In the transcriptome surveys of Laodelphax striatellus, several cDNA sequences showed a high level of similarities to the insect picorna-like virus genomes. Interestingly, there was no sequence similarity between picorna-like virus sequences from the RSV-viruliferous and those from the non-viruliferous L. striatellus. Picorna-like virus from the non-viruliferous L. striatellus was a geographical isolate of Himetobi P virus (HiPV). The genome of the HiPV was 9,272 nt in length excluding the poly(A) tail and contained two open reading frames (ORFs), which were separated by a 176 nt intergenic region that functions as an internal ribosome entry site (IRES). The 5' ORF encodes the non-structural proteins and the 3' ORF encodes the capsid proteins. The partial genomic RNA of the picorna-like virus from the RSV-viruliferous L. striatellus, LsPV-2, was 8,769 nt in length excluding the poly(A) tail and contained a single, large open reading frame (nt 1–8,535) encoding a 2,845 aa polyprotein. In terms of sequence similarity, identity, and genome organization, LsPV-2 resembled insect picornalike viruses belonging to the family Iflaviridae. A phylogenetic analysis based on RNA-dependent RNA polymerase (RdRp) sequence showed that LsPV-2 was most closely related to the deformed wing virus (DWV). The HiPV and LsPV-2 were incompatible each other in L. striatellus, suggesting that these two picorna-like viruses may have important functions in transmission of the RSV.

Rice stripe virus (RSV), the type member of the genus Tenuivirus, causes rice stripe disease and the viral transmission is mediated through the sucking by small brown planthopper, Laodelphax striatellus. Considerations have been mainly focused on the protection of rice from RSV and/or the planthopper, rather than the interaction between RSV and the insect. To clarify the interaction, in this work, mRNA was extracted from RSV-viruliferous planthopper with non-viruliferous control, and expressed sequence tag (EST) databases were generated based on 454 GS-FLX pyrosequencing technology for comparative analysis. RSV-viruliferous planthopper had ca. 2500 isotigs, which included genes on biological process (19%), cellular component (13%), molecular function (22%) and no hits (46%) from gene ontology (GO) analysis; this structure was similar to the control. However, in the viruliferous planthopper, 109 isotigs were up-regulated and 660 isotigs were down-regulated, compared to the non-viruliferous control. These RSV-dependently regulated genes may have important function in the behavior of planthopper or the transmission of RSV.

본 시험은 국화에서의 기형화 발생과 DNA 메틸레 이션에 관여하는 S-adenosyl-L-homocysteine hydrolase (SAHH) 유전자 발현과의 관련성을 검토하기 위해 수행 하였다. 스프레이 국화 ‘Lerbin’은 단일후 14일에서 27일 사이에 고온과 장일 조건에서 기형화가 발생되어, 35/20oC 의 고온과 14시간의 장일 처리 할 경우 25/20oC(12 h/ 12 h)에 비해 설상화가 2배 이상 증가하는 양상을 보였다. 스프레이 국화 ‘Lerbin’에서 분리한 전장 cDNA (DgSAHH) 는 크기가 1455 bp로 다른 작물과 90%이상의 높은 상동 성을 나타내었다. 국화의 DgSAHH 유전자 발현은 기형화 발생을 유도하는 고온과 장일 조건에서 크게 감소하였다. 이러한 결과를 통해 국화의 화아 발달에 있어서 DgSAHH 유전자 발현은 온도와 일장의 영향을 받으며, 억제된 이 유전자의 발현이 기형화 발생에 관여할 수 있을 것으로 판단된다.

Bee venom contains a variety of peptides and enzymes, including serine proteases. While the presence of serine proteases in bee venom has been demonstrated, the role of these proteins in bee venom has not been elucidated. Furthermore, there is currently no information available regarding the melanization response or the fibrin(ogen)olytic activity of bee venom serine protease, and the molecular mechanism of its action remains unknown. Here we show that bee venom serine protease (Bi-VSP) is a multifunctional enzyme. In insects, Bi-VSP acts as an arthropod prophenoloxidase (proPO)-activating factor (PPAF), thereby triggering the phenoloxidase (PO) cascade. Bi-VSP injected through the stinger induces a lethal melanization response in target insects by modulating the innate immune response. In mammals, Bi-VSP acts similarly to snake venom serine protease, which exhibits fibrin(ogen)olytic activity. Bi-VSP activates prothrombin and directly degrades fibrinogen into fibrin degradation products, defining roles forBi-VSP as a prothrombin activator, a thrombin-like protease, and a plasmin-like protease. These findings provide a novel view of the mechanism of bee venom in which the bee venom serine protease kills target insects via a melanization strategy and exhibits fibrin(ogen)olytic activity.

원자전달 라디칼 중합을 이용하여 poly(epichlorohydrine) (PECH)를 주사슬로 한 양친성 가지형 공중합체를 합성하였다. PECH로부터 poly(methyl methacrylate)(PMMA) 및 poly(butyl methacrylate)(PBMA)의 가지형 중합이 성공적임을 1H NMR과 FT-IR분석을 통해 확인하였다. 합성한 가지형 공중합체에 KI나 LiI 염을 도입하였을 때, ether 신축진동 피크가 낮은 wavenumber영역으로 이동하였으며, 이는 배위결합 상호작용 때문이다. PECH-g-PBMA 복합체의 이온 전도도는 PECH-g-PMMA 복합체에 비해 항상 높게 나타났는데, 이는 고무상인 PBMA 사슬의 높은 이동성으로부터 기인한 것으로 확인되었다. 최고 이온전도도 값은 질량비 10 wt%의 KI가 도입된 PECH-g-PBMA 전해질체에서 2.7 × 10 -5 S/cm로 나타났다.

Bee venom contains a variety of peptides and enzymes, including serine proteases. Here we describe the molecular cloning and characterization of a serine protease (Bt-VSP) isolated from the venom of the bumblebee Bombus terrestris. The Bt-VSP gene consists of six exons encoding a 358-amino acid protein. The form of Bt-VSP detected in bee venom was the 34-kDa mature protein, which is created by cleavage of the catalytic domain of Bt-proVSP between Arg111 and Val112. Bt-VSP activates prothrombin and directly degrades fibrinogen into fibrin degradation products, defining roles for Bt-VSP as a prothrombin activator, a thrombin-like protease, and a plasmin-like protease. The finding that Bt-VSP acts as a fibrin(ogen)olytic enzyme is similar to a previous finding that Bi-VSP, a venom serine protease of B. ignitus, exhibits fibrin(ogen)olytic activity. We also compared major venom components in honeybee and bumblebee, and found that bumblebee venom contains a larger amount of serine protease. Furthermore, unlike bumblebee venom, which exhibits fibrin(ogen)olytic activity owing to the presence of a serine protease, it is likely that honeybee venom lacks fibrin(ogen)olytic activity.

The purpose of this study was to investigate the distribution of trig˗ ger points(TrPs) on athletes with various sporting background. To achieve the purpose, a study was carried out through a survey from 180 athletes involved in 6 selected sports at Yong-In University. Selected sports included Judo, Taekwondo, Kendo, Ssi-reum(Korean traditional wrestling), Boxing, and Golf. An interview type survey and physical examination were conducted with each thirty athletes from each of the selected sports groups. Technical statistic(SPSS 15.0) was used to analyze the distribution of TrPs on these athletes. The most common TrPs observed in muscles of Trapezius, Quadratus Lumborum, Quadriceps in Judo. In Taekwondo, it was on the trapez˗ ius and triceps surae. Kendo athletes had TrPs at sites of trapezius, brachioradialis and triceps surae. Ssirem athletes were found to have TrPs on trapezius, deltoid and quadrates lumborum. In boxers, TrPs appearing at trapezius and brachioradialis were observed. Finally, Golf players were seen to have TrPs at trapezius, quadrates lumbo˗ rum and brachioradialis. Hence, the analysis shows that there are significant differences of the distribution of TrPs according to the different sport items of the athlete.

We report for the first time the occurrence of DWV-infected bumble bees (Bombus ignitus). For the present study, the detection of DWV virus from the female and male bumble bee was investigated in the same colony. The Deformed wing virus (DWV) of honeybee (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. Using specific RT-PCR protocols for the detection of DWV followed by sequencing of the PCR products we could demonstrate that the bumble bees were indeed infected with DWV. The virus was detected from Bombus ignitus, and its partial DWV gene was cloned and sequenced. The partial DWV gene encoding the polyprotein is 711-nt of 235 amino acid residues. The deduced nucleotide sequence of the polyprotein partial gene of DWV showed 96.9%, 96.2%, 96.8%, and 96.5% homology to other structure polyprotein partial gene of DWV from insects, respectively. Phylogenetic analysis further conformed that the deduced nucleotide sequence of the polyprotein partial gene of DWV divided to the outside tree. We describe the first time that presence of Deformed wing virus(DWV) from bumble bee(Bombus terrestris) in korea using RT-PCR.

Phospholipid-hydroperoxide glutathione peroxide (PHGPx) is an antioxidant enzyme that reduces lipid hydroperoxides in biomembranes. Here, we cloned and characterized cys-PHGPx from the bumblebee Bombus ignitus (Bi-PHGPx). The Bi-PHGPx gene consists of 4 exons, encoding 168 amino acid residues with a canonical cys-codon at residue 45 and active site residues Gln82 and Trp134. Recombinant Bi-PHGPx, expressed as a 19 kDa protein in baculovirus-infected insect cells, exhibited enzymatic activity against PLPC-OOH and H2O2 using glutathione as an electron donor. Tissue distribution analyses showed the presence of Bi-PHGPx in all tissues examined. Bi-PHGPx transcripts were upregulated by stresses, such as wounding, H2O2 exposure, external temperature shock, and starvation. Under H2O2 overload, the RNA interference (RNAi)-induced thioredoxin peroxidase (BiTPx1)-knock-down B. ignitus worker bees showed upregulated expression of Bi-PHGPx in the fat body. These results indicate that Bi-PHGPx is a stress-inducible antioxidant enzyme that acts on phospholipid hydroperoxide and H2O2.

Bee venom contains a variety of toxic enzymes and peptides. One of the major components of bumblebee venom is bombolitin, which is the most abundant venom constituent and biologically similar to melittin. Here, we first show the molecular cloning and antimicrobial activity of the venom bombolitin from the bumblebee Bombus ignitus. The B. ignitus venom bombolitin gene consists of 2 exons, encoding 56 amino acid residues. The bombolitin purified from B. ignitus venom was the 2104 Da mature peptide with 18 amino acid residues, which are created by cleavage of the probombolitin domain between Ala38 and Leu39. We examined the pattern of bombolitin expression to confirm that it is a component of bumblebee venom. B. igniutus venom bombolitin exhibits venom gland-specific expression. We also investigated the venom bombolitin for antimicrobial properties against bacteria and fungi. The venom bombolitin showed high antibacterial activity against both Gram-negative and Gram-positive bacteria. Most interestingly, the venom bombolitin showed high antifungal activity against Fulvia falva, a leaf mold, and Alternaria radicia, a black rot. These antimicrobial profiles of B. ignitus venom bombolitin reported herein will be useful in the application for potential antimicrobial agents.

We cloned and characterized two peroxiredoxins (Prxs), BiPrx1 (a 1-Cys Prx) and BiTPx1 (a 2-Cys Prx) from the bumblebee Bombus ignitus. The BiPrx1 gene consists of 5 exons, encoding 220 amino acid residues with one conserved cysteine residue. The BiTPx1 gene consists of three exons, encoding 195 amino acid residues with 2 conserved cysteine residues. Recombinant BiPrx1 (27 kDa) and BiTPx1 (25 kDa), expressed in baculovirus-infected insect Sf9 cells, reduced H2O2 in the presence of electrons donated by dithiothreitol. Unlike BiTPx1, however, BiPrx1 did not show reduction activity when thioredoxin was used as the electron donor. Both BiPrx1 and BiTPx1 protected super-coiled DNA from damage by metal-catalyzed oxidation (MCO) in vitro. Tissue distribution analyses showed the presence of BiPrx1 and BiTPx1 in the fat body, midgut, muscle and epidermis, but not in the hemolymph, suggesting that BiPrx1 and BiTPx1 are not secretable. When H2O2 was injected into B. ignitus bees, BiPrx1 and BiTPx1 transcripts were acutely up-regulated in the fat body tissues. We also demonstrated regulation of BiPrx1 and BiTPx1 expression via reduction of transcript levels in the fat body with RNA interference (RNAi). Under H2O2 overload, the RNAi-induced BiPrx1 knock-down B. ignitus worker bees showed up-regulated expression of BiTPx1. Reciprocally, BiTPx1 RNAi knockdowns showed up-regulated BiPrx1 expression in the fat body. These results indicate that loss of expression of BiPrx1 or BiTPx1 is compensated by up-regulation of expression of the other peroxidase in response to H2O2 overload.

Background: Proteolytic enzymes are involved in insect molting and metamorphosis and play a vital role in the programmed cell death of obsolete organs. Here we show the expression profile of cathepsin B in the fat body of the silkworm Bombyx mori during development. We also compared the expression profile of B. mori cathepsins B (BmCatB) and D (BmCatD) in the fat body during the larval-pupal transformation of B. mori in the BmCatB or BmCatD RNA interference (RNAi) process. Results: BmCatB is ecdysone-induced and expressed in the fat body of B. mori during the molting, and the larval-pupal and pupal-adult transformations, and its expression leads to programmed cell death. In particular, BmCatB is highly expressed in the fat body of B. mori during the larval-pupal transformation and BmCatB RNAi treatment resulted in the arrest of the larval-pupal transformation. RNAi-treated BmCatB knock-down sustained the expression of BmCatD during the larval-pupal transformation. On the other hand, BmCatD RNAi up-regulated the expression of BmCatB in the fat body of final instar larvae. Conclusion: Based on these results, we conclude that BmCatB is involved in the programmed cell death of the fat body during B. mori metamorphosis and that BmCatB and BmCatD contribute collaboratively to B. mori metamorphosis

Among bee venom proteins, phospholipase A2 (PLA2) is critical one of bee venom components to defend against predators intruders. In this study, PLA2 gene from cDNA libarary using the venom glands of Bombus ignitus worker bees(BiVn-PLA2) was cloned and characterized. BiVn-PLA2 spans 2211 bp and consists of three introns and four exons encoding 180 amino acid residues. BiVn-PLA2 shares high levels of identity with a bumblebee, B. terristris (89% protein sequence identity), B. pennsylvanicus (88%), and a honey bee, Apis mellifera (53%). Northern blot analysis revealed that BiVn-PLA2 is expressed in venom gland, indicating that BiVn-PLA2 is one of the venom components of B. ignitus. To determine BiVn-PLA2 of venom components from venom sac, N-terminal amino acid sequencing of a putative BiVn-PLA2 (the purified 18 kDa) was performed by Edman degradation. The N-terminal amino acid sequencing of the 18 kDa protein was coincident with the N-terminal amino acid residues of the mature BiVn-PLA2 and the 18 kDa protein catalysed the hydrolysis of DBPC subs trate[1-O-(6-Dabcyl-aminohexanoyl)-2-O-(12-(5-B ODIPY-entanoyl) aminododecanoyl)-sn-glyceryl phosphatidylcholine] that is a sensitive fluorogenic probe for PLA2 activation. Western blot analysis revealed that BiVn-PLA2 is expressed in the venom gland, stored in the venom sac, and then emitted throughout sting apparatus. Finally, to test BiVn-PLA2 toxicity, BiVn-PLA2 was adjusted to a insect cell (Sf9) at different concentrations (1-30 μg/2×105 cells). The apoptotic cell death assay results showed that the cell survival decreased with increasing concentrations (1-30 μg/2×105 cells).

Transferrin and ferritin are iron-binding proteins involved in transport and storage of iron as part of iron metabolism. Here, we describe the cDNA cloning and characterization of transferrin (Bi-Tf) and the ferritin heavy chain subunit (Bi-FerHCH), from the bumblebee Bombus ignitus. Bi-Tf cDNA spans 2,340 bp and encodes a protein of 706 amino acids and Bi-FerHCH cDNA spans 1,393 bp and encodes a protein of 217 amino acids. Comparative analysis revealed that Bi-Tf appears to have residues comprising iron-binding sites in the N-terminal lobe, and Bi-FerHCH contains a 5’UTR iron-responsive element and seven conserved amino acid residues associated with a ferroxidase center. The Bi-Tf and Bi-FerHCH cDNAs were expressed as 79 kDa and 27 kDa polypeptides, respectively, in baculovirus-infected insect Sf9 cells. Northern blot analysis revealed that Bi-Tf exhibits fat body-specific expression and Bi-FerHCH shows ubiquitous expression. The expression profiles of the Bi-Tf and Bi-FerHCH in the fat body of B. ignitus worker bees revealed that Bi-Tf and Bi-FerHCH are differentially induced in a time-dependent manner in a single insect by wounding, bacterial challenge, and iron overload.

Potato (Solamum tuberosum 'Dejima') plantlets were investigated on culture type and initial quantity of inoculation in bioreactor and survival rate by hydroponics for mass production. rode stems (1 to 1.5cm in length) of potato plantlets multiplied in vitro were grown for 3 weeks in liquid Murashige and Skoog (MS) medium with sucrose 30 g L-1. When plantlets (80-node inoculation) were raised in 10L balloon type bubble (BB) bioreactor, the healthiest growth of plantlets was obtained from explants cultured in ebb & flow culture with medium supplied periodically 12 times per day. The suitable inoculation quantity of 20L BB bioreactor was 120 pieces of stem segments (mean 2.2g fresh weight) in ebb & flow culture. Number of nodal shoot was eight on the average. In controlled culture room, survival rate of plantlets at 7 days after stem cutting was above 70% when they were acclimatized by hydroponics grown in deep flow and solid medium culture. The highest survival rate of the stem cutting plantlets was in nutrient solution adjusted to EC 1.4dS·m-1. Stem cutting plantlets through one culture could be obtained 670~900, when plantlets were grown in ebb & flow culture during 3 weeks using a 20L bioreactor with initial 120 pieces of nodal segments. 11 is possible In do mass production of seedlings cultured in bioreactor and hydroponics.

Safflower (Carthamus tinctorius L.) seeds have long been clinically used in Korea to promote bone formation and prevent osteoporosis. In addition, the safflower buds (SB) were found to have more useful functional ingredients than safflower seed. Thus, we investigated the preventive effects of SB diet in ovariectomized (OVX) rats. The rats were divided into five groups; sham operated group, OVX alone group, OVX plus 17β-estradiol (E2 10 ㎍/㎏, i.p.) and OVX plus SB diet feeding group (0.3% or 1%). Feeding of SB diet (0.3% or 3%) to OVX rats markedly increased trabecular formation in femur compared to OVX rats. Feeding of SB diet (0.3% or 3%) to OVX rats also decreased TRAP activity compared to OVX rats. These results suggest that SB diets have bone sparing effects by the decrease of osteoclast activity. We also observed that OVX rats fed with SB diet (0.3% or 3%) exhibited the decrease of calcium and phosphorus in serum compared to OVX-induced rats. Therefore, SB may be beneficial for the patients of osteoporosis, especially in postmenopausal women.

The vascular system of plants consists of two conducting tissues, xylem and phloem, which differentiate from procambium cells. Xylem serves as a transporting system for water and signaling molecules and is formed by sequential developmental processes, including cell division/expansion, secondary cell wall deposition, vacuole collapse, and programmed cell death (PCD). PCD during xylem differentiation is accomplished by degradation of cytoplasmic constituents, and it is required for the formation of hollow vessels, known as tracheary elements (TEs). Our recent study revealed that the small GTPase RabG3b acts as a regulator of TE differentiation through its autophagic activation. By using an Arabidopsis in vitro cell culture system, we showed that autophagy is activated during TE differentiation. Overexpression of a constitutively active RabG3b (RabG3bCA) significantly enhances both autophagy and TE differentiation, which are consistently suppressed in transgenic plants overexpressing a dominant negative form (RabG3bDN) or RabG3bRNAi (RabG3bRNAi), a brassinosteroidinsensitive mutant bri1-301, and an autophagy mutant atg5-1. Wood (called secondary xylem) is the most abundant biomass produced by land plants including Populus and Eucalyptus, and therefore is considered to be one of the most cost-effective and renewable bioenergy resources. In an attempt to enhance xylem differentiation and thus to improve biomass traits in poplars, we generated transgenic poplars overexpressing the RabG3bCA form. As notable phenotypes, both stem height and diameter were increased and xylem area in vascular bundles was significantly expanded in RabG3bCA transgenic poplars compared to control plants. Taken together, these results demonstrate that RabG3b regulates xylem differentiation in both Arabidopsis and Populus. This study enhances our understanding of biological mechanisms underlying wood formation and serve as a framework to engineer the quality and quantity of wood as useful biomass.