RNA N6-methyladenosine of DHAPAT and PAP involves in regulation of diapause of Bombyx mori via the lipid metabolism pathway.

Environment-induced epigenetics are involved in diapause regulation, but the molecular mechanism that epigenetically couples nutrient metabolism to diapause regulation remains unclear. In this study, we paid special attention to the significant differences in the level of N6-adenosine methylation (m6A) of dihydroxyacetone phosphate acyltransferase (DHAPAT) and phosphatidate phosphatase (PAP) genes in the lipid metabolism pathway of the bivoltine silkworm (Bombyx mori) strain Qiufeng developed from eggs incubated at a normal temperature (QFHT, diapause egg producer) compared to those from eggs incubated at a low temperature (QFLT, non-diapause egg producer). We knocked down DHAPAT in the pupal stage of the QFLT group, resulting in the non-diapause destined eggs becoming diapausing eggs. In the PAP knockdown group, the colour of the non-diapause destined eggs changed from light yellow to pink 3 days after oviposition, but they hatched as normal. Moreover, we validated that YTHDF3 binds to m6A-modified DHAPAT and PAP mRNAs to promote their stability and translation. These results suggest that RNA m6A methylation participates in the diapause regulation of silkworm by changing the expression levels of DHAPAT and PAP and reveal that m6A epigenetic modification can be combined with a lipid metabolism signal pathway to participate in the regulation of insect diapause traits, which provides a clearer image for exploring the physiological basis of insect diapause.

Exorista sorbillans (Diptera: Tachinidae) parasitism shortens host larvae growth duration by regulating ecdysone and juvenile hormone titers in Bombyx mori (Lepidoptera: Bombycidae).

The tachinid fly, Exorista sorbillans, is a notorious ovolarviparous endoparasitoid of the silkworm, Bombyx mori, causing severe damage to silkworm cocoon industry. Silkworm larvae show typically precocious wandering behavior after being parasitized by E. sorbillans; however, the underlying molecular mechanism remains unexplored. Herein, we investigated the changes in the levels of 20-hydroxyecdysone (20E) and juvenile hormone (JH) titer, and they both increased in the hemolymph of parasitized silkworms. Furthermore, we verified the expression patterns of related genes, which showed an upregulation of 20E signaling and biosynthesis genes but a significant downregulation of ecdysone oxidase (EO), a 20E inactivation enzyme, in parasitized silkworms. In addition, related genes of the JH signaling were activated in parasitized silkworms, while related genes of the JH degradation pathway were suppressed, resulting in an increase in JH titer. Notably, the precocious wandering behavior of parasitized silkworms was partly recoverable by silencing the transcriptions of BmCYP302A1 or BmCYP307A1 genes. Our findings suggest that the developmental duration of silkworm post parasitism could be shortened by regulation of 20E and JH titers, which may help silkworm to resist the E. sorbillans infestation. These findings provide a basis for deeper insight into the interplay between silkworms and E. sorbillans and may serve as a reference for the development of a novel approach to control silkworm myiasis.

m6A-dependent mevalonate kinase in juvenile hormone synthesis pathway regulates the diapause process of bivoltine silkworm (Bombyx mori).

BACKGROUND: Research has shown that epigenetic modification are involved the regulation of diapause in bivoltine silkworms (Bombyx mori), but it remains unclear how epigenetic modification in response to environmental signals precisely to regulate the diapause processing of bivoltine B. mori. METHODS AND RESULTS: In this study, the diapause terminated eggs of bivoltine B. mori, Qiufeng (QF) were divided into two groups: a QFHT group incubated at 25 °C with a natural day/night cycle to produce diapause eggs, and a QFLT group incubated at 16.5 °C in darkness to produce non-diapause eggs. On the 3rd day of the pupal stage, the total RNAs of the eggs were extracted and their N6-adenosine methylation (m6A) abundances were analyzed to explore the effects of m6A methylation on diapause in the silkworm. The results showed that 1984 m6A peaks are shared, 1563 in QFLT and 659 in QFHT. The m6A methylation level of the QFLT group was higher than that of the QFHT one in various signaling pathways. The m6A methylation rate of mevalonate kinase (MK) in the insect hormone synthesis pathway was significantly different between the two groups. The knockdown of MK by RNA interference in the pupae of QFLT resulted in females laying diapause eggs rather than non-diapause eggs after mating. CONCLUSIONS: m6A methylation involves in the diapause regulation of bivoltine B. mori by changing the expression levels of MK. This result provides a clearer image of the environmental signals on the regulation of diapause in bivoltine silkworms.

[Accumulation, subcellular distribution, and chemical forms of zinc in three tree species]. / 锌在3ç§ä¹”æœ¨ä¸­çš„ç§¯ç´¯åŠå ¶äºšç»†èƒžåˆ†å¸ƒå’ŒåŒ–å­¦å½¢æ€.

In order to explore the mechanism underlying zinc (Zn) accumulation and tolerance in woody garden species, the effects of different Zn concentrations (0, 250, 500, 1000, 2000 mg·kg-1) on leaf, branch, root biomass and leaf ultrastructure of Koelreuteria paniculata, Ailanthus altissima, and Ginkgo biloba were studied in a pot pollution simulation experiment. The concentration of Zn in plant organs, the subcellular distribution and chemical forms of Zn in leaves and roots were further analyzed. The results showed that all the three species could survive under diffe-rent Zn concentrations, but the biomass of leaves, stems and roots decreased compared with the control. Excessive Zn could lead to cell deformation, cell wall rupture and organelle disintegration of leaves in K. paniculata and A. altissima, while the cells in leaves of G. biloba could maintain normal morphology, indicating that G. biloba had a better tolerance to Zn than K. paniculata and A. altissima. With the increases of Zn concentration, Zn concentration in the organs of the three species showed an increasing trend, and the Zn concentration in K. paniculata and A. altissima was significantly higher than that in G. biloba, indicating that the Zn accumulation ability of K. paniculata and A. altissima was stronger than that of G. biloba. Zn was mainly distributed in the cell walls of leaves and roots, accounting for 26.9%-71.8% and 28.1%-82.6%, respectively. Under the treatment with the highest Zn concentration (2000 mg·kg-1), Zn concentration in the soluble components (mainly vacuoles) could be higher than that in the cell walls. In addition, Zn mainly existed in NaCl-, HAc- and HCl-extracted forms in leaves, accounting for 57.4%-82.7%, and Zn mainly existed in NaCl- and HAc-extracted forms in roots, accounting for 42.8%-67.2%, all of which were forms with relatively low activity. Therefore, cell wall retention, vacuoles segregation and accumulating Zn in less active forms might be important mechanisms underlying Zn accumulation and tolerance in the three trees.

Molecular cloning and comparative analysis of transcripts encoding chemosensory proteins from two plant bugs, Lygus lineolaris and Lygus hesperus.

Chemosensory proteins (CSPs) are soluble carrier proteins typically characterized by a six-helix bundle structure joined by two disulfide bridges and a conserved Cys spacing pattern (C1-X6-8 -C2-X16-21 -C3-X2 -C4). CSPs are functionally diverse with reported roles in chemosensation, immunity, development, and resistance. To expand our molecular understanding of CSP function in plant bugs, we used recently developed transcriptomic resources for Lygus lineolaris and Lygus hesperus to identify 17 and 14 CSP-like sequences, respectively. The Lygus CSPs are orthologous and share significant sequence identity with previously annotated CSPs. Three of the CSPs are predicted to deviate from the typical CSP structure with either five or seven helical segments rather than six. The seven helix CSP is further differentiated by an atypical C3-X3 -C4 Cys spacing motif. Reverse transcriptase PCR-based profiling of CSP transcript abundance in adult L. lineolaris tissues revealed broad expression for most of the CSPs with antenna specific expression limited to a subset of the CSPs. Comparative sequence analyses and homology modeling suggest that variations in the amino acids that comprise the Lygus CSP binding pockets affect the size and nature of the ligands accommodated.

BMDREDD REGULATES THE APOPTOSIS COORDINATING WITH BMDAXX, BMCIDE-B, BMFADD, AND BMCREB IN BMN CELLS.

The apoptosis mechanisms in mammals were investigated relatively clearly. However, little is known about how apoptosis is achieved at a molecular level in silkworm cells. We cloned a caspase homologous gene named BmDredd (where Bm is Bombyx mori and Dredd is death-related ced-3/Nedd2-like caspase) in BmN cells from the ovary of Bm and analyzed its biological information. We constructed the N-terminal, C-terminal, and overexpression vector of BmDredd, respectively. Our results showed that the transcriptional expression level of BmDredd was increased in the apoptotic BmN cells. Furthermore, overexpression of BmDredd increased the caspase-3/7 activity. Simultaneously, RNAi of BmDredd could save BmN cells from apoptosis. The immunofluorescence study showed that BmDredd located at the cytoplasm in normal cell otherwise is found at the nucleus when cells undergo apoptosis. Moreover, we quantified the transcriptional expressions of apoptosis-related genes including BmDredd, BmDaxx (where Daxx is death-domain associated protein), BmCide-b (where Cide-b is cell death inducing DFF45-like effector), BmFadd (Fadd is fas-associated via death domain), and BmCreb (where Creb is cAMP-response element binding protein) in BmN cells with dsRNA interferences to detect the molecular mechanism of apoptosis. In conclusion, BmDredd may function for promoting apoptosis and there are various regulatory interactions among these apoptosis-related genes.

BmPLA2 containing conserved domain WD40 affects the metabolic functions of fat body tissue in silkworm, Bombyx mori.

PLA2 enzyme hydrolyzes arachidonic acid, and other polyunsaturated fatty acids, from the sn-2 position to release free arachidonic acid and a lysophospholipid. Previous studies reported that the PLA2 in invertebrate organisms participates in lipid signaling molecules like arachidonic acid release in immune-associated tissues like hemocytes and fat bodies. In the present study, we cloned the BmPLA2 gene from fat body tissue of silkworm Bombyx mori, which has a total sequence of 1.031 kb with a 31.90 kDa protein. In silico results of BmPLA2 indicated that the protein has a putative WD40 conserved domain and its phylogeny tree clustered with Danaus plexippus species. We investigated the transcriptional expression in development stages and tissues. The highest expression of BmPLA2 was screened in fat body among the studied tissues of third day fifth instar larva, with a high expression on third day fifth instar larva followed by a depression of expression in the wandering stage of the fifth instar larva. The expression of BmPLA2 in female pupa was higher than that of male pupa. Our RNAi-mediated gene silencing results showed highest reduction of BmPLA2 expression in post-24 h followed by post-48 and post-72 h. The BmPLA2-RNAi larvae and pupa could be characterized by pharate adult lethality and underdevelopment. The phenotypic characters of fat body cells in RNAi-induced larva implied that BmPLA2 affects the metabolic functions of fat body tissue in silkworm Bombyx mori.

RNAi KNOCKDOWN OF BmRab3 LED TO LARVA AND PUPA LETHALITY IN SILKWORM Bombyx mori L.

Rab3 GTPases are known to play key a role in vesicular trafficking, and express highest in brain and endocrine tissues. In mammals, Rab3 GTPases are paralogs unlike in insect. In this study, we cloned Rab3 from the silk gland tissue of silkworm Bombyx mori, and identified it as BmRab3. Our in silico analysis indicated that BmRab3 is an isoform with a theoretical isoelectric point and molecular weight of 5.52 and 24.3 kDa, respectively. Further, BmRab3 showed the C-terminal hypervariability for GGT2 site but having two other putative guanine nucleotide exchange factor/GDP dissociation inhibitor interaction sites. Multiple alignment sequence indicated high similarities of BmRab3 with Rab3 isoforms of other species. The phylogeny tree showed BmRab3 clustered between the species of Tribolium castaneum and Aedes aegypti. Meanwhile, the expression analysis of BmRab3 showed the highest expression in middle silk glands (MSGs) than all other tissues in the third day of fifth-instar larva. Simultaneously, we showed the differential expression of BmRab3 in the early instar larva development, followed by higher expression in male than female pupae. In vivo dsRNA interference of BmRab3 reduced the expression of BmRab3 by 75% compared to the control in the MSGs in the first day. But as the worm grew to the third day, the difference of BmRab3 between knockdown and control was only about 10%. The knockdown later witnessed underdevelopment of the larvae and pharate pupae lethality in the overall development of silkworm B. mori L.

Hedgehog signaling pathway regulated the target genes for adipogenesis in silkworm Bombyx mori.

Hedgehog (Hh) signals regulate invertebrate and vertebrate development, yet the role of the pathway in adipose development remains poorly understood. In this report, we found that Hh pathway components are expressed in the fat body of silkworm larvae. Functional analysis of these components in a BmN cell line model revealed that activation of the Hh gene stimulated transcription of Hh pathway components, but inhibited the expression of the adipose marker gene AP2. Conversely, specific RNA interference-mediated knockdown of Hh resulted in increased AP2 expression. This further showed the regulation of Hh signal on the adipose marker gene. In silkworm larval models, enhanced adipocyte differentiation and an increase in adipocyte cell size were observed in silkworms that had been treated with a specific Hh signaling pathway antagonist, cyclopamine. The fat-body-specific Hh blockade tests were consistent with Hh signaling inhibiting silkworm adipogenesis. Our results indicate that the role of Hh signaling in inhibiting fat formation is conserved in vertebrates and invertebrates.

Differential ß-catenin expression levels are associated with morphological features and prognosis of colorectal cancer.

ß-catenin, an epithelial-mesenchymal transition (EMT)-associated marker, is key in the progression of colorectal cancer (CRC). However, the prognostic significance of ß-catenin expression in patients with CRC remains controversial. In the present study, the expression of ß-catenin at the tumor invasive front and the tumor center was investigated, and the correlations amongst ß-catenin differential expression patterns and the clinicopathological characteristics and prognosis of CRC patients were determined. In total, 181 patients that were diagnosed with CRC (as determined by histopathological evaluation) and subjected to surgical resection at the First Hospital of China Medical University between 2000 and 2001 were examined, and CRC specimens were obtained. Immunohistochemical (IHC) staining of ß-catenin was performed for each specimen. The nuclear ß-catenin expression levels were identified to be significantly lower in the tumor center than at the tumor invasive front (immunoreactivity score, 0.05±0.303 versus 2.18±3.917; P<0.001). The presence of nuclear ß-catenin overexpression at the tumor invasive front was found to be correlated with the tumor, node, metastasis stage (P=0.020), lymph node metastasis (P=0.016) and histological differentiation (P=0.006). Survival analysis revealed that reduced membranous expression levels and increased nuclear expression levels of ß-catenin were statistically significantly associated with poor survival times. Furthermore, differential ß-catenin expression levels were associated with aggressive morphological features, EMT and a poor prognosis in CRC. Therefore, IHC analysis of ß-catenin is considered to be a useful marker to predict the prognosis in patients with CRC.

piggyBac transposon-derived targeting shRNA interference against the Bombyx mori nucleopolyhedrovirus (BmNPV).

The Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the most destructive diseases in silkworm, which has caused the main damage to sericulture industry. In this study, we developed a system of RNAi to prevent the BmNPV infection using the piggyBac transposon-derived targeting short hairpin RNA (shRNA) interference. The shRNAs targeting the genes of i.e.-1, lef-1, lef-2 and lef-3 of BmNPV were designed and used to inhibit the intracellular replication or multiplication of BmNPV in Bm cells. The highest activity was presented in the shRNA targeting the i.e.-1c of BmNPV, of which the inhibition rate reached 94.5 % in vitro. Further a stable Bm cell line of piggyBac transposon-derived targeting shRNA interference against BmNPV was established, which has a highly efficacious suppression on virus proliferation. These results indicated that the recombinant shRNA expression system was a useful tool for resistance to BmNPV in vitro. The approach by recombinant shRNAs opens a door of RNAi technology as a strategy that offering technically simpler, cheaper, and quicker gene knockdown for promising research and biotechnology application on silkworm lethal diseases.

The role of pleomorphic adenoma gene-like 2 in gastrointestinal cancer development, progression, and prognosis.

Numerous previous studies have revealed that pleomorphic adenoma gene-like 2 (PLAGL2) is a transcription factor that is active in cancer progression. The aim of this study was to investigate the role of PLAGL2 in the development, progression and prognosis of gastrointestinal cancer. Immunohistochemical analysis revealed that PLAGL2 was expressed in gastrointestinal tumors and adjacent normal tissues. The expression of PLAGL2 was significantly higher in 225 colorectal cancer tissues than in 66 adjacent non-tumor tissues (P = 0.037). However, expression was not significantly different between 286 gastric tumors and 57 adjacent non-tumor tissues (P = 0.352). Moreover, the PLAGL2 expression level significantly correlated with the depth of tumor invasion in colorectal cancer (P = 0.030). However, the PLAGL2 expression level significantly correlated with tumor size in gastric cancer (P = 0.046). Furthermore, we performed survival analyses and found that neither higher nor lower PLAGL2 expression was a prognostic factor in gastrointestinal cancer. Our findings indicate that PALGL2 serves as a tumor oncoprotein in the development and progression of colorectal cancer. However, the role of this protein in the development, progression and prognosis of gastric cancer is uncertain. Further investigation into the molecular mechanisms of PLAGL2 activity in gastrointestinal cancer is warranted.

ILEI: a novel marker for epithelial-mesenchymal transition and poor prognosis in colorectal cancer.

AIMS: Accumulating evidence over the past decade has shown that abnormal activation of epithelial to mesenchymal transition (EMT) contributes to tumour progression and metastasis in colorectal cancer (CRC). In this study, we investigated the expression of interleukin-like EMT inducer (ILEI) and EMT-associated markers (E-cadherin, vimentin) in CRC tissues and determined the correlations between ILEI expression and clinicopathological characteristics, prognosis and EMT in CRC. METHODS AND RESULTS: In total, 194 patients diagnosed with CRC based on histopathological evaluation and those subjected to surgical resection at the First Hospital of China Medical University between 2003 and 2005 were examined. Immunohistochemical staining for ILEI, vimentin and E-cadherin was performed for each specimen. Cytoplasmic overexpression of ILEI usually accompanied down-regulation of E-cadherin and positive expression of vimentin. Conversely, ILEI was simultaneously down-regulated with overexpression of E-cadherin and negative expression of vimentin. ILEI overexpression was associated significantly with T-stage, N-stage, TNM stage and EMT phenotype (P = 0.024, <0.001, <0.001 and <0.001, respectively). Multivariate analysis revealed that ILEI expression was an independent prognostic factor for patient survival. CONCLUSIONS: Our findings indicate that cytoplasmic ILEI expression is a potential marker of EMT and tumour progression in CRC. ILEI is an independent predictive factor associated with poor prognosis in CRC.

Molecular cloning and transcription expression of 3-dehydroecdysone 3α-reductase (3de 3α-reductase) in the different tissues and the developing stage from the silkworm, Bombyx mori L.

Molting in insects is regulated by molting hormones (ecdysteroids), which are also crucial to insect growth, development, and reproduction etc. The decreased ecdysteroid in titre results from enhanced ecdysteroid inactivation reactions including the formation of 3-epiecdyson under ecdysone oxidase and 3-dehydroecdysone 3α-reductase (3DE 3α-reductase). In this paper, we cloned and characterized 3-dehydroecdysone 3α-reductase (3DE 3α-reductase) in different tissues and developing stage of the silkworm, Bombyx mori L. The B. mori 3DE 3α-reductase cDNA contains an ORF 783 bp and the deduced protein sequence containing 260 amino acid residues. Analysis showed the deduced 3DE 3α-reductase belongs to SDR family, which has the NAD(P)-binding domain. Using the Escherichia coli, a high level expression of a fusion polypeptide band of approx. 33 kDa was observed. High transcription of 3DE 3α-reductase was mainly presented in the midgut and hemolymph in the third day of fifth instar larvae in silkworm. The expression of 3DE 3α-reductase at different stages of larval showed that the activity in the early instar was high, and then reduced in late instar. This is parallel to the changes of molting hormone titer in larval. 3DE 3α-reductase is key enzyme in inactivation path of ecdysteroid. The data elucidate the regulation of 3DE 3α-reductase in ecdyteroid titer of its targeting organs and the relationship between the enzyme and metamorphosis.

Clinicopathological significance of claudin-4 in gastric carcinoma.

BACKGROUND: Aberrant expression of claudin proteins has been reported in a variety of cancers. Previous studies have demonstrated that overexpression of claudin may promote tumorigenesis and metastasis through increased invasion and survival of tumor cells. However, the prognostic significance of claudin-4 in gastric cancer remains unclear. METHODS: Immunohistochemistry was used to analyze the expression of claudin-4 in 329 clinical gastric cancer specimens and 44 normal stomach samples, 21 intestinal metaplasia samples, and 21 adjacent precursor lesions dysplasia samples. Statistical analysis methods were used to evaluate the relationship between claudin-4 expression and various clinicopathological parameters. Univariate and multivariate analyses were performed, respectively, to detect the independent predictors of survival. RESULTS: Claudin-4 expression was present in only 7(15.9%) normal gastric samples, but expression of claudin-4 in the intestinal metaplasia lesions and dysplasia lesions was 90.5% and 95.2%, respectively. The expression of claudin-4 was significantly associated with histological differentiation (P < 0.001) and tumor growth patterns (P < 0.001) but not associated with patient survival. However, intermediate type staining of claudin-4 exhibited a trend of correlation with patients' survival (P = 0.023). The five-year survival rate with low expression of claudin-4 in intermediate type (76.4%) was similar to expanding type (64.5%), while the high expression group (46.6%) was closer to infiltrative type (50.7%). CONCLUSIONS: The findings in this study demonstrate claudin-4 aberrant expression in gastric cancer and precursor lesions. The expression of claudin-4 could serve as a basis for identifying gastric cancer of the intermediate type.

Expression and functions of dopa decarboxylase in the silkworm, Bombyx mori was regulated by molting hormone.

Insect molting is an important developmental process of metamorphosis, which is initiated by molting hormone. Molting includes the activation of dermal cells, epidermal cells separation, molting fluid secretion, the formation of new epidermis and old epidermis shed and other series of continuous processes. Polyphenol oxidases, dopa decarboxylase and acetyltransferase are necessary enzymes for this process. Traditionally, the dopa decarboxylase (BmDdc) was considered as an enzyme for epidermal layer's tanning and melanization. This work suggested that dopa decarboxylase is one set of the key enzymes in molting, which closely related with the regulation of ecdysone at the time of biological molting processes. The data showed that the expression peak of dopa decarboxylase in silkworm is higher during molting stage, and decreases after molting. The significant increase in the ecdysone levels of haemolymph was also observed in the artificially fed silkworm larvae with ecdysone hormone. Consistently, the dopa decarboxylase expression was significantly elevated compared to the control. BmDdc RNAi induced dopa decarboxylase expression obviously declined in the silkworm larvae, and caused the pupae appeared no pupation or incomplete pupation. BmDdc was mainly expressed and stored in the peripheral plasma area near the nucleus in BmN cells. In larval, BmDdc was mainly located in the brain and epidermis, which is consisted with its function in sclerotization and melanization. Overall, the results described that the dopa decarboxylase expression is regulated by the molting hormone, and is a necessary enzyme for the silkworm molting.

The clinical significance of mesenchyme forkhead 1 (FoxC2) in gastric carcinoma.

AIMS: Mesenchyme forkhead 1 (FoxC2) is an epithelial-mesenchymal transition (EMT)-inducing factor. Previous studies have demonstrated that FoxC2 binds directly to the promoter region of p120-catenin (p120ctn). The aim of this study was to investigate the clinical significance of FoxC2 expression and the inter-relationship between FoxC2 and p120ctn, in gastric cancer. METHODS AND RESULTS: Immunohistochemistry was used to examine the expression of FoxC2 and p120ctn proteins in 325 gastric cancer samples. Staining for FoxC2 in cancer tissues was markedly stronger than in normal tissues. High FoxC2 expression was associated significantly with differentiation, invasion depth, lymph node metastasis and tumour stage. Patients with high FoxC2 expression or low p120ctn expression had a poor prognosis. In the high p120ctn expression group, the prognosis for patients with low FoxC2 expression was better than for the high FoxC2 group. Moreover, stepwise Cox regression showed that p120ctn was an independent prognostic factor, but FoxC2 in combination with p120ctn was not correlated significantly with survival. CONCLUSIONS: We found that FoxC2 and p120ctn play important roles in the progression and prognosis of gastric cancer. Moreover, FoxC2 and p120ctn should be evaluated further as novel biomarkers and therapeutic targets for gastric cancer treatment.

Characterization of adaptor protein complex-1 in the silkworm, Bombyx mori.

To investigate the function of adaptor protein complex-1 (AP-1) in the silkworm, we characterized AP-1 in the silkworm by RNAi technique and co-localization methods. As a result, AP-1 was found to exist as cytosolic form and membrane-bound form distinguished by phosphate status, showing molecular mass difference. There was relatively more cytosolic form of AP-1 than its membrane-bound counterpart in the silkworm. However, AP-1 distributed predominantly as cytosolic form in BmN cells. Interruption of AP-1 expression via DsRNA was more efficient in BmN cells than in the insect larval, which led to a tendency to dissociation between subcellular organelles like the Golgi apparatus and the mitochondria. Environmental condition changes like relatively higher temperature and treatment with dimethyl sulfoxide can lead to expression variance of AP-1 both in mRNA and protein level. In BmN cells, both the heavy chain γ and light chain σ could clearly co-localize with AP-1 ß, mostly forming pits in cytoplasm. Two isoforms of AP-1 σ corresponded to distinct subcellular distribution pattern, possibly due to C-terminal amino acids difference.

Phenol oxidase is a necessary enzyme for the silkworm molting which is regulated by molting hormone.

Insect molting is an important developmental process of metamorphosis, which is initiated by molting hormone. The molting process includes the activation of dermal cells, epidermal cells separation, molting fluid secretion, the formation of new epidermis and old epidermis excoriation etc. Polyphenol oxidases (PPOs), dopa decarboxylase and acetyltransferase are necessary enzymes for this process. Traditionally, the phenol oxidase was considered as an enzyme for epidermal layer's tanning and melanization. This work suggested that polyphenol oxidases are one set of the key enzymes in molting, which closely related with the role of ecdysone in regulation of molting processes. The data showed that the expression peak of phenol oxidase in silkworm is higher during molting stage, and decreases after molting. The significant increase in the ecdysone levels of haemolymph was observed in the artificially fed silkworm larvae with ecdysone hormone. Consistently, the phenol oxidase expression was significantly elevated compared to the control. PPO1 RNAi induced phenol oxidase expression obviously declined in the silkworm larvae, and caused the pupae incomplete pupation. Overall, the results described that the phenol oxidase expression is regulated by the molting hormone, and is a necessary enzyme for the silkworm molting.

Structural Modification of Graphene Sheets to Create a Dense Network of Defect Sites.

Pt/graphene composites were synthesized by loading platinum nanoparticles onto graphene and etched at 1000 °C in a hydrogen atmosphere. This results in the formation of a dense array of nanostructured defect sites in the graphene, including trenches, nanoribbons, islands, and holes. These defect sites result in an increase in the number of unsaturated carbon atoms and, consequently, enhance the interaction of the CO2 molecules with the etched graphene. This leads to a high capacity for storing CO2; 1 g of the etched samples can store up to 76.3 cm(3) of CO2 at 273 K under ambient pressure.