Paola Lucca

Bio: Paola Lucca is an academic researcher from École Polytechnique Fédérale de Lausanne. The author has contributed to research in topics: Endosperm & Golden rice. The author has an hindex of 7, co-authored 7 publications receiving 3164 citations.

Engineering the Provitamin A (β-Carotene) Biosynthetic Pathway into (Carotenoid-Free) Rice Endosperm

Abstract: Rice (Oryza sativa), a major staple food, is usually milled to remove the oil-rich aleurone layer that turns rancid upon storage, especially in tropical areas. The remaining edible part of rice grains, the endosperm, lacks several essential nutrients, such as provitamin A. Thus, predominant rice consumption promotes vitamin A deficiency, a serious public health problem in at least 26 countries, including highly populated areas of Asia, Africa, and Latin America. Recombinant DNA technology was used to improve its nutritional value in this respect. A combination of transgenes enabled biosynthesis of provitamin A in the endosperm.

Role of EDEM in the release of misfolded glycoproteins from the calnexin cycle.

Abstract: The mechanisms that determine how folding attempts are interrupted to target folding-incompetent proteins for endoplasmic reticulum-associated degradation (ERAD) are poorly defined. Here the alpha-mannosidase I-like protein EDEM was shown to extract misfolded glycoproteins, but not glycoproteins undergoing productive folding, from the calnexin cycle. EDEM overexpression resulted in faster release of folding-incompetent proteins from the calnexin cycle and earlier onset of degradation, whereas EDEM down-regulation prolonged folding attempts and delayed ERAD. Up-regulation of EDEM during ER stress may promote cell recovery by clearing the calnexin cycle and by accelerating ERAD of terminally misfolded polypeptides.

Golden Rice: Introducing the β-Carotene Biosynthesis Pathway into Rice Endosperm by Genetic Engineering to Defeat Vitamin A Deficiency

Abstract: To obtain a functioning provitamin A (beta-carotene) biosynthetic pathway in rice endosperm, we introduced in a single, combined transformation effort the cDNA coding for phytoene synthase (psy) and lycopene beta-cyclase (beta-lcy) both from Narcissus pseudonarcissus and both under the control of the endosperm-specific glutelin promoter together with a bacterial phytoene desaturase (crtI, from Erwinia uredovora under constitutive 35S promoter control). This combination covers the requirements for beta-carotene synthesis and, as hoped, yellow beta-carotene-bearing rice endosperm was obtained in the T(0)-generation. Additional experiments revealed that the presence of beta-lcy was not necessary, because psy and crtI alone were able to drive beta-carotene synthesis as well as the formation of further downstream xanthophylls. Plausible explanations for this finding are that these downstream enzymes are constitutively expressed in rice endosperm or are induced by the transformation, e.g., by enzymatically formed products. Results using N. pseudonarcissus as a model system led to the development of a hypothesis, our present working model, that trans-lycopene or a trans-lycopene derivative acts as an inductor in a kind of feedback mechanism stimulating endogenous carotenogenic genes. Various institutional arrangements for disseminating Golden Rice to research institutes in developing countries also are discussed.

Effective selection and regeneration of transgenic rice plants with mannose as selective agent

Abstract: A new method for the selection of transgenic rice plants without the use of antibiotics or herbicides has been developed. The phosphomannose isomerase (PMI) gene from Escherichia coli has been cloned and consitutively expressed in japonica rice variety TP 309. The PMI gene was transferred to immature rice embryos by Agrobacterium-mediated transformation, which allowed the selection of transgenic plants with mannose as selective agent. The integration and expression of the transgene was confirmed by Southern and northern blot analysis and the activity of PMI indirectly proved with the chlorophenol red assay. The results of genetic analysis showed that the transgenes were segregated in a Mendelian fashion in the T1 generation. The establishment of this selection system in rice provides an efficient way for producing transgenic plants without using antibiotics or herbicides with a transformation frequency of up to 41%.

Engineering the Provitamin A (b-Carotene) Biosynthetic Pathway into (Carotenoid-Free)

Abstract: ). Protoplasts were lysed and recombi-nant protein was immunoprecipitated with GFP pep-tide antibodies (Clontech, Palo Alto, CA) and proteinA—Sepharose CL-4B (Amersham Pharmacia Biotech,Picataway, NJ). Immunoprecipitated proteins wereassayed for kinase activity using histone III-S (Sigma)as substrate (

XBP-1 Regulates a Subset of Endoplasmic Reticulum Resident Chaperone Genes in the Unfolded Protein Response

Abstract: The mammalian unfolded protein response (UPR) protects the cell against the stress of misfolded proteins in the endoplasmic reticulum (ER). We have investigated here the contribution of the UPR transcription factors XBP-1, ATF6alpha, and ATF6beta to UPR target gene expression. Gene profiling of cell lines lacking these factors yielded several XBP-1-dependent UPR target genes, all of which appear to act in the ER. These included the DnaJ/Hsp40-like genes, p58(IPK), ERdj4, and HEDJ, as well as EDEM, protein disulfide isomerase-P5, and ribosome-associated membrane protein 4 (RAMP4), whereas expression of BiP was only modestly dependent on XBP-1. Surprisingly, given previous reports that enforced expression of ATF6alpha induced a subset of UPR target genes, cells deficient in ATF6alpha, ATF6beta, or both had minimal defects in upregulating UPR target genes by gene profiling analysis, suggesting the presence of compensatory mechanism(s) for ATF6 in the UPR. Since cells lacking both XBP-1 and ATF6alpha had significantly impaired induction of select UPR target genes and ERSE reporter activation, XBP-1 and ATF6alpha may serve partially redundant functions. No UPR target genes that required ATF6beta were identified, nor, in contrast to XBP-1 and ATF6alpha, did the activity of the UPRE or ERSE promoters require ATF6beta, suggesting a minor role for it during the UPR. Collectively, these results suggest that the IRE1/XBP-1 pathway is required for efficient protein folding, maturation, and degradation in the ER and imply the existence of subsets of UPR target genes as defined by their dependence on XBP-1. Further, our observations suggest the existence of additional, as-yet-unknown, key regulators of the UPR.

Roles of N-Linked Glycans in the Endoplasmic Reticulum

Abstract: From a process involved in cell wall synthesis in archaea and some bacteria, N-linked glycosylation has evolved into the most common covalent protein modification in eukaryotic cells. The sugars are added to nascent proteins as a core oligosaccharide unit, which is then extensively modified by removal and addition of sugar residues in the endoplasmic reticulum (ER) and the Golgi complex. It has become evident that the modifications that take place in the ER reflect a spectrum of functions related to glycoprotein folding, quality control, sorting, degradation, and secretion. The glycans not only promote folding directly by stabilizing polypeptide structures but also indirectly by serving as recognition "tags" that allow glycoproteins to interact with a variety of lectins, glycosidases, and glycosyltranferases. Some of these (such as glucosidases I and II, calnexin, and calreticulin) have a central role in folding and retention, while others (such as alpha-mannosidases and EDEM) target unsalvageable glycoproteins for ER-associated degradation. Each residue in the core oligosaccharide and each step in the modification program have significance for the fate of newly synthesized glycoproteins.

Enrichment of cereal grains with zinc: Agronomic or genetic biofortification?

Abstract: Zinc deficiency is a well-documented problem in food crops, causing decreased crop yields and nutritional quality. Generally, the regions in the world with Zn-deficient soils are also characterized by widespread Zn deficiency in humans. Recent estimates indicate that nearly half of world population suffers from Zn deficiency. Cereal crops play an important role in satisfying daily calorie intake in developing world, but they are inherently very low in Zn concentrations in grain, particularly when grown on Zn-deficient soils. The reliance on cereal-based diets may induce Zn deficiency-related health problems in humans, such as impairments in physical development, immune system and brain function. Among the strategies being discussed as major solution to Zn deficiency, plant breeding strategy (e.g., genetic biofortification) appears to be a most sustainable and cost-effective approach useful in improving Zn concentrations in grain. The breeding approach is, however, a long-term process requiring a substantial effort and resources. A successful breeding program for biofortifying food crops with Zn is very much dependent on the size of plant-available Zn pools in soil. In most parts of the cereal-growing areas, soils have, however, a variety of chemical and physical problems that significantly reduce availability of Zn to plant roots. Hence, the genetic capacity of the newly developed (biofortified) cultivars to absorb sufficient amount of Zn from soil and accumulate it in the grain may not be expressed to the full extent. It is, therefore, essential to have a short-term approach to improve Zn concentration in cereal grains. Application of Zn fertilizers or Zn-enriched NPK fertilizers (e.g., agronomic biofortification) offers a rapid solution to the problem, and represents useful complementary approach to on-going breeding programs. There is increasing evidence showing that foliar or combined soil+foliar application of Zn fertilizers under field conditions are highly effective and very practical way to maximize uptake and accumulation of Zn in whole wheat grain, raising concentration up to 60 mg Zn kg−1. Zinc-enriched grains are also of great importance for crop productivity resulting in better seedling vigor, denser stands and higher stress tolerance on potentially Zn-deficient soils. Agronomic biofortification strategy appears to be essential in keeping sufficient amount of available Zn in soil solution and maintaining adequate Zn transport to the seeds during reproductive growth stage. Finally, agronomic biofortification is required for optimizing and ensuring the success of genetic biofortification of cereal grains with Zn. In case of greater bioavailability of the grain Zn derived from foliar applications than from soil, agronomic biofortification would be a very attractive and useful strategy in solving Zn deficiency-related health problems globally and effectively.

ER stress and the unfolded protein response.

Abstract: Conformational diseases are caused by mutations altering the folding pathway or final conformation of a protein. Many conformational diseases are caused by mutations in secretory proteins and reach from metabolic diseases, e.g. diabetes, to developmental and neurological diseases, e.g. Alzheimer's disease. Expression of mutant proteins disrupts protein folding in the endoplasmic reticulum (ER), causes ER stress, and activates a signaling network called the unfolded protein response (UPR). The UPR increases the biosynthetic capacity of the secretory pathway through upregulation of ER chaperone and foldase expression. In addition, the UPR decreases the biosynthetic burden of the secretory pathway by downregulating expression of genes encoding secreted proteins. Here we review our current understanding of how an unfolded protein signal is generated, sensed, transmitted across the ER membrane, and how downstream events in this stress response are regulated. We propose a model in which the activity of UPR signaling pathways reflects the biosynthetic activity of the ER. We summarize data that shows that this information is integrated into control of cellular events, which were previously not considered to be under control of ER signaling pathways, e.g. execution of differentiation and starvation programs.

Biofortification of crops with seven mineral elements often lacking in human diets--iron, zinc, copper, calcium, magnesium, selenium and iodine.

Abstract: Summary The diets of over two-thirds of the world's population lack one or more essential mineral elements. This can be remedied through dietary diversification, mineral supplementation, food fortification, or increasing the concentrations and/or bioavailability of mineral elements in produce (biofortification). This article reviews aspects of soil science, plant physiology and genetics underpinning crop biofortification strategies, as well as agronomic and genetic approaches currently taken to biofortify food crops with the mineral elements most commonly lacking in human diets: iron (Fe), zinc (Zn), copper (Cu), calcium (Ca), magnesium (Mg), iodine (I) and selenium (Se). Two complementary approaches have been successfully adopted to increase the concentrations of bioavailable mineral elements in food crops. First, agronomic approaches optimizing the application of mineral fertilizers and/or improving the solubilization and mobilization of mineral elements in the soil have been implemented. Secondly, crops have been developed with: increased abilities to acquire mineral elements and accumulate them in edible tissues; increased concentrations of ‘promoter’ substances, such as ascorbate, β-carotene and cysteine-rich polypeptides which stimulate the absorption of essential mineral elements by the gut; and reduced concentrations of ‘antinutrients’, such as oxalate, polyphenolics or phytate, which interfere with their absorption. These approaches are addressing mineral malnutrition in humans globally.