Structured to reduce the mitogenicity of anti-CD3 antibody based on computer-guided molecular design

The mouse anti-human CD3 monoclonal antibody such as OKT3 is a potent immunosuppressive agent used in clinical transplantation to manipulate T-cell functions and prevent acute allograft rejection. However, the broad use of anti-CD3 antibody in clinical treatment was severely limited by the side effects of human anti-mouse antibody response and cytokine release syndrome. In this study, on the basis of a murine anti-human CD3 antibody yCD3 obtained in our previous work, a novel engineered anti-human CD3 antibody fragment (i.e. V(H)-Linker-V(L)-Hinge-CH(3)) was constructed with computer-guided molecular design method to avoid the clinical side effects. According to the distance geometry and intra-molecular interaction, the hinge region was re-designed and different from the parental hinge region in human IgG1. With the novel hinge region, the cysteine residues in hinge were exposure and prone to form the disulfide bond. Therefore, a novel bivalent antibody fragment named as mini-yCD3 was obtained. Mini-yCD3 displayed similar antigen-binding affinity and specificity to yCD3. Importantly, mini-yCD3 was shown to be much less potent in the induction of T-cell proliferation, cytokine release (interferon-gamma and interleukin-2) and early activation marker expression on the cell surface (CD69 and CD25) than parental yCD3. Furthermore, mini-yCD3 was effective in modulating T-cell receptor/CD3 and inhibiting mixed lymphocyte reaction with similarity as yCD3. In conclusion, the constructed mini-yCD3 was much less mitogenic to T cells but retained potent immunosuppression, suggesting it might be an alternative to yCD3 as an immunosuppressive drug with less immunogenicity and toxicity for clinical application.     

Expression of phytoene synthase gene (Psy) is enhanced during fruit ripening of Cara Cara navel orange (Citrus sinensis Osbeck)

Citrus is an important fruit crop as regards accumulation of carotenoids. In plant carotenoid biosynthesis, phytoene synthase gene (Psy) plays a key role in catalyzing the head-to-head condensation of geranylgeranyl diphosphate molecules to produce colorless phytoene. In the present paper, we reported the phytoene contents determination and characterization of Psy during fruit ripening of “Washington” navel orange and its red-fleshed mutant “Cara Cara”. Results showed that phytoene was exclusively accumulated in peel and pulp of “Cara Cara”. Although phytoene was observed accumulating with fruit ripening of “Cara Cara”, the contents in pulp were 10 times higher than those in peel. The isolated two Psy cDNAs were both 1520 bp in full length, containing 436 deduced amino acid residues, with a different amino acid at 412th. Genomic hybridization results showed that one or two copies might be present in “Cara Cara” and “Washington” genomes. During “Cara Cara” and “Washington” fruit coloration, expression of Psy was observed to be up-regulated, as revealed by tissue specific profiles in the flavedo, albedo, segment membrane and juice sacs. However, Psy expression in albedo of “Cara Cara” was higher than that in “Washington”, as evidenced by phytoene accumulation in the peel.     

Interleukin-17F signaling requires ubiquitination of interleukin-17 receptor via TRAF6

Interleukin-17F (IL-17F), together with interleukin-17A (IL-17 or IL-17A), is a marker of T(H)17 cells, a new lineage of effector CD4(+) T cells to contribute to pathogenesis of a growing list of autoimmune and inflammatory diseases, such as experimental autoimmune encephalitis (EAE) and collagen-induced arthritis (CIA). IL-17F, similar to IL-17A, was reported to employ interleukin-17 receptor (IL-17R or IL-17RA) for signaling but the downstream cascades remain largely elusive. Here we report that TRAF6 interacts with IL-17R and mediates ubiquitination of the receptor. We observed that IL-17F and IL-17A could induce IL-17R ubiquitination and DN-TRAF6, a dominant-negative mutant, could block IL-17F- but not IL-17A-triggered ubiquitination of IL-17R. Moreover, we showed that the ubiquitination of IL-17R was positively correlated with the downstream signaling, as evaluated by a luciferase reporter driven by a putative native promoter of 24p3, an IL-17 targeted gene. Our results indicate that ubiquitination of IL-17R mediated by TRAF6 plays a critical role in IL-17F signaling. This study, for the first time, reveals a possible molecular mechanism that the initiation of the IL-17F/IL-17R signaling pathway requires the receptor ubiquitination by TRAF6.     

Taurine promotes connective tissue growth factor (CTGF) expression in osteoblasts through the ERK signal pathway

Summary. Taurine is found in bone tissue, but its function in skeletal tissue is not fully understood. The present study was undertaken to investigate regulation of gene expression of connective tissue growth factor (CTGF), and the roles of mitogen-activated protein kinases (MAPKs) in murine osteoblast MC3T3-E1 cells treated with taurine. Western blot analysis showed taurine stimulated CTGF protein secretion in a dose- and time-dependent manner. Taurine induced activation of extracellular signal-regulated kinase (ERK), but not p38 and c-jun N-terminal Kinase (JNK), in osteoblasts. Furthermore, pretreatment of osteoblasts with the ERK inhibitor PD98059 abolished the taurine-induced CTGF production. These data indicate that taurine induces CTGF secretion in MC3T3-E1 cells mediated by the ERK pathway, and suggest that osteoblasts are direct targets of taurine.     

A comparative location database (CompLDB): map integration within and between species

We have adapted the Location Database (LDB) map-integration strategy of Morton et al. [Ann Hum Genet 56:223–232] (1992) as above to create an integrated map for each of several species for which fully annotated genome sequences are not yet available (sheep, cattle, pig, wallaby), using all types of partial maps for that species, including cytogenetic, linkage, somatic-cell hybrid, and radiation hybrid maps. An integrated map provides not only predictions of the kilobase location of every locus, but also predicts locations (in cM) and cytogenetic band locations for every locus. In this way a comprehensive linkage map and a comprehensive cytogenetic map are created, including all loci, irrespective of whether they have ever been linkage mapped or physically mapped, respectively. High-resolution physical maps from annotated sequenced species have also been placed alongside the integrated maps. This has created a powerful tool for comparative genomics. The LDB map-integration strategy has been extended to make use of zoo-FISH comparative information. It has also been extended to enable the creation of a “virtual” map for each species not yet sequenced by using mapping data from fully sequenced species. All of the partial maps, together with the integrated map, for each species have been placed in a database called Comparative Location Database (CompLDB), which is available for querying, browsing, or download in tabular form at . Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

反式作用干扰小RNA

反式作用干扰小RNA（trans—acting siRNA,tasiRNA）是植物中一类内源性siRNA,其成熟过程由miRNA所介导,而且不产生扩增效应。     

Microbial hydroxylation of 16α,17α-dimethyl-17β-(l-oxopropyl)androsta-l,4-dien-3-one to rimexolone by Curvularia lunata AS 3.4381

A novel synthetic route to rimexolone, a corticosteroid for treatment of ocular inflammation without significant elevation of intraocular pressure, was described. An investigation has been undertaken of the microbial transformation of 16α,17α-dimethyl-17β-(l-oxopropyl)androsta-l,4-dien-3-one by microorganisms known to hydroxylate conventional steroids, using Curvularia lunata AS 3.4381 gave rimexolone, the product of 11β-hydroxylation, respectively. The target compound was characterized with reference substance rimexolone by TLC, HPLC, elemental analysis, MS, IR, and NMR.     

Characterization of hypothetical proteins Cpn0146, 0147, 0284 &; 0285 that are predicted to be in the <Emphasis Type="Italic">Chlamydia pneumoniae</Emphasis> inclusion membrane

Background  

Although more than 100 Chlamydia pneumoniae hypothetical proteins have been predicted to be inclusion membrane proteins, only a few have been experimentally demonstrated to be in the inclusion membrane. Using antibodies raised with fusion proteins, we characterized four such hypothetical proteins encoded by two gene clusters (Cpn0146-147 and Cpn0284-285) in the C. pneumoniae genome.     

A strong constitutive ethylene-response phenotype conferred on Arabidopsis plants containing null mutations in the ethylene receptors <Emphasis Type="Italic">ETR1</Emphasis> and <Emphasis Type="Italic">ERS1</Emphasis>

Background  

The ethylene receptor family of Arabidopsis consists of five members, falling into two subfamilies. Subfamily 1 is composed of ETR1 and ERS1, and subfamily 2 is composed of ETR2, ERS2, and EIN4. Although mutations have been isolated in the genes encoding all five family members, the only previous insertion allele of ERS1 (ers1-2) is a partial loss-of-function mutation based on our analysis. The purpose of this study was to determine the extent of signaling mediated by subfamily-1 ethylene receptors through isolation and characterization of null mutations.