GPC3 and PEG10 peptides associated with placental gp96 elicit specific T cell immunity against hepatocellular carcinoma.

The placenta and tumors can exhibit a shared expression profile of proto-oncogenes. The basis of placenta-derived heat shock protein gp96, which induces prophylactic and therapeutic T cell responses against cancer including hepatocellular carcinoma (HCC), remains unknown. Here, we identified the associated long peptides from human placental gp96 using matrix-assisted laser desorption/ionization-time-of-flight and mass spectrometry and analyzed the achieved proteins through disease enrichment analysis. We found that placental gp96 binds to numerous peptides derived from 73 proteins that could be enriched in multiple cancer types. Epitope-harboring peptides from glypican 3 (GPC3) and paternally expressed gene 10 (PEG10) were the major antigens mediating anti-HCC T cell immunity. Molecular docking analysis showed that the GPC3- and PEG10-derived peptides, mainly obtained from the cytotrophoblast layer of the mature placenta, bind to the lumenal channel and client-bound domain of the gp96 dimer. Immunization with bone marrow-derived dendritic cells pulsed with recombinant gp96-GPC3 or recombinant gp96-PEG10 peptide complex induced specific T cell responses, and T cell transfusion led to pronounced growth inhibition of HCC tumors in nude mice. We demonstrated that the chaperone gp96 can capture antigenic peptides as an efficient approach for defining tumor rejection oncoantigens in the placenta and provide a basis for developing GPC3 and PEG10 peptide-based vaccines against HCC. This study provides insight into the underlying mechanism of the antitumor response mediated by embryonic antigens from fetal tissues, and this will incite more studies to identify potential tumor rejection antigens from placenta.

Climate change did not alter the effects of Bt maize on soil Collembola in northeast China.

Bt maize is being increasingly cultivated worldwide as the effects of climate change are increasing globally. Bt maize IE09S034 and its near-isogenic non-Bt maize Zong 31 were used to investigate whether climate change alters the effects of Bt maize on soil Collembola. Warming and drought conditions were simulated using open-top chambers (OTC), and their effects on soil Collembola were evaluated. We found that the maize type had no significant effect on Collembola; however, the abundance and diversity of Collembola were significantly higher in the OTC than outside at the seedling stage; they were significantly lower in the OTC at the heading and mature stages. The interactions of the maize type with the OTC had no effect on these parameters. Therefore, Bt maize had no significant effect on soil Collembola, and the effects of climate warming and drought on soil Collembola depended on the ambient climatic conditions. When the temperature was low, collembolan abundance and diversity were promoted by warming; however, when the temperature was high and the humidity was low, collembolan abundance and diversity were inhibited by warming and drought. The climate changes simulated by the OTC did not alter the effects of Bt maize on soil Collembola.

Biochemical analyses demonstrate that Bt maize has no adverse effects on Eisenia fetida.

The potential effects of Bt (Bacillus thuringiensis) maize on non-target organisms should be evaluated before such maize is commercially planted. Earthworms play an indispensable role in the soil ecosystem; act as important bio-indicators of soil quality and environmental pollution. Therefore, earthworms are often used as the object to evaluate the non-target effect of Bt maize. To accelerate the commercialization of transgenic maize in China, a 90-day Eisenia fetida feeding experiment was conducted to evaluate the potential effects of Bt maize line, BT799-which was developed by China Agricultural University and contains the Cry1Ac gene-and its non-Bt conventional isoline-Zheng 58-on E. fetida. Our results showed that the Bt maize line had no significant effects on the growth, reproduction, or enzymatic activities of these earthworms. In summary, Bt maize had no toxic effects on E. fetida.

Construction of a Novel Degradation Model of Bacillus thuringiensis Protein in Soil and Its Application in Estimation of the Degradation Dynamics of Bt-Cry1Ah Protein.

Bacillus thuringiensis (Bt) protein expressed by genetically modified (GM) crops is released into the soil ecosystem, where it accumulates for a long time; therefore, degradation of Bt protein has gained increased attention for environmental risk assessments. A first-order kinetic model (Y = ae-b*X) is usually used to evaluate the degradation of Bt proteins, including Bt-Cry1Ab and Bt-Cry1Ac; this has some limitations regarding the precise fitting and explanation of the influence of various factors on Bt protein degradation in the later stage. Therefore, to amend these limitations, we report a new degradation model Y = Y0 + ae-b*X. The effects of soil temperature, water content, soil types, and soil sterilization on the degradation of Bt-Cry1Ah protein in soil were estimated in a 96d long laboratory study using a GM maize leaf-soil mixture. The results showed that the Bt-Cry1Ah protein degraded rapidly in the early stage and then slowly in the middle and late stages. Temperature was identified as the key factor affecting the degradation of Cry1Ah protein-a relatively higher temperature favored the degradation. The degradation rate of Cry1Ah protein was the fastest when the water content was 33 and 20% in the early and later stages, respectively. The soil types had a significant effect on the degradation of Cry1Ah protein. Moreover, soil sterilization slowed down the rate of protein degradation in both the early and later stages. In conclusion, the model Y = Y0 + ae-b*X established in this study provided a more robust model for exploring and simulating the degradation of Bt protein in soil growing GM crops and overcame the shortcomings of the Y = ae-b*X model. The findings of this study enriched the understanding of Bt protein degradation in soil ecosystems. They would be helpful for evaluating the environmental safety of GM crops.

Use of Taxonomic and Trait-Based Approaches to Evaluate the Effect of Bt maize Expressing Cry1Ie Protein on Non-Target Collembola: A Case Study in Northeast China.

To evaluate the effect of Bt maize expressing Cry1Ie protein on non-target soil Collembola, a two-year field study was conducted in Northeast China. Bt maize line IE09S034 and its near isoline Zong 31 were selected as experimental crops; we investigated the collembolan community using both taxonomic and trait-based approaches, and elucidated the relationship between environmental variables and the collembolan community using redundancy analysis (RDA).The ANOVA results showed that maize variety neither had significant effect on the parameters based on taxonomic approach (abundance, species richness, Shannon-Wiener index, Pielou's evenness index), nor on the parameters based on trait-based approach (ocelli number, body length, pigmentation level, and furcula development) in either year. The results of RDA also showed that maize variety did not affect collembolan community significantly. These results suggest that two years cultivation of cry1Ie maize does not affect collembolan community in Northeast China.

Toxicological and biochemical analyses demonstrate no toxic effect of Bt maize on the Folsomia candida.

The potential effects of Bt (Bacillus thuringiensis) maize on non-target organisms must be conducted before the Bt maize is commercially planted. Folsomia candida is one of the non-target organisms of Bt maize, also as an important indicator of soil quality and environmental pollution. In this study, a 90-day F. candida feeding test were conducted to evaluate the potential effects of two Bt maize lines IE09S034 and BT799 and their non-Bt conventional isolines Zong 31 and Zheng 58. The results show that Bt maize lines had no significant effects on the survival rate, reproduction, adult body length, larval body length, and the activities of acetyl cholinesterase, catalase and superoxide dismutase on the F. candida. Namely, Bt maize had no toxic effects on the F. candida.

Use of Taxonomic and Trait-Based Approaches to Evaluate the Effects of Transgenic Cry1Ac Corn on the Community Characteristics of Soil Collembola.

Nontarget soil fauna in fields planted with transgenic Bt crops may be influenced by the Bt protein following crop establishment. Here, we investigated the effects of transgenic Cry1Ac corn Bt-799 on soil Collembola using both taxonomic and trait-based approaches in an experimental field in northeast China from 2014 to 2015. The relationship between the collembolan community and environmental variables (including corn type) was investigated using redundancy analysis (RDA). In May 2015, we found that collembolan species richness, Shannon-Wiener's index, and all trait values examined in transgenic Cry1Ac corn (Event Bt-799, Bt corn) were lower than in its near isoline (Zheng 58, non-Bt corn). However, this significant influence did not persist in the subsequent corn growth period. For all other sampling periods over 2-yr study, all indices based on taxonomic (abundance, species richness, and Shannon-Wiener's index) and trait-based (number of ocelli, body size, pigmentation, furcula development, and antennae length) approaches in Bt corn were not significantly different from non-Bt corn. The results showed no significant difference between the two corn types in the repeated-measures analysis of variance (ANOVA). Moreover, redundancy analysis (RDA) showed that corn type only explained 1% of the variation in the collembolan community. These results suggested that Bt corn did not affect collembolan community characteristics in the short term.

No impact of transgenic cry1Ie maize on the diversity, abundance and composition of soil fauna in a 2-year field trial.

Soil fauna play an essential role in the soil ecosystem, but they may be influenced by insecticidal Cry proteins derived from Bacillus thuringiensis (Bt) maize. In this study, a 2-year field trial was conducted to study the effects of transgenic cry1Ie maize, a type of Bt maize (Event IE09S034), on soil fauna, with the near-isogenic line non-Bt maize (Zong 31) as a control. The soil animals were collected with Macfadyen heat extractor and hand-sorting methods, respectively, and their diversity, abundance and community composition were calculated. Then, the effects of maize type, year, sampling time and soil environmental factors on the soil fauna were evaluated by repeated-measures ANOVA, redundancy analysis (RDA) and nonmetric multidimensional scaling (nMDS). Repeated-measures ANOVA showed that the diversity and abundance of the soil fauna were not affected by maize type, while they were significantly influenced by year and sampling time. Furthermore, for both the Macfadyen and hand-sorting methods, RDA indicated that soil fauna community composition was not correlated with maize type (Bt and non-Bt maize) but was significantly correlated with year, sampling time and root biomass. In addition, it was significantly related to soil pH according to the hand-sorting method. nMDS indicated that soil fauna community composition was significantly correlated with year and sampling time; however, it was not associated with maize type. In this study, we collected soil faunal samples according to the Macfadyen and hand-sorting methods and processed the obtained data with ANOVA, RDA, and nMDS in three ways, and our data indicate that transgenic cry1Ie maize (Event IE09S034) had no substantial influence on the diversity, abundance or community composition of the soil fauna.