GM crops: science, politics and communication

As the public debate in Europe about genetically modified (GM) crops heats up and the trade row between the United States and the European Union over GM food escalates, what better time to examine the issues with an international group of experts (Box 1). Their views are diverse, but they all agree that we need more impartial communication, less propaganda and an effective regulatory regime that is based on a careful case-by-case consideration of GM technology. It seems that GM crops are here to stay, so let us hope that these requirements are met and that the developing nations that perhaps have the most to gain from this technology can start to reap its benefits.     

The transmembrane domains of the sensor kinase KdpD of Escherichia coli are not essential for sensing K+ limitation

The sensor kinase/response regulator system KdpD/KdpE of Escherichia coli regulates the expression of the kdpFABC operon, which encodes the high affinity K+ transport system KdpFABC. The membrane-bound sensor kinase KdpD consists of four transmembrane domains, a large cytoplasmic N-terminal domain and a cytoplasmic C-terminal transmitter domain. To elucidate the role of the four transmembrane domains, various deletions were introduced in kdpD and the activities of the resulting truncated derivatives of KdpD were determined. A KdpD protein lacking all four transmembrane domains was able to sense low K+ concentrations, whereas at higher K+ concentrations kdpFABC expression was constitutive. These and further results with various truncated KdpD proteins lacking distinct parts of the transmembrane domains or derivatives in which a linker peptide or two transmembrane domains of PutP, the Na+/proline transporter of Escherichia coli, replaced the missing part indicated that the transmembrane domains are not essential for sensing of K+ limitation, but may be important for the correct positioning of the large N- and C-terminal cytoplasmic domains to each other.     

Loss of actin cytoskeletal function and EDS1 activity,in combination,severely compromises non-host resistance in Arabidopsis against wheat powdery mildew

Plant immunity against the majority of the microbial pathogens is conveyed by a phenomenon known as non-host resistance (NHR). This defence mechanism affords durable protection to plant species against given species of phytopathogens. We investigated the genetic basis of NHR in Arabidopsis against the wheat powdery mildew fungus Blumeria graminis f. sp. tritici (Bgt). Both primary and appressorial germ tubes were produced from individual Bgt conidia on the surface of the Arabidopsis leaves. Attempted infection occasionally resulted in successful penetration, which led to the development of an abnormal unilateral haustorium. Inoculation of a series of Arabidopsis defence-related mutants with Bgt resulted in the attenuation of reactive oxygen intermediate (ROI) production and salicylic acid (SA)-dependent defence gene expression in eds1, pad4 and nahG plants, which are known to be defective in some aspects of host resistance. Furthermore, Bgt often developed bilateral haustoria in the mutant Arabidopsis lines that closely resembled those formed in wheat. A similar decrease in NHR was observed following treatment of the wild-type Arabidopsis plants with cytochalasin E, an inhibitor of actin microfilament polymerisation. In eds1 mutants, inhibition of actin polymerisation severely compromised NHR in Arabidopsis against Bgt. This permitted completion of the Bgt infection cycle on these plants. Therefore, actin cytoskeletal function and EDS1 activity, in combination, are major contributors to NHR in Arabidopsis against wheat powdery mildew.     

Detection and quantitation of cellularly derived amyloid β peptides by immunoprecipitation-HPLC-MS

A quantitative method for detection of amyloid β peptides using immunoprecipitation-HPLC-mass spectrometry (IP-LC-MS) is described. Comparison of IP-LC-MS with sandwich ELISA revealed comparable results in the analysis of Aβ 1–40 and Aβ 1–42 derived from fetal guinea pig cell media and cell lysates. The use of IP-LC-MS not only allows a quantitative method for Aβ 1–40 and Aβ 1–42 peptides present in Alzheimer's disease (AD), but allows detection of other Aβ peptide species that may also play a role in the onset of AD in humans.     

Post-Translational Modifications of β Subunits of Voltage-Dependent Calcium Channels

Different post-translational modifications of Ca channel subunits have been identified. Recent studies have characterized the palmitoylation of the Ca channel _2a subunit, as well as one effect of this modification on channel function. The potential importance of palmitoylation on other channel properties is discussed. Other studies have addressed the role of phosphorylation of subunits in the regulation of voltage-dependent Ca channels. Phosphorylation of subunits by second messenger-activated protein kinases, as well as by unidentified protein kinases, may affect interactions between channel subunits and other aspects of channel function. The differential modification of Ca channel subunit isoforms by post-translational events likely results in diversely regulated channels with unique properties.     

Directed evolution of a secretory leader for the improved expression of heterologous proteins and full‐length antibodies in Saccharomyces cerevisiae

Because of its eukaryotic nature, simple fermentation requirements, and pliable genetics, there have been many attempts at improving recombinant protein production in Saccharomyces cerevisiae. These strategies typically involve altering the expression of a native protein thought to be involved in heterologous protein trafficking. Usually, these approaches yield three‐ to tenfold improvements over wild‐type strains and are almost always specific to one type of protein. In this study, a library of mutant alpha mating factor 1 leader peptides (MFα1pp) is screened for the enhanced secretion of a single‐chain antibody. One of the isolated mutants is shown to enhance the secretion of the scFv up to 16‐fold over wild type. These leaders also confer a secretory improvement to two other scFvs as well as two additional, structurally unrelated proteins. Moreover, the improved leader sequences, combined with strain engineering, allow for a 180‐fold improvement over previous reports in the secretion of full‐length, functional, glycosylated human IgG₁. The production of full‐length IgG₁ at milligram per liter titers in a simple, laboratory‐scale system will significantly expedite drug discovery and reagent synthesis while reducing antibody cloning, production, and characterization costs. Biotechnol. Bioeng. 2009;103: 1192–1201. © 2009 Wiley Periodicals, Inc.