TIP-1 Translocation onto the Cell Plasma Membrane Is a Molecular Biomarker of Tumor Response to Ionizing Radiation

Tumor response to treatment has been generally assessed with anatomic and functional imaging. Recent development of in vivo molecular and cellular imaging showed promise in time-efficient assessment of the therapeutic efficacy of a prescribed regimen. Currently, the in vivo molecular imaging is limited with shortage of biomarkers and probes with sound biological relevance. We have previously shown in tumor-bearing mice that a hexapeptide (HVGGSSV) demonstrated potentials as a molecular imaging probe to distinguish the tumors responding to ionizing radiation (IR) and/or tyrosine kinase inhibitor treatment from those of non-responding tumors.In this study we have studied biological basis of the HVGGSSV peptide binding within the irradiated tumors by use of tumor-bearing mice and cultured cancer cells. The results indicated that Tax interacting protein 1 (TIP-1, also known as Tax1BP3) is a molecular target that enables the selective binding of the HVGGSSV peptide within irradiated xenograft tumors. Optical imaging and immunohistochemical staining indicated that a TIP-1 specific antibody demonstrated similar biodistribution as the peptide in tumor-bearing mice. The TIP-1 antibody blocked the peptide from binding within irradiated tumors. Studies on both of human and mouse lung cancer cells showed that the intracellular TIP-1 relocated to the plasma membrane surface within the first few hours after exposure to IR and before the onset of treatment associated apoptosis and cell death. TIP-1 relocation onto the cell surface is associated with the reduced proliferation and the enhanced susceptibility to the subsequent IR treatment.This study by use of tumor-bearing mice and cultured cancer cells suggested that imaging of the radiation-inducible TIP-1 translocation onto the cancer cell surface may predict the tumor responsiveness to radiation in a time-efficient manner and thus tailor radiotherapy of cancer

Contrasting plant ecological benefits endowed by naturally occurring EPSPS resistance mutations under glyphosate selection

Concurrent natural evolution of glyphosate resistance single- and double-point EPSPS mutations in weed species provides an opportunity for the estimation of resistance fitness benefits and prediction of equilibrium resistance frequencies in environments under glyphosate selection. Assessment of glyphosate resistance benefit was conducted for the most commonly identified single Pro-106-Ser and less-frequent double TIPS mutations in the EPSPS gene evolved in the global damaging weed Eleusine indica. Under glyphosate selection at the field dose, plants with the single Pro-106-Ser mutation at homozygous state (P106S-rr) showed reduced survival and compromised vegetative growth and fecundity compared with TIPS plants. Whereas both homozygous (TIPS-RR) and compound heterozygous (TIPS-Rr) plants with the double TIPS resistance mutation displayed similar survival rates when exposed to glyphosate, a significantly higher fecundity in the currency of seed number was observed in TIPS-Rr than TIPS-RR plants. The highest plant fitness benefit was associated with the heterozygous TIPS-Rr mutation, whereas plants with the homozygous Pro-106-Ser and TIPS mutations exhibited, respectively, 31% and 39% of the fitness benefit revealed by the TIPS-Rr plants. Populations are predicted to reach stable allelic and genotypic frequencies after 20 years of glyphosate selection at which the WT allele is lost and the stable genotypic polymorphism is comprised by 2% of heterozygous TIPS-Rr, 52% of homozygous TIPS-RR and 46% of homozygous P106S-rr. The high inbreeding nature of E. indica is responsible for the expected frequency decrease in the fittest TIPS-Rr in favour of the homozygous TIPS-RR and P106S-rr. Mutated alleles associated with the glyphosate resistance EPSPS single EPSPS Pro-106-Ser and double TIPS mutations confer contrasting fitness benefits to E. indica under glyphosate treatment and therefore are expected to exhibit contrasting evolution rates in cropping systems under recurrent glyphosate selection.Fil: Vila Aiub, Martin Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentina. University of Western Australia; AustraliaFil: Han, Heping. University of Western Australia; AustraliaFil: Yu, Qin. University of Western Australia; AustraliaFil: García, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; ArgentinaFil: Powles, Stephen B.. University of Western Australia; Australi

On the discovery of genes involved in metabolism-based resistance to herbicides using RNA-Seq transcriptome analysis in Lolium rigidum

Weed control failures due to herbicide resistance are an increasing and worldwide problem significantly impacting crop yields. Herbicide resistance due to increased herbicide metabolism in weeds is not well characterized at the genetic level. An RNA-Seq transcriptome analysis was used to identify genes conferring metabolism-based herbicide resistance (MBHR) in a population (R) of a major global weed (Lolium rigidum), in which resistance to the herbicide diclofop-methyl was experimentally evolved through recurrent selection from a susceptible (S) progenitor population. A reference transcriptome of 19,623 contigs was assembled using 454 sequencing technology on a cDNA library and annotated using UniProt and Pfam databases. Transcriptomic-level gene expression was measured using Illumina 100 bp reads from untreated control, mock, and diclofop-methyl treatments of R and S. Due to the established importance of cytochrome P450 (CytP450), glutathione-S-transferase (GST), and glucosyltransferase (GT) genes in MBHR, 11 contigs with these annotations and higher constitutive expression in untreated R than in untreated S were selected as candidate genes for hypothesis testing, along with 17 additional differentially expressed contigs with annotations related to metabolism or signal transduction. In a forward genetics validation experiment, higher constitutive expression of nine contigs co-segregated with the resistance phenotype in an F2 population, including 3 CytP450, 3 GST, and 1 GT. At least nine genes with heritable increased constitutive expression are associated with MBHR trait. In a physiological validation experiment where 2, 4-D pre-treatment induced diclofop-methyl protection in S individuals due to increased metabolism, seven of the nine genetically-validated contigs were significantly induced. These data help explain accumulation of resistance-endowing genes and rapid evolution of MBHR, and provide the opportunity to improve diagnostics of MBHR using molecular tools such as transcriptional markers. Keywords: 2,4-Dichlorophenoxyacetic acid (2,4-D), diclofop-methyl, evolution, herbicide metabolism, herbicide resistance, next-generation sequencing, transcriptional markers Zur Entdeckung der beteiligten Gene an der metabolischen Herbizidresistenz in Lolium rigidum durch RNA-Seq Transkriptom Analyse. Zusammenfassung Herbizidresistenz ist weltweit ein zunehmendes Problem in der Landwirtschaft, vor allem der enzymatische Abbau von Wirkstoffen bzw. die metabolische Herbizidresistenz (MBHR) ist bislang noch weitgehend unbekannt, besonders auf genetischer Ebene. Um die an einer MBHR beteiligten Gene zu identifizieren wurde das Transkriptom von Herbizid-resistenten und sensitiven Weidelgräsern (Lolium rigidum) in einem RNA-Seq-Ansatz verglichen. Die verwendete Diclofop-Methyl herbizidresistente Population wurde experimentell aus einer sensitiven Population rekurrent selektiert. Ein 19623 Contig umfassendes Referenztranskriptom wurde aus einer cDNA Bibliothek der resistenten Pflanzen durch 454 Sequenzierung erstellt und mit Hilfe von UniProt und Pfam annotiert. Mit 100 bp Illumina-Reads wurde die Genexpression in unbehandelten, mit Blindformulierung und mit formuliertem Herbizid Diclofop-methyl behandelten resistenten und sensitiven Pflanzen untersucht. Es ist bekannt, dass Cytochrom P450 (CytP450), Glutathion-S-transferasen (GST) oder Glycosyltransferasen (GT) eine wichtige Rolle in der MBHR spielen. Elf Contigs, die als solche annotiert und in unbehandelten resistenten Pflanzen konstitutiv höher exprimiert sind als in Sensitiven, wurden als Kandidatengene ausgewählt und getestet. Zusätzlich wurden 17 weitere Contigs untersucht, die zwischen resistenten und sensitiven Pflanzen unterschiedlich exprimiert und durch ihre Annotation in Metabolismus oder Signaltransduktion eingebunden sind. Eine höhere konstitutive Expression konnte in neun selektierten Contigs (u.a. 3 CytP450, 3 GST, 1 GT) auch in der F2 Generation bestätigt werden, cosegregierend mit den resistenten Phänotypen. Daher ist die Expression von mindestens neun Genen mit metabolischer Diclofop-methyl Herbizidresistenz gekoppelt und vererbbar. Eine Vorbehandlung sensitiver Weidelgras-Pflanzen mit 2,4-D induziert metabolische Resistenz gegenüber dem ACCase Inhibitor Diclofop-methyl. Dies wurde verwendet, um die Expression der 28 selektierten Contigs zusätzlich physiologisch zu validieren. Sieben der neun zuvor beschriebenen Contigs sind in der physiologischen Validierung auch in sensitiven Pflanzen durch 2, 4-D induziert. Diese Ergebnisse bieten neue Diagnosemöglichkeiten und verdeutlichen den Zusammenhang zwischen einer vererbbaren Expressionsregulierung von Resistenz vermittelnden Genen und der schnellen Entwicklung von MBHR. Stichwörter: 2,4-Dichlorophenoxyessig Säure (2, 4-D), Diclofop-Methyl, Evolution, Herbizidmetabolisierung, Herbizidresistenz, Next-Generation Sequenzierung, transkriptionelle Marker, Transkriptomi

MSM HIV testing following an online testing intervention in China.

BACKGROUND: Scaling up HIV testing is the first step in the HIV treatment continuum which is important for controlling the HIV epidemic among men who have sex with men (MSM). Following an online HIV testing intervention among MSM, we aim to examine sociodemographic and spatial factors associated with HIV testing. METHODS: We conducted a secondary analysis on data from an online HIV testing intervention among MSM who had never-tested for HIV. The survey was distributed through online networks connected to all provinces and regions of China. Univariate and multivariable analyses were performed to examine factors associated with testing three weeks post-intervention. RESULTS: At three weeks after the intervention, 36% of 624 followed-up MSM underwent HIV testing, 69 men reported positive HIV test results. Having money for sex, ever tested for sexually transmitted infections and intimate partner violence experience were significant factors of post-intervention HIV testing. Students were less likely to undergo HIV testing at follow-up compared to others (adjusted odds ratio=0.69, 95% C.I.=0.47-0.99), adjusted by age and type of intervention. Moderate provincial spatial variation of testing was observed. CONCLUSIONS: While high risk men generally had higher HIV testing rates, some MSM like students had lower testing rates, suggesting the need for further ways to enhance HIV testing in specific MSM communities

Deciphering Signaling Outcomes from a System of Complex Networks

Cellular signal transduction machinery integrates information from multiple inputs to actuate discrete cellular behaviors. Interaction complexity exists when an input modulates the output behavior that results from other inputs. To address whether this machinery is iteratively complex—that is, whether increasing numbers of inputs produce exponential increases in discrete cellular behaviors—we examined the modulated secretion of six cytokines from macrophages in response to up to five-way combinations of an agonist of Toll-like receptor 4, three cytokines, and conditions that activated the cyclic adenosine monophosphate pathway. Although all of the selected ligands showed synergy in paired combinations, few examples of nonadditive outputs were found in response to higher-order combinations. This suggests that most potential interactions are not realized and that unique cellular responses are limited to discrete subsets of ligands and pathways that enhance specific cellular functions

Herbicide resistance endowed by enhanced rates of herbicide metabolism in wild oat (Avena spp.)

The biochemical basis of resistance to the acetyl-coenzyme A carboxylase (ACCase)-inhibiting herbicide diclofop-methyl was investigated in a resistant wild oat population (R1), which does not exhibit a resistant ACCase. Rates of foliar uptake and translocation of [14C]-diclofop were the same in the R1 vs. susceptible (S) populations. However, the level of phytotoxic diclofop acid was always found to be lower in the R1 vs. S plants, with a concomitant higher level (up to 1.7-fold) of nontoxic polar diclofop metabolites in R1 relative to the S plants. These results indicate that a non–target-site-based mechanism of enhanced rate of diclofop acid metabolism confers resistance in population R1. Moreover, the high-performance liquid chromotography elution profile of the major diclofop metabolites in R1 is similar to that of wheat, suggesting resistance in individuals of population R1 involves a wheat-like detoxification system mediated by cytochrome P450 monooxygenases. In addition, lower level of tissue diclofop acid was also observed using nonradioactive ultra-performance liquid chromatography–mass spectrometry analysis in resistant individuals of three other resistant wild oat populations (R2, R3, and R4) known to posses ACCase gene resistance mutations. These results establish that either one or at least two independent resistance mechanisms (target-site ACCase resistance mutations and non–target-site enhanced rates of herbicide metabolism) can be present in individual wild oat plants

Cytosolic and Nucleosolic Calcium Signaling in Response to Osmotic and Salt Stresses Are Independent of Each Other in Roots of Arabidopsis Seedlings

Calcium acts as a universal second messenger in both developmental processes and responses to environmental stresses. Previous research has shown that a number of stimuli can induce [Ca2+] increases in both the cytoplasm and nucleus in plants. However, the relationship between cytosolic and nucleosolic calcium signaling remains obscure. Here, we generated transgenic plants containing a fusion protein, comprising rat parvalbumin (PV) with either a nuclear export sequence (PV-NES) or a nuclear localization sequence (NLS-PV), to selectively buffer the cytosolic or nucleosolic calcium. Firstly, we found that the osmotic stress-induced cytosolic [Ca2+] increase (OICIcyt) and the salt stress-induced cytosolic [Ca2+] increase (SICIcyt) were impaired in the PV-NES lines compared with the Arabidopsis wildtype (WT). Similarly, the osmotic stress-induced nucleosolic [Ca2+] increase (OICInuc) and salt stress-induced nucleosolic [Ca2+] increase (SICInuc) were also disrupted in the NLS-PV lines. These results indicate that PV can effectively buffer the increase of [Ca2+] in response to various stimuli in Arabidopsis. However, the OICIcyt and SICIcyt in the NLS-PV plants were similar to those in the WT, and the OICInuc and SICInuc in the PV-NES plants were also same as those in the WT, suggesting that the cytosolic and nucleosolic calcium dynamics are mutually independent. Furthermore, we found that osmotic stress- and salt stress-inhibited root growth was reduced dramatically in the PV-NES and NLS-PV lines, while the osmotic stress-induced increase of the lateral root primordia was higher in the PV-NES plants than either the WT or NLS-PV plants. In addition, several stress-responsive genes, namely CML37, DREB2A, MYB2, RD29A, and RD29B, displayed diverse expression patterns in response to osmotic and salt stress in the PV-NES and NLS-PV lines when compared with the WT. Together, these results imply that the cytosolic and nucleosolic calcium signaling coexist to play the pivotal roles in the growth and development of plants and their responses to environment stresses