The tomato Cf-9 disease resistance gene functions in tobacco and potato to confer responsiveness to the fungal avirulence gene product avr 9

The Cf-9 gene encodes an extracytoplasmic leucine-rich repeat protein that confers resistance in tomato to races of the fungus Cladosporium fulvum that express the corresponding avirulence gene Avr 9. We investigated whether the genomic Cf-9 gene functions in potato and tobacco. Transgenic tobacco and potato plants carrying Cf-9 exhibit a rapid hypersensitive cell death response (HR) to Avr 9 peptide injection. Cf 9 tobacco plants were reciprocally crossed to Avr 9-producing tobacco. A developmentally regulated seedling lethal phenotype occurred in F1 progeny when Cf9 was used as the male parent and Avr 9 as the female parent. However, when Cf9 was inherited in the maternal tissue and a heterozygous Avr 9 plant was used as the pollen donor, a much earlier reaction was caused, leading to no germination of any F1 seed. Detailed analysis of the Avr 9-induced responses in Cf 9 tobacco leaves revealed that (1) most mesophyll cells died within 3 hr (compared with 12 to 16 hr in tomato); (2) the macroscopic HR was visible at an Avr 9 titer five times lower than that which caused visible symptoms in tomato; (3) the HR invariably extended into noninjected panels of the tobacco leaf; (4) no HR occurred in leaves of young tobacco plants; (5) in older plants, the HR was dramatically enhanced by sequential Avr 9 challenges; and (6) coexpression of a salicylate hydroxylase transgene (nahG) from Pseudomonas putida reduced the severity of the macroscopic leaf HR and also restored germination to Cf 9 x 35S:Avr 9 F1 seedlings. Simultaneous introduction of Cf-9 homologs (Hcr 9-9 genes A and B or D) along with the native Cf-9 gene did not alter the responses that were specifically induced by Avr 9. Various ways to use the Cf-9-Avr 9 gene combination to engineer broad-spectrum disease resistance in several solanaceous species are discussed.     

Flicker noise of ion-selective membranes and turbulent convection in the depleted layer

Flicker noise of electric currents through ion-selective membranes is explained. It is attributed to the depletion of salt on one side of the membrane, which creates a thin layer of high resistance. Joule heating in this depletion layer and the ensuing temperature gradient, as well as the concentration gradient, give rise to buoyant forces which may create a turbulent convection current. The turbulence mixes the depletion layer so that the electric resistance fluctuates, and consequently the current flickers.Experiments with ion-selective membranes support this conjecture. They show that 1) Noise is coincident with the increase of the electric resistance by the depletion process. 2) When the current density is reduced, it reaches a critical value, below which the convection current changes from turbulent to laminar, and the noise disappears. 3) Noise reduces with temperature, because the expansion coefficient of water decreases with temperature, and its viscosity increases. 4) A non-ionic water-soluble polymer added to the compartment on the side of the depletion layer reduces the noise, by increasing the bulk viscosity of the solution. 5) Noise depends on the membrane's orientation in the gravitational field. 6) The convection-current in the depletion layer can be observed directly, using a laser-beam, by adding latex particles which create optical noise as they drift with the convection current across the beam. The optical noise is observed only coincidently with the current noise.A. Katzir-Katchalsky Fellow     

Solvent effects on the helix–coil transition in polypeptides

A simple way to incorporate the solvent–peptide interaction in any available theory of the helix–coil transition is developed. The competition between the intramolecular hydrogen bonding and the solvent–polymer hydrogen bonding is considered in multi-component solvents where some of the components have hydrogen-bonding capacity. Molecular averages are computed by using the theory of Lifson and Roig. The experimental data of Yang are analyzed, and the range of acceptable values of the equilibrium constants of hydrogen bond formation is deduced. The enthalpy of the transition in multicomponent solvents is calculated.     

Protection Afforded by an HIV Vaccine Candidate in Macaques Depends on the Dose of SIVmac251 at Challenge Exposure

We used the simian immunodeficiency virus mac251 (SIV_mac251) macaque model to study the effect of the dose of mucosal exposure on vaccine efficacy. We immunized macaques with a DNA prime followed by SIV gp120 protein immunization with ALVAC-SIV and gp120 in alum, and we challenged them with SIV_mac251 at either a single high dose or at two repeated low-dose exposures to a 10-fold-lower dose. Infection was neither prevented nor modified following a single high-dose challenge of the immunized macaques. However, two exposures to a 10-fold-lower dose resulted in protection from SIV_mac251 acquisition in 3 out of 12 macaques. The remaining animals that were infected had a modulated pathogenesis, significant downregulation of interferon responsive genes, and upregulation of genes involved in B- and T-cell responses. Thus, the choice of the experimental model greatly influences the vaccine efficacy of vaccines for human immunodeficiency virus (HIV).     

CD8 T Cell Response Maturation Defined by Anentropic Specificity and Repertoire Depth Correlates with SIVΔnef-induced Protection

The live attenuated simian immunodeficiency virus (LASIV) vaccine SIVΔnef is one of the most effective vaccines in inducing protection against wild-type lentiviral challenge, yet little is known about the mechanisms underlying its remarkable protective efficacy. Here, we exploit deep sequencing technology and comprehensive CD8 T cell epitope mapping to deconstruct the CD8 T cell response, to identify the regions of immune pressure and viral escape, and to delineate the effect of epitope escape on the evolution of the CD8 T cell response in SIVΔnef-vaccinated animals. We demonstrate that the initial CD8 T cell response in the acute phase of SIVΔnef infection is mounted predominantly against more variable epitopes, followed by widespread sequence evolution and viral escape. Furthermore, we show that epitope escape expands the CD8 T cell repertoire that targets highly conserved epitopes, defined as anentropic specificity, and generates de novo responses to the escaped epitope variants during the vaccination period. These results correlate SIVΔnef-induced protection with expanded anentropic specificity and increased response depth. Importantly, these findings render SIVΔnef, long the gold standard in HIV/SIV vaccine research, as a proof-of-concept vaccine that highlights the significance of the twin principles of anentropic specificity and repertoire depth in successful vaccine design.