Quintín Rascón-Cruz

Lactoferrin: structure, function and applications.

Lactoferrin (LF) is an 80 kDa iron-binding glycoprotein of the transferrin family that is expressed in most biological fluids and is a major component of the mammalian innate immune system. Its protective effects range from direct antimicrobial activities against a large panel of microorganisms, including bacteria, viruses, fungi and parasites, to anti-inflammatory and anticancer activities. These extensive activities are made possible by mechanisms of action utilising not only the capacity of LF to bind iron but also interactions of LF with molecular and cellular components of both host and pathogens. This review summarises the putative antimicrobial mechanisms, clinical applications and heterologous expression models for LF.

Lactoferrin a multiple bioactive protein: an overview.

Abstract Background Lactoferrin (Lf) is an 80kDa iron-binding glycoprotein of the transferrin family. It is abundant in milk and in most biological fluids and is a cell-secreted molecule that bridges innate and adaptive immune function in mammals. Its protective effects range from anticancer, anti-inflammatory and immune modulator activities to antimicrobial activities against a large number of microorganisms. This wide range of activities is made possible by mechanisms of action involving not only the capacity of Lf to bind iron but also interactions of Lf with molecular and cellular components of both hosts and pathogens. Scope of review This review summarizes the activities of Lf, its regulation and potential applications. Major conclusions The extensive uses of Lf in the treatment of various infectious diseases in animals and humans has been the driving force in Lf research however, a lot of work is required to obtain a better understanding of its activity. General significance The large potential applications of Lf have led scientists to develop this nutraceutical protein for use in feed, food and pharmaceutical applications. This article is part of a Special Issue entitled Molecular Mechanisms of Iron Transport and Disorders.

Accumulation, assembly, and digestibility of amarantin expressed in transgenic tropical maize.

An amaranth (Amaranthus hypochondriacus) 11S globulin cDNA, encoding one of the most important storage proteins (amarantin) of the seed, with a high content of essential amino acids, was used in the transformation of CIMMYT tropical maize genotype. Constructs contained the amarantin cDNA under the control of a tissue-specific promoter from rice glutelin-1 (osGT1) or a constitutive (CaMV 35S) promoter with and without the first maize alcohol dehydrogenase intron (AdH). Southern-blot analysis confirmed the integration of the amarantin cDNA, and copy number ranged from one to more than ten copies per maize genome. Western-blot and ultracentrifugation analyses of transgenic maize indicate that the expressed recombinant amarantin precursors were processed into the mature form, and accumulated stably in maize endosperm. Total protein and some essential amino acids of the best expressing maize augmented 32% and 8–44%, respectively, compared to non-transformed samples. The soluble expressed proteins were susceptible to digestion by simulated gastric and intestinal fluids, and it is suggested that they show no allergenic activity. These findings demonstrate the feasibility of using genetic engineering to improve the amino acid composition of grain crops.

Immunomodulatory effects of lactoferrin

Lactoferrin (Lf) is an iron-binding glycoprotein of the transferrin family, which is expressed in most biological fluids with particularly high levels in mammalian milk. Its multiple activities lie in its capacity to bind iron and to interact with the molecular and cellular components of hosts and pathogens. Lf can bind and sequester lipopolysaccharides, thus preventing pro-inflammatory pathway activation, sepsis and tissue damages. Lf is also considered a cell-secreted mediator that bridges the innate and adaptive immune responses. In the recent years much has been learned about the mechanisms by which Lf exerts its activities. This review summarizes the recent advances in understanding the mechanisms underlying the multifunctional roles of Lf, and provides a future perspective on its potential prophylactic and therapeutic applications.

High-Level Expression of Recombinant Bovine Lactoferrin in Pichia pastoris with Antimicrobial Activity

In this study, bovine lactoferrin (bLf), an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf) in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin) demonstrated antibacterial activity against Escherichia coli (E. coli) BL21DE3, Staphylococcus aureus (S. aureus) FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa) ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly.

Genetic transformation of blue grama grass with the rolA gene from Agrobacterium rhizogenes: regeneration of transgenic plants involves a "hairy embryo" stage

Until recently, information about the effects of transforming plants with the rolA gene of Agrobacterium rhizogenes has been restricted mainly to dicots in which a severely wrinkled phenotype, reduced internode distances, and abnormal reproductive development were commonly observed. In this work, we analyzed the effects associated with the expression of this gene in a new genetic context: the forage grass genome. Transgenic P35S•rolA plants of blue grama grass (Bouteloua gracilis) were obtained by a biolistic approach employing embryogenic chlorophyllic cells as the target material. Four independent transgenic lines with regeneration capacity were recovered, which showed stable integration of this transgene as demonstrated by polymerase chain reaction and Southern blot hybridization. Growth of the rolA-transformed lines under greenhouse conditions provided evidence for a new biotechnological application for the rolA gene in plants, namely, the improvement of biomass production in forage grasses. Additionally, we described here a new phenotypic marker (referred to here as the “hairy embryo” syndrome) that can be instrumental for the early identification of transformation events when transforming grasses with this gene.

Identification of miRNA from Bouteloua gracilis, a drought tolerant grass, by deep sequencing and their in silico analysis

BACKGROUND MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate signal transduction, development, metabolism, and stress responses in plants through post-transcriptional degradation and/or translational repression of target mRNAs. Several studies have addressed the role of miRNAs in model plant species, but miRNA expression and function in economically important forage crops, such as Bouteloua gracilis (Poaceae), a high-quality and drought-resistant grass distributed in semiarid regions of the United States and northern Mexico remain unknown. RESULTS We applied high-throughput sequencing technology and bioinformatics analysis and identified 31 conserved miRNA families and 53 novel putative miRNAs with different abundance of reads in chlorophyllic cell cultures derived from B. gracilis. Some conserved miRNA families were highly abundant and possessed predicted targets involved in metabolism, plant growth and development, and stress responses. We also predicted additional identified novel miRNAs with specific targets, including B. gracilis ESTs, which were detected under drought stress conditions. CONCLUSIONS Here we report 31 conserved miRNA families and 53 putative novel miRNAs in B. gracilis. Our results suggested the presence of regulatory miRNAs involved in modulating physiological and stress responses in this grass species.

Comparison of the structure and organization of the rrna operons of Bouteloua gracilis and Zea mays

Aguado-Santacruz, G. A., Betancourt-Guerra, D. A., Siquerios-Cendón, T., Arévalo-Gallegos, S., Rivera-Chavira, B. E., Nevarez-Moorillon, G. V., Moreno-Gómez, B. and Rascón-Cruz, Q. 2011. Comparison of the structure and organization of therrnaoperons of Bouteloua gracilisandZea mays. Can. J. Plant Sci. 91: 107-116. We studied the genomic structure of Bouteloua gracilis chloroplast DNA (cpDNA) and compared it with the sequenced ribosomal RNA spacer region from other cereals. This will allow us to understand chloroplast topology and the recombination ability of cpDNA. The development of potential tools for biotechnology applied to cereals can be focused through the study of cpDNA in family related grasses, such as B. gracilis. cpDNA was prepared from green B. gracilis and Zea mays plants using a modified NaCl method. A 2332 bp intergenic spacer (IGS) region (rrna16S-trnI-trnA-rrna23S) from B. gracilis was sequenced, which showed great similarity (at least 92%) to IGS region from Z. mays, Oryza sativa and Saccharum officinarum. A physical map constructed by Southern hybridization using petA, psbA, psbD, ndhA, rbcL, 16S and 23S rDNA digoxigenin-labelled probes showed low organizational resemblance to maize cpDNA. Moreover, when compared to a similar fragment of Z. mays, a 239 bp intron deletion was found in the trnI gene in the B. gracilis cpDNA. Restriction and hybridization analyses suggested that the B. gracilis cpDNA has a molecular weight of 130 Kb. We expect that the findings reported in this work can be a baseline for increasing our knowledge in chloroplast organization in grasses and for the development of molecular tools.