Calcineurin is expressed and plays a critical role in inflammatory arthritis

Calcineurin is a calcium-activated phosphatase to mediate lymphocyte activation and neuron signaling, but its role in inflammatory arthritis remains largely unknown. In this study, we demonstrate that calcineurin was highly expressed in the lining layer, infiltrating leukocytes, and endothelial cells of rheumatoid synovium. The basal expression levels of calcineurin were higher in the cultured synoviocytes of rheumatoid arthritis patients than those of osteoarthritis patients. The calcineurin activity in the synoviocytes was increased by the stimulation with proinflammatory cytokines such as IL-1beta and TNF-alpha. Moreover, rheumatoid arthritis synoviocytes had an enlarged intracellular Ca(2+) store and showed a higher degree of [Ca(2+)](i) release for calcineurin activity than osteoarthritis synoviocytes when stimulated with either TNF-alpha or phorbol myristate acetate. IL-10, an anti-inflammatory cytokine, failed to increase the Ca(2+) and calcineurin activity. The targeted inhibition of calcineurin by the overexpression of calcineurin-binding protein 1, a natural calcineurin antagonist, inhibited the production of IL-6 and matrix metalloproteinase-2 by rheumatoid synoviocytes in a similar manner to the calcineurin inhibitor, cyclosporin A. Moreover, the abundant calcineurin expression was found in the invading pannus in the joints of mice with collagen-induced arthritis. In these mice, calcineurin activity in the cultured synovial and lymph node cells correlated well with the severity of arthritis, but which was suppressed by cyclosporin A treatment. Taken together, our data suggest that the abnormal activation of Ca(2+) and calcineurin in the synoviocytes may contribute to the pathogenesis of chronic arthritis and thus provide a potential target for controlling inflammatory arthritis.     

Hox genes in the echiuroid Urechis unicinctus

An echiuroid species, Urechis unicinctus, was surveyed for Hox genes using polymerase chain reaction with homeobox-specific degenerate primers. We identified nine distinct homeodomain-containing gene fragments. These nine fragments were classified by comparative analysis. This analysis revealed that this echiuroid possessed at least three Hox genes from the anterior group, five from the central group, and one from the posterior group.Sung-Jin and Dae-Hee Lee contributed equally to this work.An erratum to this article can be found at     

Dachs: an unconventional myosin that functions downstream of Fat to regulate growth, affinity and gene expression in Drosophila

The dachs gene was first identified almost a century ago based on its requirements for appendage growth, but has been relatively little studied. Here, we describe the phenotypes of strong dachs mutations, report the cloning of the dachs gene, characterize the localization of Dachs protein, and investigate the relationship between Dachs and the Fat pathway. Mutation of dachs reduces, but does not abolish, the growth of legs and wings. dachs encodes an unconventional myosin that preferentially localizes to the membrane of imaginal disc cells. dachs mutations suppress the effects of fat mutations on gene expression, cell affinity and growth in imaginal discs. Dachs protein localization is influenced by Fat, Four-jointed and Dachsous, consistent with its genetic placement downstream of fat. However, dachs mutations have only mild tissue polarity phenotypes, and only partially suppress the tissue polarity defects of fat mutants. Our results implicate Dachs as a crucial downstream component of a Fat signaling pathway that influences growth, affinity and gene expression during development.     

Negative regulation of Hedgehog signaling by the cholesterogenic enzyme 7-dehydrocholesterol reductase

Cholesterol regulates Hedgehog (Hh) signaling during early vertebrate development. Smith-Lemli-Opitz syndrome (SLOS) is caused by defects in 7-dehydrocholesterol reductase (DHCR7), an enzyme catalyzing the final step of cholesterol biosynthesis. Many developmental malformations attributed to SLOS occur in tissues and organs where Hh signaling is required for development, but the precise role of DHCR7 deficiency in this disease remains murky. We report that DHCR7 and Sonic Hedgehog (Shh) are co-expressed during midline development in Xenopus embryos. DHCR7 has previously been implicated to function as a positive regulator of Hh signaling that acts to regulate the cholesterol adduction of Hh ligand or to affect Hh signaling in the responding cell. We present gain- and loss-of-function analyses suggesting that DHCR7 functions as a negative regulator of Hh signaling at the level or downstream of Smoothened (Smo) and affects intracellular Hh signaling. Our analysis also raises the possibility that the human condition SLOS is caused not only by disruption of the enzymatic role of DHCR7 as a reductase in cholesterol biosynthesis, but may also involve defects in DHCR7 resulting in derepression of Shh signaling.     

The existence of all three ParaHox genes in the clitellate annelid, Perionyx excavatus

A ParaHox gene cluster is composed of three genes (Gsx, Xlox, and Cdx). It has been proposed that all three ParaHox genes were present in the last common ancestor to the lophotrochozoan protostomes and the deuterostomes and that gene loss event has occurred in the ecdysozoan lineage. In this paper, we report the existence of all three ParaHox genes in Perionyx excavatus, a clitellate annelid. Although orthologs of each of the three ParaHox genes were previously discovered from other lopotrochozoan taxa, this study constitutes the first reported isolation of all three ParaHox genes in the same clitellate species.Bum Joon Park and Sung-Jin Cho contributed equally to this work.     

Expression, purification and transduction of PEP-1-botulinum neurotoxin type A (PEP-1-BoNT/A) into skin

Botulinum neurotoxin A (BoNT/A) has been used therapeutically to treat muscular hypercontractions and sudomotor hyperactivity and it has been reported that BoNT/A might have analgesic properties in headache. PEP-1 peptide is a known carrier peptide that delivers full-length native proteins in vitro and in vivo. In this study, a BoNT/A gene were fused with PEP-1 peptide in a bacterial expression vector to produce a genetic in-frame PEP-1-BoNT/A fusion protein. The expressed and purified PEP-1-BoNT/A fusion proteins were efficiently transduced into cells in a time- and dose-dependent manner when added exogenously in a culture medium. In addition, immunohistochemical analysis revealed that PEP-1-BoNT/A fusion protein efficiently penetrated into the epidermis as well as the dermis of the subcutaneous layer, when sprayed on mice skin. These results suggest that PEP-1-BoNT/A fusion protein provide an efficient strategy for therapeutic delivery in various human diseases related to this protein.     

One-step enzymatic synthesis of blue pigments from geniposide for fabric dyeing

In this study, we describe a one-step chemoenzymatic reaction for the production of natural blue pigments, in which the geniposide from Gardenia extracts is transformed by glycosidases to genipin. Genipin is then allowed to react with amino acids, thereby generating a natural blue pigment. The β-glycosidases, most notably isolase (a variant of β-glucanase), recombinant β-glucosidase, Cellulase T, and amylases, were shown to hydrolyze geniposide to produce the desired pigments, whereas the α-glycosidases did not. Among the 20 tested amino acids, glycine and tyrosine were associated with the highest dye production yields. The optimal molar ratio of geniposide to glycine, two reactants relevant to pigment production, was unity. The natural blue pigments produced in this study were used to dye cotton, silk, and wool. The color yields of the pigments were determined to be significantly higher than those of other natural dyes. Furthermore, the color fastness properties of these dyes were fairly good, even in the absence of mordant.     

Identification and expression of the cym, cmt, and tod catabolic genes from Pseudomonas putida KL47: expression of the regulatory todST genes as a factor for catabolic adaptation

Pseudomonas putida KL47 is a natural isolate that assimilates benzene, 1-alkylbenzene (C(1)-C(4)), biphenyl, p-cumate, and p-cymene. The genetic background of strain KL47 underlying the broad range of growth substrates was examined. It was found that the cym and cmt operons are constitutively expressed due to a lack of the cymR gene, and the tod operon is still inducible by toluene and biphenyl. The entire array of gene clusters responsible for the catabolism of toluene and p-cymene/p-cumate has been cloned in a cosmid vector, pLAFR3, and were named pEK6 and pEK27, respectively. The two inserts overlap one another and the nucleotide sequence (42,505 bp) comprising the cym, cmt, and tod operons and its flanking genes in KL47 are almost identical (>99%) to those of P. putida F1. In the cloned DNA fragment, two genes with unknown functions, labeled cymZ and cmtR, were newly identified and show high sequence homology to dienelactone hydrolase and CymR proteins, respectively. The cmtR gene was identified in the place of the cmtI gene of previous annotation. Western blot analysis showed that, in strains F1 and KL47, the todT gene is not expressed during growth on Luria Bertani medium. In minimal basal salt medium, expression of the todT gene is inducible by toluene, but not by biphenyl in strain F1; however, it is constantly expressed in strain KL47, indicating that high levels of expression of the todST genes with one amino acid substitution in TodS might provide strain KL47 with a means of adaptation of the tod catabolic operon to various aromatic hydrocarbons.     

WebCell: a web-based environment for kinetic modeling and dynamic simulation of cellular networks

SUMMARY: WebCell is a web-based environment for managing quantitative and qualitative information on cellular networks and for interactively exploring their steady-state and dynamic behaviors in response to systemic perturbations. It is designed as a user-friendly web interface, allowing users to efficiently construct, visualize, analyze and store reaction network models, thereby facilitating kinetic modeling and in silico simulation of biological systems of interest. A collected model library is also available to provide comprehensive implications for cellular dynamics of the published models.