Activation of the Pathogen-Inducible Gst1 Promoter of Potato after Elicitation by Venturia inaequalis and Erwinia amylovora in Transgenic Apple ( Malus × Domestica )

Rather than using a constitutive promoter to drive transgenes for resistance against fungal and bacterial diseases in genetic engineering of apple (Malus × domestica) cultivars, a promoter induced only after infection was preferred. The ability of the Pgst1 promoter from potato (Solanum tuberosum L.) to drive expression of the gusA reporter gene was determined in two genotypes of apple: the fruit cultivar Royal Gala and the M.26 rootstock. β-glucuronidase activity in the transgenic lines grown in a growth chamber was determined quantitatively using fluorometric assays and compared to the activity in Cauliflower Mosaic Virus (CaMV) 35S promoter-driven transgenic lines. In both apple genotypes, the Pgst1 promoter exhibited a low level of expression after bacterial and fungal inoculation compared to the level obtained with the PCaMV35S promoter (15% and 8% respectively). The Pgst1 promoter was systematically activated in apple at the site of infection with a fungal pathogen. It was also activated after treatment with salicylic acid, but not after wounding. Taken together, these data show that, although the Pgst1 promoter is less active than the PCaMV35S promoter in apple, its pathogen responsiveness could be useful in driving the expression of transgenes to promote bacterial and fungal disease resistanc

239. Ocena efektu przeciwnowotworowego genetycznie modyfikowanej szczepionki komórkowej w mysim modelu raka nerki

CelGenetycznie modyfikowane szczepionki komórkowe (GMTV) mają za zadanie indukcję efektywnej odpowiedzi przeciwnowotworowej. Postanowiliśmy ocenić efekt protekcyjny dwóch różnych GMTV w mysim modelu raka jasnokomórkowego nerki, oraz rolę komórek dendrytycznych w fazie indukcji przeciwnowotworowej odpowiedzi komórkowej.MetodyPrzy wykorzystaniu wektorów retrowirusowych DCCMV-IRES-Neo-H-6 oraz DCCMV-IRES-Neo-IL-6 wprowadzono do komórek mysiego raka jasnokomórkowego (RenCa) skonstruowano dwa rodzaje GMTV: (i) Komórki RenCa wykazujące ekspresję genu Interleukiny-6, (ii) komórki RenCa wykazujące ekspresję genu Hyper-lnterleukiny-6 (sztuczna cytokina będąca białkiem fuzyjnym składającym się z IL-6 powiązanej sztucznym linkerem z agonistycznym rozpuszczalnym receptorem) W celu oceny efektu protekcyjnego GMTV, myszy Balb/c w wieku 8–12 tygodni (8 osobników w jednej grupie eksperymentalnej) immunizowano podając podskórnie w lewe udo, 1×10^6 naświetlonych (80 Gy) komórek (RenCa w/t, RenCa-IL-6, Renca-H6). Po 14 dniach myszom podawano podskórnie w prawe udo wyjściowe komórki RenCa w/t w ilości 5×10^5. Następnie oceniano dynamikę pojawiania się guzów oraz kinetykę ich wzrostu. W celu oceny mechanizmów indukcji odpowiedzi immunologicznej postanowiono ocenić in situ wpływ poszczególnych rodzajów GMTV na komórki dendrytyczne. Myszy Balb/c otrzymywały w okolicy śródbrzusza, podskórnie 2×10^6 napromienionych (80 Gy) komórek Renca w/t, Renca-IL-6, Renca H-6 zawieszonych w Matrigelu™. Po 7 dniach przy pomocy cytometru przepływowego analizowano komórki naciekające Matrigel.Wyniki i podsumowanieImmunizacja myszy komórkami RenCa-H6 okazała się najbardziej efektywna w porównaniu do komórek RenCa w/t i RenCa-IL-6. Jakkolwiek nie przeciwdziałała wzrostowi guzów. Aktywowane komórki DC naciekały najsilniej komórki RenCa-H6. Efekt protekcyjny wyraźnie korelował z ilością aktywowanych komórek DC naciekających miejsce podania GMTV. Intensywna infiltracja miejsca podania GMTV przez komórki DC o wysokim poziomie aktywacji wskazuje na silną role Hyper-Interleukiny-6 w procesie indukcji funkcjonalnej przeciwnowotworowej odpowiedzi immunologicznej

Wetting layer states of InAs/GaAs self-assembled quantum dot structures. Effect of intermixing and capping layer

The authors present a modulated reflectivity study of the wetting layer (WL) states in mol. beam epitaxy grown InAs/GaAs quantum dot (QD) structures designed to emit light in the 1.3-1.5 micro m range. A high sensitivity of the technique has allowed the observation of all optical transitions in the QD system, including low oscillator strength transitions related to QD ground and excited states, and the ones connected with the WL quantum well (QW). The support of WL content profiles, detd. by transmission electron microscopy, has made it possible to analyze in detail the real WL QW confinement potential which was then used for calcg. the optical transition energies. In spite of a very effective WL QW intermixing, mainly due to the Ga-In exchange process (causing the redn. of the max. indium content in the WL layer to about 35% from nominally deposited InAs), the transition energies remain almost unaffected. The latter effect could be explained in effective mass envelope function calcns. taking into account the intermixing of the QW interfaces described within the diffusion model. We have followed the WL-related transitions of 2 closely spaced QD layers grown at different temps., as a function of the In content in the capping layer. Changing the capping layer from pure GaAs to In0.236Ga0.764As has no significant influence on the compn. profile of the WL itself and the WL QW transitions can be usually interpreted properly when based on the cap-induced modification of the confinement potential within a squarelike QW shape approxn. However, some of the obsd. features could be explained only after taking into consideration the effects of intermixing and InGaAs cap layer decompn. [on SciFinder (R)

Epigenetic Regulation of Histone H3 Serine 10 Phosphorylation Status by HCF-1 Proteins in C. elegans and Mammalian Cells

BACKGROUND: The human herpes simplex virus (HSV) host cell factor HCF-1 is a transcriptional coregulator that associates with both histone methyl- and acetyltransferases, and a histone deacetylase and regulates cell proliferation and division. In HSV-infected cells, HCF-1 associates with the viral protein VP16 to promote formation of a multiprotein-DNA transcriptional activator complex. The ability of HCF proteins to stabilize this VP16-induced complex has been conserved in diverse animal species including Drosophila melanogaster and Caenorhabditis elegans suggesting that VP16 targets a conserved cellular function of HCF-1. METHODOLOGY/PRINCIPAL FINDINGS: To investigate the role of HCF proteins in animal development, we have characterized the effects of loss of the HCF-1 homolog in C. elegans, called Ce HCF-1. Two large hcf-1 deletion mutants (pk924 and ok559) are viable but display reduced fertility. Loss of Ce HCF-1 protein at reduced temperatures (e.g., 12 degrees C), however, leads to a high incidence of embryonic lethality and early embryonic mitotic and cytokinetic defects reminiscent of mammalian cell-division defects upon loss of HCF-1 function. Even when viable, however, at normal temperature, mutant embryos display reduced levels of phospho-histone H3 serine 10 (H3S10P), a modification implicated in both transcriptional and mitotic regulation. Mammalian cells with defective HCF-1 also display defects in mitotic H3S10P status. CONCLUSIONS/SIGNIFICANCE: These results suggest that HCF-1 proteins possess conserved roles in the regulation of cell division and mitotic histone phosphorylation

Classification and Prediction of Survival in Patients with the Leukemic Phase of Cutaneous T Cell Lymphoma

We have used cDNA arrays to investigate gene expression patterns in peripheral blood mononuclear cells from patients with leukemic forms of cutaneous T cell lymphoma, primarily Sezary syndrome (SS). When expression data for patients with high blood tumor burden (Sezary cells >60% of the lymphocytes) and healthy controls are compared by Student's t test, at P < 0.01, we find 385 genes to be differentially expressed. Highly overexpressed genes include Th2 cells–specific transcription factors Gata-3 and Jun B, as well as integrin β1, proteoglycan 2, the RhoB oncogene, and dual specificity phosphatase 1. Highly underexpressed genes include CD26, Stat-4, and the IL-1 receptors. Message for plastin-T, not normally expressed in lymphoid tissue, is detected only in patient samples and may provide a new marker for diagnosis. Using penalized discriminant analysis, we have identified a panel of eight genes that can distinguish SS in patients with as few as 5% circulating tumor cells. This suggests that, even in early disease, Sezary cells produce chemokines and cytokines that induce an expression profile in the peripheral blood distinctive to SS. Finally, we show that using 10 genes, we can identify a class of patients who will succumb within six months of sampling regardless of their tumor burden

Parallel Retention of Pdx2 Genes in Cartilaginous Fish and Coelacanths

The Pdx1 or Ipf1 gene encodes an important homeodomain-containing protein with key roles in pancreas development and function. Mutations in human PDX1 are implicated in developmental defects and disease of the pancreas. Extensive research, including genome sequencing, has indicated that Pdx1 is the only member of its gene family in mammals, birds, amphibians, and ray-finned fish, and with the exception of teleost fish, this gene forms part of the ParaHox gene cluster along with Gsx1 and Cdx2. The ParaHox cluster, however, is a remnant of a 4-fold genome duplication; the three other ParaHox paralogues lack a Pdx-like gene in all vertebrate genomes examined to date. We have used bacterial artificial chromosome cloning and synteny analysis to show that the ancestor of living jawed vertebrates in fact had more ParaHox genes, including two Pdx genes (Pdx1 and Pdx2). Surprisingly, the two Pdx genes have been retained in parallel in two quite distantly related lineages, the cartilaginous fish (sharks, skates, and chimeras) and the Indonesian coelacanth, Latimeria menadoensis. The Pdx2 gene has been lost independently in ray-finned fish and in tetrapods

The Suppressor of AAC2 Lethality SAL1 Modulates Sensitivity of Heterologously Expressed Artemia ADP/ATP Carrier to Bongkrekate in Yeast

The ADP/ATP carrier protein (AAC) expressed in Artemia franciscana is refractory to bongkrekate. We generated two strains of Saccharomyces cerevisiae where AAC1 and AAC3 were inactivated and the AAC2 isoform was replaced with Artemia AAC containing a hemagglutinin tag (ArAAC-HA). In one of the strains the suppressor of ΔAAC2 lethality, SAL1, was also inactivated but a plasmid coding for yeast AAC2 was included, because the ArAACΔsal1Δ strain was lethal. In both strains ArAAC-HA was expressed and correctly localized to the mitochondria. Peptide sequencing of ArAAC expressed in Artemia and that expressed in the modified yeasts revealed identical amino acid sequences. The isolated mitochondria from both modified strains developed 85% of the membrane potential attained by mitochondria of control strains, and addition of ADP yielded bongkrekate-sensitive depolarizations implying acquired sensitivity of ArAAC-mediated adenine nucleotide exchange to this poison, independent from SAL1. However, growth of ArAAC-expressing yeasts in glycerol-containing media was arrested by bongkrekate only in the presence of SAL1. We conclude that the mitochondrial environment of yeasts relying on respiratory growth conferred sensitivity of ArAAC to bongkrekate in a SAL1-dependent manner. © 2013 Wysocka-Kapcinska et al