leaf 6 of sugarcane culms of a high sucrose storing variety (Saccharum spp. hybrid cv. Nco376). All plants were of similar age (12 months) and were grown under similar conditions. The movement and metabolic fate of radiolabeled sucrose was determined at four time points, (6 hours, 24 hours, 7 days and 6 weeks) during a 6 week period. The metabolic fate of sucrose was determined in internodes number 3, number 6 and number 9. Internode 3 was found to have a relatively high hexose sugar content of 42 mg glc&fruc fw g-1 and a low sucrose content of 14 mg suc fw g-1. In contrast the sucrose content of internode 9 was much higher at 157 mg suc fw g-1 and the hexose sugar content much lower at 4.3 mg glc&fruc fw g-1. Based on previous work, the sugar content of internode 3 and internode 9 are characteristic of immature and mature tissues respectively. Internode 6 occupies an intermediary position between internode 3 and 6 with its sucrose content higher than its hexose sugar content, but with the hexose sugar content still being notable at 15 mg glc&fruc fw g-1. Although the metabolic fate of sucrose within sink tissue was the focal point of the study, the experimental design also allowed for certain aspects of sucrose production in the source to be investigated. The average photosynthetic rate for leaf 6 in full sunlight was estimated at 48 mg CO2 dm-2 s -1. During photosynthesis, only 30% of the fixed carbon was partitioned into the storage carbohydrate pool while the remaining 70% was partitioned into sucrose for immediate export from the leaf. This high rate of carbon fixation combined with a high rate of carbon export is characteristic of C4 plants such as sugarcane. On entering the culm, translocation of radiolabeled sucrose was predominantly basipetal with relatively little acropetal translocation. The majority of the radiolabeled carbon was found to be stored in mature internodes. No significant loss of radiolabeled carbon was observed in mature and elongating internodes over the study period. A 22% loss of total radiolabeled carbon was observed in immature internodes over the same period. This can probably be attributed to the higher rates of cellular respiration known to occur in immature tissues. There appear to be three phases of sucrose accumulation in the developing culm. Initially, the accumulation rate in rapidly growing tissue, as internode 3 develops into internode 6, is relatively low. This is followed by a rapid increase in the rate of sucrose accumulation during internode elongation, as internode 6 becomes internode 9. Finally, a decrease in the rate of sucrose accumulation is observed during late maturation, as internode 9 becomes internode 12. Determination of the sucrose content in internodes 3, 6 and 9 revealed that there is a notable increase in sucrose content during internode maturation. It is proposed that the higher sucrose content of mature tissue is not merely a consequence of the longer growth period of mature tissue, but is due to the increased rate of sucrose accumulation observed during internode elongation. Short-term (24 hours) analysis of carbon partitioning revealed that intemodal maturation was associated with a redirection of carbon from non-sucrose cellulal organic fractions to sucrose storage. In immature internodes only 20% of the total radiolabeled carbon was present in the sucrose pool 24 hours after feeding. In elongating internodes the figure increased to 54% while in mature internodes as much as 77% of the total radiolabeled carbon was retained in the sucrose pool. Concomitant with the increased carbon partitioning into stored sucrose down the developing culm is a decrease in carbon partitioning into the hexose sugar pool. In immature tissue, 42 % of the total radiolabel is present in the hexose sugar pool, while in mature tissue the percentage drops to 11%. This decrease is probably indicative of decreased levels of carbon cycling between the sucrose and hexose sugar pool as a result of internode maturation. Internode maturation was also found to be associated with a decrease in the amount of carbon in the water insoluble matter pool and the amino acid/ organic acid/ sugar phosphate pool. Thus, internode maturation is associated with a redirection of carbon from total respiration to sucrose storage. Long-term (6 weeks) analysis of carbon partitioning confirmed that sucrose storage in mature tissue is greater than that in immature tissue. From the 6 hour time point to the 6 week time point, an 87% reduction in the stored radiolabeled sucrose content was observed in immature internodes. During the same period only a 25% reduction in the stored radiolabeled sucrose was observed in mature internodes. Radiolabel loss from the radiolabeled sucrose pool in both mature and immature internodes was accounted for by relative radiolabel gains in other cellular organic fractions. At all time points during the study, and in all three tissues studied, radiolabel was found in the sucrose pool, the hexose sugars pool, the ionic pool and the water insoluble matter pool. The occurrence of radiolabel in the non-sucrose tissue constituents suggests that sucrose turnover is occurring in both immature, and mature internodal tissue. / Thesis (M.Sc.)-University of Natal, Durban, 2000.