Neutral Bystander, Intrusive Micromanager, or Useful Catalyst?

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73380/1/j.1541-0072.1995.tb01746.x.pd

HIV-1 Vpu Protein Mediates the Transport of Potassium in Saccharomyces cerevisiae

Human immunodeficiency virus type 1 (HIV-1) Vpu is an integral membrane protein that belongs to the viroporin family. Viroporins interact with cell membranes, triggering membrane permeabilization and promoting release of viral particles. In vitro electrophysiological methods have revealed changes in membrane ion currents when Vpu is present; however, in vivo the molecular mechanism of Vpu at the plasma membrane is still uncertain. We used the yeast Saccharomyces cerevisiae as a genetic model system to analyze how Vpu ion channel impacts cellular homeostasis. Inducible expression of Vpu impaired cell growth, suggesting that this viral protein is toxic to yeast cultures. This toxicity decreased with extracellular acidic pH. Also, Vpu toxicity diminished as the extracellular K(+) concentration was increased. However, expression of the Vpu protein suppresses the growth defect of K(+) uptake-deficient yeast (Δtrk1,2). The phenotype rescue of these highly hyperpolarized cells was almost total when they were grown in medium supplemented with high concentrations of KCl (100 mM) at pH 7.0 but was significantly reduced when the extracellular K(+) concentration or pH was decreased. These results indicate that Vpu has the ability to modify K(+) transport in both yeast strains. Here, we show also that Vpu confers tolerance to the aminoglycoside antibiotic hygromycin B in Δtrk1,2 yeast. Our results suggest that Vpu interferes with cell growth of wild-type yeast but improves proliferation of the hyperpolarized trk1,2 mutant by inducing plasma membrane depolarization. Furthermore, evaluation of the ion channel activity of the Vpu protein in Δtrk1,2 yeast could aid in the development of a high-throughput screening assay for molecules that target the retroviral protein.This study was supported by Grants PI PI05/00013 and PI08/0912 from Fondo de Investigación Sanitaria. L.H. and N.M. were holders of Predoctoral Fellowships from Instituto de Salud Carlos III.S

Determination of mycotoxin concentration by ELISA and near-infrared spectroscopy in Fusarium-inoculated maize

Ear rots of maize caused by Fusarium spp. reduce grain yield and produce mycotoxins, which are harmful to humans and animals. To breed maize cultivars resistant to Fusarium spp., reliable large-scale phenotyping is essential. Our objectives were to (i) examine the precision of the ELISA method for determination of important mycotoxins, namely deoxynivalenol (DON) and fumonisins (FUM), (ii) evaluate the potential of near-infrared reflectance spectroscopy (NIRS) to estimate concentrations of DON and FUM in grain produced in inoculated maize plants, and (iii) compare the efficiency of ELISA, NIRS, and visual rating of disease severity for estimation of mycotoxin concentrations. Insignificant variation was observed between duplicate evaluations of DON and FUM by ELISA, showing the high repeatability of this method. DON and FUM determinations by ELISA were more closely correlated with mycotoxin concentrations predicted through NIRS than with visual rating of disease severity. For the prediction of DON, NIRS had very high magnitude of the coefficients of determination of calibration and cross validation (R 2 = 0.90–0.88). Thus, NIRS has a promising potential to predict DON concentration in grain samples of inoculated maize genotypes evaluated in resistance breeding programs