No abstract

Plant virus or viroid infection stimulates the phosphorylation of a plant-encoded protein of M, 68,000 to 70,000 (now termed pPKR) that is associated with double-stranded RNA-stimulated protein kinase activity. Using various biochemical and immunological comparisons, we have demonstrated that this plant protein is an analog of the mammalian PKR enzymes. pPKR is both cytosolic and ribosome associated, similar to mammalian PKR, and appears to be capable of phosphorylating exogenous histones. Monoclonal antiserum to the human PKR as well as antiserum to a conserved double-stranded RNA-binding domain present on mammalian PKR demonstrated cross-reactivity with pPKR. Likewise, polyclonal antiserum to the pPKR detected the mouse and human PKR in western blot analysis. Northern blot analysis of a mammalian PKR cDNA detected a specific 2.5-kb transcript present in plant poly(A)' RNA.

Regulation of protein synthesis by eukaryotic initiation factor-2alpha (eIF-2alpha) phosphorylation is a highly conserved phenomenon in eukaryotes that occurs in response to various stress conditions. Protein kinases capable of phosphorylating eIF-2alpha have been characterized from mammals and yeast. However, the phenomenon of eIF2-alpha-mediated regulation of protein synthesis and the presence of an eIF-2alpha kinase has not been demonstrated in higher plants. We show that plant eIF-2alpha (peIF-2alpha) and mammalian eIF-2alpha (meIF-2alpha) are phosphorylated similarly by both the double-stranded RNA-binding kinase, pPKR, present in plant ribosome salt wash fractions and the meIF-2alpha kinase, PKR. By several criteria, phosphorylation of peIF-2alpha is directly correlated with pPKR protein and autophosphorylation levels. Significantly, pPKR is capable of specifically phosphorylating Ser51 in a synthetic eIF-2alpha peptide, a key characteristic of the eIF-2alpha kinase family. Taken together, these data support the concept that pPKR is a member of the eIF-2alpha kinase family. In addition, the inhibition of brome mosaic virus RNA in vitro translation in wheat germ lysates by the addition of double-stranded RNA, phosphorylated peIF-2alpha, meIF-2alpha, or activated human PKR suggests that plant protein synthesis may be regulated via phosphorylation of eIF-2alpha.

BackgroundProliferation and expansion of security risks necessitates new measures to ensure authenticity and validation of GMOs. Watermarking and other cryptographic methods are available which conceal and recover the original signature, but in the process reveal the authentication information. In many scenarios watermarking and standard cryptographic methods are necessary but not sufficient and new, more advanced, cryptographic protocols are necessary.ResultsHerein, we present a new crypto protocol, that is applicable in broader settings, and embeds the authentication string indistinguishably from a random element in the signature space and the string is verified or denied without disclosing the actual signature. Results show that in a nucleotide string of 1000, the algorithm gives a correlation of 0.98 or higher between the distribution of the codon and that of E. coli, making the signature virtually invisible.ConclusionsThis algorithm may be used to securely authenticate and validate GMOs without disclosing the actual signature. While this protocol uses watermarking, its novelty is in use of more complex cryptographic techniques based on zero knowledge proofs to encode information.

No abstract

Two different "antisense" oligodeoxynucleotides and their RNA analogues, each complementary to non-overlapping sequences of 51 bases near the 5' end of TMV RNA, inhibit in vitro translation of the genomic RNA in a rabbit reticulocyte lysate. Inhibition is dependent upon complementarity, concentration, and hybridization of the oligomers with TMV RNA. Inhibition is observed at molar ratios of TMV RNA to antisense oligomers as low as 1:1.5. A plateau of inhibition at which 10-25% of the control signal remains is achieved by molar ratios of TMV RNA:antisense DNA or RNA greater than or equal to 1:15. The extent of inhibition is not increased by the simultaneous presence of both complementary fragments. Oligodeoxynucleotides and their RNA analogues identical to the same regions of TMV RNA have no direct effect on translation, however, they can block inhibition by the antisense fragments. Translation of BMV RNA is not affected by any of the oligodeoxynucleotides. Polyacrylamide gel electrophoresis shows translation of TMV p126 is selectively inhibited. We conclude that the observed inhibition of translation is due to direct interference with ribosome function.

Indirect immunofluorescent staining for ml of a 3 X 108 cells/ ml suspension) were detection and identification of Xanthomonas campestris pv. phaseoli in naturally infected bean injected into marginal ear veins of New seed. Plant Disease 67:645-647. Zealand white rabbits (2.2 kg). Repeat injections using antigen (0.5 ml of a 3 Indirect immunofluorescence (IF) was evaluated for detecting and identifying Xanthomonas X 108 cells/ ml suspension) as prepared campestris pv.phaseoli in pure and mixed culture and in Pinto and Navy bean seed. IF was specific before were made at weekly intervals. for 24 X. c. pv. phaseoli isolates tested that were collected from diverse geographical locations. After 6 wk, the rabbits were test-bled and Strong positive cross-reacting fluorescence was not observed when antiserum was tested against the agglutination titer determined by tube numerous X. campestris pathovars as well as other pseudomonads and common bean seed the in titer determin edb bacteria. Cells of X. c. pv. phaseoli were detected on prepared slides in mixed bacterial populations agglutination (2). Antisera with endcontaining as few as 250 colony-forming units (cfu) per milliliter of X. c. pv.phaseoli and 108 cfu/ ml point titers of 5,120 or greater were used of common bean seed bacteria. Cells of X. c. pv. phaseoli were detected in naturally infected bean for immunofluorescence reactions. seed lots with infection levels of 0.01% and in bean seed leachate to which 102 cfu/ ml of X. c. pv. Antigen preparation. Antigen consisted phaseoli were added. IF provides a reliable method for rapid screening of bean seed lots for X. c. pv. of cells of X. c. pv. phaseoli from axenic phaseoli although it is not viewed as the sole test for final determination of seed certification. cultures (as described for immunogen preparation) from naturally infected bean seed collected from symptomatic Pinto bean pods in commercial fields and Common bacterial blight incited by Saettler (13) combined a semiselective from artificially infested bean seed. Bean Xanthomonas campestris pv. phaseoli is enrichment medium and a double-seed samples (1.6 kg) were washed a major seedborne disease of dry beans diffusion assay to detect X. c. pv.phaseoli vigorously on a rotary shaker for 5 mm in (Phaseolus vulgaris L.) (17). Disease in Navy bean seed. 0.05% Tween 80 suspension and surfacemanagement is based primarily on This paper presents information on disinfested with 5.25% NaOCI for 5 min. planting of pathogen-free bean seed. application of an immunofluorescent Seed was then rinsed three times in sterile Although no data exist correlating staining procedure for detecting and PBS and soaked for 24 hr in sterile PBS. percent seed lot infection or inoculum identifying X. c. pv. phaseoli in pure and Leachate (about 150 ml) was concentrated density within individual seeds with field mixed cultures and in naturally infected to about 10 ml by centrifugation at 12,000 losses, a zero tolerance for X. c. pv. bean seed. A preliminary report has been g for 15 min. This ce...

The aberrantly expressed signal transducer and activator of transcription 3 (STAT3) predicts poor prognosis, primarily in estrogen receptor positive (ER(+)) breast cancers. Activated STAT3 is overexpressed in luminal A subtype cells. The mechanisms contributing to the prognosis and/or subtype relevant features of STAT3 in ER(+) breast cancers are through multiple interacting regulatory pathways, including STAT3-MYC, STAT3-ERα, and STAT3-MYC-ERα interactions, as well as the direct action of activated STAT3. These data predict malignant events, treatment responses and a novel enhancer of tamoxifen resistance. The inferred crosstalk between ERα and STAT3 in regulating their shared target gene-METAP2 is partially validated in the luminal B breast cancer cell line-MCF7. Taken together, we identify a poor prognosis relevant gene set within the STAT3 network and a robust one in a subset of patients. VEGFA, ABL1, LYN, IGF2R and STAT3 are suggested therapeutic targets for further study based upon the degree of differential expression in our model.