G protein-coupled receptors (GPCRs) belong to cell membrane protein family, which regulate various physiological process such as reproduction, behavior and immune etc. In other to identify the GPCRs in pheromone gland of Maruca vitrata, we carried out transcriptome analysis from both females. Transcriptome analysis in the pheromone gland yielded approximately 22Gb and 47,528 transcripts showed positive FPKM value. 48 Genes involved in GPCRs were identified such as pheromone biosynthesis activating neuropeptide receptor (PBANr), prostaglandin receptors, neuropeptide receptor, 5-hydroxytryptamine receptor, galanin receptor, calcitonin gene-related peptide receptor, diuretic hormone receptor, gonadotropin-releasing hormone receptor, frizzled and orphan receptors, etc. Various expression of GPCRs in the pheromone gland indicates the role of pheromone gland may not be limited to the production of pheromone.

The native human saliva obtained through the centrifugation of whole saliva showed characteristic salivary protein complex (SPC) peaks in gel filtration high performance liquid chromatography (HPLC) using Superose 12 column1,2). In the previous study the SPC peaks in chromatography were explored to know their composition and functions by different detection methods, but still the nature of SPCs was not clearly elucidated so far. In this study the SPC peaks were examined by direct antibody interaction in order to target different antimicrobial and protective proteins distributed in the SPCs via gel filtration HPLC. As the SPC peak shape and migration speed can be changed by antibody binding to specific proteins of SPC, it was found that mucin1 is evenly distribution in all SPCs, while PRPs are more abundant in the late dominant SPC than the early dominant SPC and also in the intermediated SPCs. Most of antimicrobial proteins including lysozyme, LL-37, lactoferrin, β-defensin-1, -2, -3, IgA, mucocidin, and α1-antitrypsin were more abundant in the late dominant SPC than the early dominant SPC, while histatin showed relatively even distribution in all SPCs. Therefore, it was presumed that the late dominant SPC containing abundant antimicrobial and protective proteins could be applied as a biomarker to measure the defensive potential of whole saliva in oral diseases.

Glucose-regulated protein 78 (GRP78) is a member of the heat shock protein 70 (HSP70) family that is specific to endoplasmic reticulum (ER). It is known as chaperones and signaling regulators that respond to ER stresses in vertebrates. However, its function in invertebrates, including insects, is uncertain. Here we determined a full cDNA sequence and the expression patterns of grp78 of Aphis gossypii, which is a major pest of numerous crop plants worldwide. Its cDNA had highly conserved motifs of the HSP70 family and the C-terminal motif of KDEL characteristic of ER-specific HSPs. It showed 98.5% identity with the GRP78 of the pea aphid Acyrthosiphon pisum. Real-time RT-PCR analysis showed that the grp78 level was higher in the fourth instar nymph than in the younger instar and adult stages. Its level was not affected by thermal stress of 10 to 40°C for 1 h. The grp78 level was proportional to the ingestion of a sucrose solution ranging in concentration from 0% to 30% in a Parafilm feeding chamber. In addition, the grp78 level varied among aphids feeding on leaves from 14 different host plants for 24 h; it was higher with eggplant and pepper but lower with pigweed and tobacco than any other plants. Our study suggests that the grp78 level is regulated by nutritional condition of A. gossypii.

Cotton aphid infests more than 700 plants and a major pest of various horticultural crops worldwide. The glucose regulated protein 78 (GRP78) is a member of heat shock protein 70. Its expression is associated with the nutritional changes as well as environmental stresses. The full sequences of grp78 cDNA of Aphis gossypii was determined. It had conserved motifs of hsp genes and terminated in KDEL which is common to GRP78. Quantitative realtime PCR showed that its level was changed during development and also upregulated by starvation. However, its level was not much changed by heat stress. The level of grp78 can be use to understand nutritional physiology on insects.

Interferon induced transmembrane protein-1 (IFITM1) is one of transmembrane protein which is differentially expressed in uterus during estrus cycle and pregnancy, that IFITM1 gene is highly expressed in estrus stage by the effect of estrogen, and in parturition by the effect of PGF2 alpha. This genes are also up-regulated in cells with hyperactivation of the WNT/β-catenin signaling pathway. In this study, to identify a function of IFITM1, the binding partner of IFITM1 were determined using immunoprecipitation and LC- MASSMASS methods. 1, 3 and 5 ug of polyclonal anti-IFITM1 antisera were used for immunoprecipitation, and the 75 kDa of specific band was detected in silver stained polyacylamide gel. This band were chracterized using LC-MASS-MASS, and revealed this band is glucose regulated protein 75 (GRP75) which binds to p53 and inhibits the p53 action in nucleus. To identify the localization of GRP75 in cells, immunocytochemical approach has been applied, and GRP75 is expressed in mitochondria of L929 murine connective tissue cells. Co-localization study between IFITM1 and GRP75 in L929 cell identified that these two proteins were closely expressed in mitochondria. Although the role of the interaction of these two protein need to be clarified in various biological phenomena, this data suggest that close interaction of IFITM1 and GRP75 may regulate cellular functions in uterus on sets of estrus cycle and pregnancy.

Insulin/insulin-like peptide-binding protein (IBP) is abundantly found in venom of the solitary hunting wasp, Eumenes pomiformis (Hymenoptera: Eumenidae). E. pomiformis IBP (EpIBP) is most similar to insect IBP-like proteins that are known to inhibit insect growth and insulin signaling. To investigate the toxicity and target protein, EpIBP was in vivo expressed by Escherichia coli. Spodoptera exigua (Lepidoptera: Noctuidae) larvae injected with EpIBP showed a 20% lower pupation rate than the control larvae, although their body weight was not significantly different from the control when the larvae were provided artificial diet after the injection. EpIBP extended the larval stage without inducing paralysis of S. exigua larvae. To investigate the effects of EpIBP on caterpillar under a starvation condition, survivorship and body weight of the EpIBP-injected were evaluated without providing artificial diet until all the larvae died. The survivorship of the EpIBP-injected larvae was 24-36% higher than the control larvae at 4-5 d post-injection. The body weight of the control larvae reduced to 59% that is approximately 10% lower than the body weight of the EpIBP-injected larvae. These results suggest that EpIBP might inhibit the metabolism of the caterpillars, which is likely related with insulin-like peptide signaling pathway, suppress the loss of body weight and eventually extend the larval stage. An EpIBP-binding protein (EpIBPBP) isolated by immunoprecipitation was matched with a coiled-coil domain-containing protein of the fruit fly. The full-length sequence analysis of EpIBPBP is in progress.

The hemipteran whitefly Bemisia tabaci (Gennadius) is one of the most destructive pests damaging more than 600 agricultural crop species worldwide. The B and Q biotypes are most widely spread in Korea but they are not distinguishable based on morphological characters. In order to search for protein markers that can be employed for rapid and accurate diagnosis of biotypes, two-dimensional PAGE (2DE) in conjunction with mass spectroscopic analysis were conducted. Eleven biotype-specific spots were repeatedly identified during three repetitions of 2DE and analyzed by Q-TOF. One of the B type-specific protein spots was identified as carboxylesterase 2 (Coe2). The transcript level of coe2 was determined to be 6 times higher in B type than in Q type by quantitative real-time PCR. In addition, comparison of genomic DNA sequence of coe2 between B and Q types identified a biotype-specific intron, from which specific primer sets were designed. One-step PCR using these biotype-specific primers successfully distinguished the two biotypes in a high accuracy. Availability of the biotype-specific protein and DNA markers will greatly improve the detection of B. tabaci biotype in the field.

Surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF MS) is one of the recently developed proteomic technologies which is based on capturing proteins and peptides by chemically modified surfaces and highly sensitive for the analysis of complex biological samples. In the present study, to gain insights into oocyte maturation and early embryo development, SELDI-TOF-MS was used to find the protein candidates that are specifically or prominently expressed in porcine oocytes at the in vitro matured metaphase II (MIIl) and germinal vesicle (GV) stages. By selected CM10 chip, 16 candidates were found to be up-regulated in GV stage oocytes compared with in MII stage oocytes, their molecular weights were 8,180 (2 candidates), 10,226 (5 candidates), 15,767 (5 candidates) and 16,770 (4 candidates) Da respectively. And the expression of 29 candidates were higher in MII than in GV stage oocytes, their molecular weight were 10,832 (3 candidates), 17,743 (8 candidates), 20,122 (3 candidates), 22,131 (3 candidates), 24,857 (7 candidates) and 33,507 (5 candidates) Da, respectively. The expression of selected 13 candidates (0.2 and 1.0 % error tolerances) were analyzed using real time RT-PCR. The proteins that differentially regulated during oocyte in vitro maturation in the pigs may be potential biomarkers of oocyte maturation and quality.

Gene reg비 at i o n during the human craniofacial development is not well understood In effort to understand n ewly identifï ed genes that may play role(s) in the human craniofacial development, non-redundan t genes were isolated from the s ubtracted cDNA libra ry of human embryonal craniofacial tissues and examined their possible structu ral rolc in parallcl with thosc gcncs from isolatecl human c h o nclroc)πes cDNA library. Fifty genes were init ia ll y chosen from 398 clones iso latecl were used for selective dominant expression in both chondrocytes and the craniofacial sections of 10 weeks old human embryo by in situ hybridization method. Based upon the high levels 。f expression, we have identifi ecl seven unknown genes; ch89, ch96. ch129. ch 153. ch 276 ch285. and ch334 . In 。rder to unde rs tancl the possi ble role of these genes‘ the structural simulation of the expressed proteins were constructecl by Sybyl 6.6 program. Ch 276 gene was same with a clone, c14 0 1' f173. registered in GenBank(NM_022489) a nd is composed 0 1' 323 amino acids having a reverse s ignaling domain from the extra- cellular matrix(C-terminal) to cell membrane(N-terminal) and 12 turns of helical structure. Gene protein also r etains a famil iar fïbronectin binding domain(RGD). three s ites 0 1' Ca ion binding motifs. cAMP- and cGMP-dep endent protein kinase phos phorylation site, two regions of protein kinase C phosphorylation s ites. glyco- saminoglycan attachment s ite ancl N-glycosylation site. transmembrane and Al kaline Phosphatase active s ite domains This newly iclentifï ed human protein from human choncl rocytes cDNA library appearecl to be related to a known calcification s ignaling protein. was named as Ca lsin(Ch276) . Ch153 appeared to be related a family of anti-microbial peptide acting as an inflammation mediator and Ch334 clone as a zinc finger protein whose expression in creases in human adult ti ssue‘ These results suggest that these novel genes ident i!ï ed from human chondrocytes rnay provide a new path 0 1' embryonic cartilage development and human craniofacial development.

Soybean [Glycine max (L.) Merr.] seeds are abundant in high-quality proteins and fats. In addition, soybean seeds are also rich in secondary metabolites, such as isoflavones, lecithin, and saponins. Triterpene saponins are major components of these physiologically active metabolites in soybean seeds. Soybean saponins are classified as group A and DDMP saponins. Among them group A saponins are undesirable component of food products due to bitterness and astringency and also cause foaming in tofu production. Whereas, DDMP saponins and their derivatives are less bitter and astringent and beneficial to human health when consumed as regular diet. Therefore, reducing the group A saponins or increasing the DDMP saponins are required to improve the food quality. The present study focused to identify and characterize the gene which is encoding a protein responsible for biosynthesis of DDMP saponins. EMS mutant lines (sg-7-1 & sg-7-2) which lack DDMP saponins were developed. The breeding cross has been made with these two mutants with two cultivars, Pungsannamul and Wooram to study the segregation and genetic linkage analysis, respectively. The segregation analysis showed that the mutant phenotype is controlled by single recessive gene. TLC analysis for phenotyping F2 population of Wooram X sg-7-1 showed mutant, wild and heterozygous types. To surprise two more patterns were detected and they were named as strange type1 (ST1) and strange type2 (ST2). Further, SSR marker analysis will be carried out to locate the gene which encoding a protein responsible for biosynthesis of DDMP saponins.

Although it is well known that low-molecular-weight glutenin subunits (LMW-GS) affects bread and noodle processing quality, the function of specific LMW-GS proteins mostly remain unclear. It is important to find a corresponding gene for a specific LMW-GS protein in order to understand the function of the specific LMW-GS protein. The objective of this study was to identify LMW-GS genes and haplotypes using well known Glu-A3, Glu-B3 and Glu-D3 gene specific primers and to interlink their protein products by proteomic approaches in a wheat variety. A total of 36 LMW-GS genes and pseudo-genes were amplified including 11 Glu-3 gene haplotypes, designated as GluA3-13K and GluA3-22K (pseudogene) at Glu-A3 loci, GluB3-33K and GluB3-43K at Glu-B3 loci and GluD3-11K, GluD3-21K, GluD3-31K, GluD3-42K, GluD3-5K, GluD3-6K and GluD3-393K (pseudogene) at Glu-D3 loci. To determine the relationship between gene haplotypes and their protein products (to identify the corresponding LMW-GS proteins), we conducted N-terminal amino acid sequencing and tandem mass spectrometry (MS/MS) analysis of the 17 LMW-GS spots separated by 2-DGE. Successfully, LMW-GS proteins of the Glu-3 gene haplotypes except pseudo-genes mentioned above were identified. This is the first report on comprehensive characterization of LMW-GS genes and their corresponding proteins and establishment of specific correspondence between each other in a single wheat cultivar. Our approach will be useful to understand the molecular basis of the LMW-GS and to study their contribution to the end-use quality of flour.

Sessile organism, plants constitutively challenged with pathogens have been developed various strategies for protection, such as preformed and inducible defense mechanisms. Receptor-like Proteins(RLPs) play critical roles in defense response as well as in plant development and growth. The domain structure of RLPs consists of extracellular leucine–rich repeats, a transmembrane domain, and a short cytoplasmic tail. Here, we identified putative 170 RLP genes from pepper genome using in-house bioinformatics pipeline. The distribution of RLPs on pepper pseudomolecule showed uneven spread and a number of RLPs were physically clustered by tandem array in the specific chromosome. Motifs analysis of pepper RLPs showed conserved LRR sequences (LxxLxxLDLxxNxxxGxIP). To understand further functional and evolutionary characteristics, evolutional relationship and gene profiling analysis are on progress.

Fibroin silk proteins from spider or silkworm are attractive biomaterials that are of particular biotechnological interest for industrial and medical purposes because of their unique physical and mechanical properties. In this study, we generated and characterized the transgenic rice plant expressing a spider silk protein. Spider silks have great potential as biomaterials with extraordinary properties. Here, we report the cloning and characterization of the major ampullate silk protein gene from the spider Araneus ventricosus. A cDNA encoding the partial major ampullate silk protein (AvMaSp) was cloned from A. ventricosus. An analysis of the cDNA sequence shows that AvMaSp consists of a 240 amino acid repetitive region and a 99 amino acid C-terminal non-repetitive domain. The peptide motifs that were found in the spider major ampullate silk proteins, (A)n, (GA)n, and (GGX)n, were conserved in the repetitive region of AvMaSp. Phylogenetic analysis further confirmed that AvMaSp belongs to the spider major ampullate spidroin family of proteins. Recombinant AvMaSp-R was degraded abruptly by trypsin. However, AvMaSp-R was stable at 100 °C for at least 30 min. Additionally, the AvMaSp-R was stable at pH values from 2 to 12 for at least 1 h. Taken together, our findings describe the molecular structure and biochemical properties of the A. ventricosus major ampullate silk protein and demonstrate its potential as a biomaterial.

In this study, we generated and characterized the transgenic rice plant expressing a spider silk protein. Spider silks have great potential as biomaterials with extraordinary properties. We report the cloning and characterization of the major ampullate silk protein gene from the spider Araneus ventricosus. A cDNA encoding the partial major ampullate silk protein (AvMaSp) was cloned from A. ventricosus. An analysis of the cDNA sequence shows that AvMaSp consists of a 240 amino acid repetitive region and a 99 amino acid C-terminal non-repetitive domain. The peptide motifs that were found in the spider major ampullate silk proteins, (A)n, (GA)n, and (GGX)n, were conserved in the repetitive region of AvMaSp. Phylogenetic analysis further confirmed that AvMaSp belongs to the spider major ampullate spidroin family of proteins. The AvMaSp-R cDNA, which encodes the 240 amino acid repetitive domain, was expressed as a soluble 22 kDa polypeptide in baculovirus-infected insect cells. To produce transgenic rice plant with high contents of glycine and alanine, the prolamin promoter-driven AvDrag was introduced into rice plant via agrobacterium tumefaciens-mediated gene transformation. The introduction and copy number of the AvDrag gene in transgenic rice plants were determined by PCR and Southern blot analysis. AvDrag expression in transgenic rice seeds was examined by Northern blot and Western blot analysis. Immunofluorescence staining with the AvDrag antiserum revealed that the recombinant AvDrag protein were localized in transgenic rice seed. Furthermore, the amino acid content analysis showed that transgenic rice seeds were greatly increased in glycine and alanine as compared to controls