A New Member of the TBC1D15 Family from Chiloscyllium plagiosum: Rab GTPase-Activating Protein Based on Rab7 as a Substrate

APSL (active peptide from shark liver) is a hepatic stimulator cytokine from the liver of Chiloscyllium. It can effectively protect islet cells and improve complications in mice with alloxan-induced diabetes. Here, we demonstrate that the APSL sequence is present in the N-terminus of novel TBC (Tre-2, Bub2 and Cdc16) domain family, member 15 (TBC1D15) from Chiloscyllium plagiosum. This shark TBC1D15 gene, which contains an ORF of 2088 bp, was identified from a cDNA library of regenerating shark liver. Bioinformatic analysis showed that the gene is highly homologous to TBC1D15 genes from other species. Moreover, the N-terminus of shark TBC1D15 contains a motif of unknown function (DUF3548), which encompasses the APSL fragment. Rab-GAP activity analysis showed that shark TBC1D15 is a new member of the TBC1D15 family. These results demonstrated that shark TBC1D15 possesses Rab-GAP activity using Rab7 as a substrate, which is a common property of the TBC1D15 family. The involvement of APSL at the N-terminus of TBC1D15 also demonstrates that this protein might be involved in insulin signaling and may be associated with the development of type 2 diabetes. The current findings pave the way for further functional and clinical studies of these proteins from marine sources.     

Characterisation and physical mapping of cyclophilin A genes and identification of new classes of cyclophilins in wheat

The cyclophilins, a class of ‘foldase’ enzymes that catalyse the rate-limiting step of peptidyl-prolyl cis–trans isomerization, are up-regulated during wheat endosperm development and suggested to be important for folding of the storage proteins, thus influencing wheat quality. However, little information exists on the types of cyclophilins expressed, the genes encoding these, and their possible functions in wheat endosperm. We have characterised three isoforms of genes encoding cyclophilin A from eight wheat cultivars, using previously isolated cDNAs. The genes are small, intronless, comprise a small multi-gene family, lack any inter-cultivar polymorphisms and localise to chromosomal arms 6AS, 6BS and 6DS, in a region where genes for other quality traits are localised, the locus at 6AS possibly having duplicated genes. Further, cDNAs encoding two novel, endosperm-expressed classes of cyclophilins have been isolated, one being potentially plastid-localised and the other, a nuclear protein with cyclophilin-like domains. In-silico analyses have further led to identification of another form of cyclophilin A and a potentially ER-localised, endosperm-expressed cyclophilin B. The plastid and ER-localised forms are of particular relevance to events occurring during endosperm maturation. The results thus provide valuable data and molecular tools for isolation and analysis of these genes, to address their roles and any association with wheat quality traits.     

PPR蛋白在玉米子粒发育中的研究进展

PPR蛋白在真核生物中广泛存在，是最大的蛋白家族之一。该蛋白由一系列重复单位(形成两个反向平行的α螺旋)串联而成。PPR蛋白可与RNA特异性结合，参与细胞器RNA的加工和编辑。玉米中有约521个PPR蛋白成员，他们在子粒发育、叶片发育及胁迫响应等生物学过程中发挥重要作用，但多数分子功能尚未被鉴定。另外，PPR蛋白编码基因的突变体大多导致发育异常表型，如发育迟缓、子粒缺陷、叶色异常等。     

Identifying and Characterizing a Novel Peritrophic Matrix Protein (MdPM-17) Associated With Antibacterial Response From the Housefly,Musca domestica (Diptera: Muscidae)

Peritrophic matrix/membrane (PM) critically prevents the midgut of insects from external invasion by microbes. The proteins in the peritrophic membrane are its major structural components. Additionally, they determine the formation and function of this membrane. However, the role of PM proteins in immune regulation is unclear. Herein, we isolated a novel PM protein (MdPM-17) from Musca domestica larvae. Further, the function of MdPM-17 in regulating host innate immunity was identified. Results showed that the cDNA of MdPM-17 full is 635 bp in length. Moreover, it consists of a 477-bp open reading frame encoding 158 amino acid residues. These amino acid residues are composed of two Chitin-binding type-2 domain (ChtBD2) and 19 amino acids as a signal peptide. Moreover, tissue distribution analysis indicates that MdPM-17 was enriched expressed in midgut, and moderate levels in the fat body, foregut, and malpighian tubule. Notably, MdPM-17 recombinant protein showed high chitin-binding capacity, thus belongs to the Class III PM protein group. MdPM-17 protein silencing via RNA interference resulted in the expression of antimicrobial peptide (defensin, cecropins, and diptericin) genes, and this occurred after oral inoculation with exogenous microbes Escherichia coli (Enterobacteriales:Enterobacteriaceae), Staphylococcus aureus (Bacillales:Staphylococcaceae), and Candida albicans (Endomycetales:Saccharomycetaceae)). Therefore, all the antimicrobial peptide (AMP) gene expression levels are high in MdPM-17-depleted larvae during microbial infection compared to controls. Consequently, these findings indicate that MdPM-17 protein is associated with the antibacterial response from the housefly.     

Albumin and Globulin Proteins of Wheat Flour: Immunological and N-terminal Sequence Characterisation

Several non-storage proteins are encoded by genes on individual wheat chromosomes and these have been used as genome-specific markers. In this study, a library of monoclonal antibodies with differing specificities to water- and salt-soluble proteins has been developed in order to obtain markers for different wheat chromosomes. Two antibodies, BCSAU 9D1 and JGM 1B4 were found to bind polypeptides encoded by chromosomes 3D and 4D respectively. Other antibodies bound to small numbers of polypeptides encoded by more than one chromosome, such as 5A, 7D, 5B and 5D. The proteins recognised by some of the antibodies were isolated by immunoaffinity chromatography, and one protein was identified as an alpha -amylase inhibitor by N-terminal sequence characterisation. In addition, 16 water-soluble and eight salt-soluble proteins were isolated using SDS-PAGE, isoelectric focusing, two-dimensional electrophoresis and reversed-phase high performance liquid chromatography, with the main objective being to confirm the identity of particular proteins, especially those which were able to be assigned to particular chromosomes. The N-terminal sequencing characterisation identified 19 proteins, whereas four proteins were found to be blocked and one did not match with any proteins in the database. Most of the water-soluble proteins belonged to a family of alpha -amylase inhibitors. One protein, assigned to chromosome 4BS, was homologous to serine carboxypeptidase III. N-terminal sequences of some of salt-soluble proteins matched with internal sequences of barley embryo globulin. Other proteins were identified as lipid transfer protein, peroxidase BP-1 precursor and histone H4 proteins. The protein sequences could also potentially be used for making antibody or DNA probes for use in selection in breeding programmes.     

甘蔗线条花叶病毒RNA沉默抑制子的寄主互作蛋白鉴定

甘蔗线条花叶病毒（Sugarcane streak mosaic virus, SCSMV）是引起甘蔗花叶病的一种主要病原。SCSMV编码的P1蛋白是RNA沉默抑制子,在SCSMV抑制寄主的RNA沉默防御中发挥关键作用,但其作用机制尚未清楚。与寄主蛋白互作是RNA沉默抑制子发挥其抑制作用的主要途径之一,因此鉴定与病毒RNA沉默抑制子互作的寄主蛋白是研究抑制子作用机制的一个重要方法。为探究SCSMV P1抑制寄主RNA沉默的机制,本研究首先将SCSMV P1基因连接到质粒pGBKT7上构建诱饵质粒pGBKT7-P1,然后对pGBKT7-P1进行毒性和自激活检测,最后以pGBKT7-P1为诱饵,采用酵母双杂交技术从甘蔗cDNA文库中筛选与P1互作的寄主蛋白。结果显示,成功构建pGBKT7-P1诱饵质粒。将pGBKT7-P1诱饵质粒转入Y2H Gold酵母菌株后,酵母菌株在SD/-Trp平板及液体培养基中生长良好,表明pGBKT7-P1诱饵质粒对Y2H Gold酵母菌株无毒性。含pGBKT7-P1诱饵质粒的酵母菌株在SD/-Trp/X-α-Gal平板上长出白色菌落未变蓝,在SD/-Trp/X-α-Gal/AbA和SD/-Trp/-Leu/X-α-Gal/AbA平板上无菌落生长,表明pGBKT7-P1诱饵质粒无自激活活性。用pGBKT7-P1诱饵质粒对甘蔗cDNA文库进行筛选,经SD/-Trp/-Leu/X-α-Gal/AbA平板筛选1次及SD/-Trp/-Leu/-His/-Ade/X-α-Gal/AbA平板筛选3次后,获得42个阳性酵母克隆,提取阳性酵母质粒并导入大肠杆菌中扩大培养,经测序及blastx比对分析,获得13个可能与SCSMV P1互作的寄主甘蔗蛋白,分别是生长素应答蛋白IAA1、IAA15,转录因子NAC、GATA4、OFP4,真核翻译起始因子eIF5A,分子伴侣DnaJ,辅分子伴侣SBA1,重金属相关异戊二烯化植物蛋白HIPP35,mec-8和unc-52蛋白同系物抑制子SMU2,外膜孔蛋白OEP24,及2个未表征蛋白。基于这些蛋白的功能,推测P1可能通过与寄主蛋白互作来调控寄主防御反应相关基因的表达,和/或通过与寄主蛋白互作来影响寄主RNA沉默相关蛋白的合成、加工或转运,从而发挥其RNA沉默抑制子的功能。研究结果为后续深入解析P1抑制RNA沉默的作用机制奠定了重要基础。     

Biomedical and Clinical Importance of Mussel-Inspired Polymers and Materials

The substance secreted by mussels, also known as nature’s glue, is a type of liquid protein that hardens rapidly into a solid water-resistant adhesive material. While in seawater or saline conditions, mussels can adhere to all types of surfaces, sustaining its bonds via mussel adhesive proteins (MAPs), a group of proteins containing 3,4-dihydroxyphenylalanine (DOPA) and catecholic amino acid. Several aspects of this adhesion process have inspired the development of various types of synthetic materials for biomedical applications. Further, there is an urgent need to utilize biologically inspired strategies to develop new biocompatible materials for medical applications. Consequently, many researchers have recently reported bio-inspired techniques and materials that show results similar to or better than those shown by MAPs for a range of medical applications. However, the susceptibility to oxidation of 3,4-dihydroxyphenylalanine poses major challenges with regard to the practical translation of mussel adhesion. In this review, various strategies are discussed to provide an option for DOPA/metal ion chelation and to compensate for the limitations imposed by facile 3,4-dihydroxyphenylalanine autoxidation. We discuss the anti-proliferative, anti-inflammatory, anti-microbial activity, and adhesive behaviors of mussel bio-products and mussel-inspired materials (MIMs) that make them attractive for synthetic adaptation. The development of biologically inspired adhesive interfaces, bioactive mussel products, MIMs, and arising areas of research leading to biomedical applications are considered in this review.     

Molecular Basis and Regulation of Ammonium Transporter in Rice

Rice grows in flooded paddy fields and takes up ammonium as the preferred nitrogen (N) source. Ammonium uptake is facilitated by a family of integral membrane proteins known as ammonium transporters found in all domains of life. However, the molecular mechanism and functional characteristics of the ammonium transporters (AMT) in rice have not been determined in detail yet. In this review, we report a genome-wide search for AMT genes in rice, resulting in the increase of the number of potential AMT proteins to at least 12, including members of both the alpha and beta sub-groups. Analysis of the predicted protein sequences for the 12 OsAMT proteins identified many conserved phosphorylation sites in both the alpha and beta group members, which could potentially play a role in controlling the activity of the transporters. Present knowledge of the expression of rice AMT genes is also summarized in detail. Future studies should focus on the structural and functional characteristics of OsAMT proteins to provide insight into the mechanism of ammonium uptake and its regulation in rice. Such research could improve utilization and decrease wastage of N fertilizer in rice cultivation.     

Systematic Identification of Rice ABC1 Gene Family and Its Response to Abiotic Stress

Members of the activity of bc1 complex (ABC1) family are protein kinases that are widely found in prokaryotes and eukaryotes. Previous studies showed that several plant ABC1 genes participated in the abiotic stress response. Here, we present the systematic identification of rice and Arabidopsis ABC1 genes and the expression analysis of rice ABC1 genes. A total of 15 and 17 ABC1 genes from the rice and Arabidopsis genomes, respectively, were identified using a bioinformatics approach. Phylogenetic analyses o...     

Variant high-molecular-weight glutenin subunits arising from biolistic transformation of wheat

Genetic transformation via the biolistic method has been used to introduce genes encoding natural and novel high-molecular-weight glutenin subunits (HMW-GS) into wheat. The appearance of new seed proteins of sizes not predicted by the transgene coding sequences was noted in some experiments. In this report, the identities of thirteen of these novel proteins were determined by tandem mass spectrometry (MS/MS). Seven different proteins larger than and two proteins smaller than the native protein were shown to contain peptides from 1Dx5. A novel protein found in some progeny of crosses between a transgenic plant and Great Plains winter wheats was larger than but contained several peptides from 1Dy10. In one line, a protein larger than and a protein smaller than HMW-GS each contained peptides from the N- and C-terminus of 1Dx5 and from the repeat region of 1Dy10. In a sixth transgenic line, the native Bx7 gene was apparently replaced by a gene that encodes a larger version of 1Bx7. The variant proteins accumulate in the polymeric protein fraction, indicating that they can form inter-molecular disulfide bonds. These results show that novel proteins found in some transformants are encoded by altered versions of either the transforming or endogenous HMW-GS genes.     

龙眼体胚发生过程中RNA甲基化相关基因全基因组鉴定及表达分析

为了解龙眼RNA甲基化相关基因的生物学功能,本研究基于基因组和转录组数据,采用生物信息学分析方法,进行龙眼基因组RNA甲基化相关基因成员鉴定,蛋白结构域及特性分析,分子进化树分析,体胚发生过程和组织器官基因表达水平分析以及ABA处理下的定量表达分析。结果显示：龙眼中有7个参与RNA甲基化过程中的关键基因,包括5个RNA甲基转移酶基因和2个去RNA甲基化酶基因;各成员内含子数量差别较大,为4~28个不等;进化树分析显示,龙眼RNA甲基转移酶和去甲基化酶蛋白与甜橙亲缘关系最近;蛋白结构域分析显示,RNA甲基化相关酶具有保守的蛋白结构域;龙眼RNA甲基化相关基因启动子区域中主要含有光响应功能元件、激素响应功能元件、胁迫响应功能元件和分生组织表达响应元件,推测RNA甲基化相关基因可能对植物的生长发育和适应不良环境有一定调节作用;RNA甲基化相关基因部分成员在龙眼体胚不同发生阶段和不同组织器官中的表达有明显区别,推测龙眼RNA甲基化相关基因可能参与不同体胚发育阶段和组织器官形态建成;不同浓度ABA处理下,龙眼EC中RNA甲基化相关基因的表达情况各异,其中DlVIR、DlALKBH10B和DlMTB的相对表达量受到一定的抑制作用,而DlFIP37则呈现上调表达,暗示龙眼RNA甲基化相关基因参与激素响应途径。本研究表明,龙眼RNA甲基化相关基因除了可能在叶的形成、花的发育和植株的形态建成等方面发挥重要的作用外,还可能参与调控胚胎发育和响应非生物胁迫,推测龙眼RNA甲基化相关基因成员在功能上具有差异性和多样性。     

Phenotypic and allelic variation for wort protein Z in Australian and Canadian barleys

Protein Z is a major component in beer foam. Two-dimensional electrophoresis was used to analyze wort proteins of two Australian (Buloke and Commander) and two Canadian (CDC Meredith and Bentley) varieties. The Canadian barley contained more abundant proteins from MW 40–45 kDa (pI 5 to 7). These proteins were identified as either protein Z4 or protein Z7 using liquid chromatography–mass spectrometry. Full-length gene of protein Z4 and Z7 were sequenced from Canadian and Australian barleys. Sequence differences were identified in the coding region and upstream regions of the two genes, resulting in protein sequence and expression variations. Molecular markers were designed according to the indels in the upstream regions of protein Z4 and Z7 genes. These markers were highly correlated to wort protein Z content in Canadian and Australian varieties. The Canadian barleys contained ‘high level’ genotypes for protein Z4 and Z7 while most Australian barleys had ‘low level’ genotypes for protein Z4, Z7 or both. The markers identified in this study provide a valuable tool for the selection of protein Z alleles in marker-assisted breeding. Total protein Z content was assessed using different steeping conditions, and increasing air-rest time increased protein Z content in 15 varieties.     

Constitutive expression of <Emphasis Type="Italic">REL1</Emphasis> confers the rice response to drought stress and abscisic acid

Leaf rolling is one of the most significant symptoms of drought stress in plant. Previously, we identified a dominant negative mutant, termed rolled and erect 1 (hereafter referred to rel1-D), regulating leaf rolling and erectness in rice. However, the role of REL1 in drought response is still poorly understood. Here, our results indicated that rel1-D displayed higher tolerance to drought relative to wild type, and the activity of superoxide dismutase (SOD) and drought responsive genes were significantly up-regulated in rel1-D. Moreover, our results revealed that rel1-D was hypersensitive to ABA and the expression of ABA associated genes was significantly increased in rel1-D, suggesting that REL1 likely coordinates ABA to regulate drought response. Using the RNA-seq approach, we identified a large group of differentially expressed genes that regulate stimuli and stresses response. Consistently, we also found that constitutive expression of REL1 alters the expression of biotic and abiotic stress responsive genes by the isobaric tags for relative and absolute quantification (iTRAQ) analysis. Integrative analysis demonstrated that 8 genes/proteins identified by both RNA-seq and iTRAQ would be the potential targets in term of the REL1-mediated leaf morphology. Together, we proposed that leaf rolling and drought tolerance of rel1-D under normal condition might be caused by the endogenously perturbed homeostasis derived from continuous stressful dynamics.     

龙眼miRNA合成途径相关基因的鉴定及表达分析

为了解龙眼miRNA合成途径相关基因的生物学功能,本研究依据龙眼第3代基因组数据对龙眼miRNA合成途径相关基因进行了鉴定和生物信息学分析,并利用龙眼转录组对其在体胚发生早期定量表达进行分析.研究结果如下:共鉴定到12条龙眼miRNA合成途径相关基因,分别是DlDDL1、DlDDL2、DlDDL3、DlDCL1、DlS...     

The Complete Mitochondrial Genome of Brachmia macroscopa (Lepidoptera: Gelechiidae) and Its Related Phylogenetic Analysis

The sweet potato leaf folder, Brachmia macroscopa, is an important pest in China. The complete mitogenome, which consists of 13 protein-coding genes (PCGs), 22 transfer RNA genes, two ribosomal RNA genes, and an A + T-rich region, was sequenced and found to be 15,394 bp in length (GeneBank no. KT354968). The gene order and orientation of the B. macroscopa mitogenome were similar to those of other sequenced lepidopteran species. All of the PCGs started with ATN as the canonical start codon except for cox1, which started with CGA. In regard to stop codons, most PCGs stopped at TAA except for cox2, which stopped at TA, and nad4, which stopped at a single T. Thirteen PCGs of the available species (33 taxa) were used to demonstrate phylogenetic relationships. The ditrysian cluster was supported as a monophyletic clade at high levels by using maximum likelihood and Bayesian methods. The apoditrysian group, covering the Gelechioidea, formed a monophyletic clade with a bootstrap value of 88% and a posterior probability of 1.00. The superfamily Gelechioidea was supported as a monophyletic lineage by a posterior probability of 1.00.     

Variation in kernel characteristics and protein molecular weight distribution of Langdon durum-wild emmer wheat chromosome substitution lines

Triticum turgidum L. var. dicoccoides (DIC) provides a useful source of genes to improve agronomic and quality characteristics of durum wheat. Research was performed to identify DIC chromosomes that carry useful genes for quality improvement. Langdon-T. dicoccoides substitution lines consisting of 13 lines based on DIC accession PI 481521, 10 lines on PI 478742, and 2 lines on Israel A were evaluated for kernel characteristics and protein molecular weight distribution. DIC chromosomes 3A from PI 481521, and 1A and 7A from PI 478742 increased kernel hardness. Chromosome 2A from PI 481521 increased kernel weight, which resulted in increased semolina yield. Some substituted DIC chromosomes also affected grain protein concentration and protein molecular weight distribution. For example, chromosomes 2A, 5B, and 7B from PI 481521, and 6B from Israel A increased total protein concentration, which was primarily attributed to an increase of SDS soluble gliadins. Chromosome 6B from PI 478742 was unique in that it led to an increase in SDS insoluble high molecular weight polymeric proteins, which contributed to increased dough mixing strength. Results from this research indicated that chromosome 6B from PI 478742 is a potential gene source to improve dough characteristics of durum wheat by increasing insoluble high molecular weight polymeric protein concentration.     

Fish Synucleins: An Update

Synucleins (syns) are a family of proteins involved in several human neurodegenerative diseases and tumors. Since the first syn discovery in the brain of the electric ray Torpedo californica, members of the same family have been identified in all vertebrates and comparative studies have indicated that syn proteins are evolutionary conserved. No counterparts of syns were found in invertebrates suggesting that they are vertebrate-specific proteins. Molecular studies showed that the number of syn members varies among vertebrates. Three genes encode for α-, β- and γ-syn in mammals and birds. However, a variable number of syn genes and encoded proteins is expressed or predicted in fish depending on the species. Among biologically verified sequences, four syn genes were identified in fugu, encoding for α, β and two γ (γ1 and γ2) isoforms, whereas only three genes are expressed in zebrafish, which lacks α-syn gene. The list of “non verified” sequences is much longer and is often found in sequence databases. In this review we provide an overview of published papers and known syn sequences in agnathans and fish that are likely to impact future studies in this field. Indeed, fish models may play a key role in elucidating some of the molecular mechanisms involved in physiological and pathological functions of syn proteins.     

Analysis of Differential Proteins in Two Wing-Type Females of Sogatella furcifera (Hemiptera: Delphacidae)

Sogatella furcifera (Horvath) is an important rice pest with the wing dimorphism, including macropterous and brachypterous morphs. The protein expression profiles in two wing-type adults and two wing-type disc fifth-instar nymphs were analyzed using two-dimensional gel protein electrophoresis and mass spectrometry. In adults and fifth-instar nymphs, 127 and 162 protein spots were detected, respectively. Fifty-five differentially expressed protein spots were identified between the long-winged adults and the short-winged adults, and 62 differentially expressed protein spots were found between the long-winged disc fifth-instar nymphs and short-winged disc fifth-instar nymphs. In long-winged and short-winged adults, six and seven specific protein spots were identified, respectively, with five and seven protein spots having more than threefold increased level, respectively. In long-winged and short-winged disc morph nymphs, 8 and 12 specific protein spots were identified, respectively, with 11 and 17 spots containing more than threefold increased level, respectively. Among the 16 identified proteins, five proteins are associated with muscle function, suggesting that muscle is a main tissue where the genes were differentially expressed between the two wing types. In addition, the content of a peptidase with an insulinase domain was higher (by 3.02 ± 0.59 fold) in the short-winged fifth-instar nymphs than in the long-winged fifth-instar nymphs, which suggests that this peptidase may be involved in wing differentiation by regulating insulin receptors. The results of this study provide some genetic clues for the wing differential development in S. furcifera and provide more references for future studies.     

Analyses of albumins, globulins and amphiphilic proteins by proteomic approach give new insights on waxy wheat starch metabolism

Starch is composed of two types of glucose polymers: amylose and amylopectin. The Waxy (Wx) locus controls amylose synthesis in the wheat kernel. Hexaploid wheat has three Wx loci located on chromosomes 7A (Wx-A1), 4A (Wx-B1), and 7D (Wx-D1). Eight near isogenic lines (NILs) of Triticum aestivum cv. Tremie with one, two or three Wx null alleles were used. The albumin–globulin fraction, and amphiphilic proteins were separated using 2-dimensional electrophoresis (2DE) allowing the changes in the waxy kernel to be identified. Albumin–globulin fraction showed overexpression of sucrose synthases in the waxy NILs compared to the normal form of Tremie and a decrease in many proteins related to stress and defence metabolism such as serpins. A subunit of ADP-glucose pyrophosphorylase (AGPase), which is known to play a major role in starch synthesis, was also shown to be down regulated in the waxy NILs. Amphiphilic proteins confirmed the observations made on the albumin–globulin fraction with a decrease in a stress-related protein. These different regulations linked to observations made on wheat kernel (thousand kernel weight (TKW), protein amount per grain, size and distribution of the starch granules) led to formulation of the hypothesis that waxy endosperm does not reach maturity of the wild-type endosperm.