ATL-derived factor (ADF), an IL-2 receptor/Tac inducer homologous to thioredoxin; possible involvement of dithiol-reduction in the IL-2 receptor induction.

HTLV-I transformed T cells not only express a large number of interleukin-2 receptors [IL-2R/p55(Tac)], but also produce a factor named ATL-derived factor (ADF) that augments the expression of IL-2R/p55(Tac). Based on a partial N-terminal amino acid sequence, complementary DNA (cDNA) clones for human and mouse ADF were isolated and sequenced. Recombinant ADF produced by COS-7 monkey kidney cells showed IL-2R/Tac inducing activity on YT cells, which are sensitive for ADF. ADF mRNA was strongly expressed in HTLV-I(+) T cells lines, but not in inactivated cells (THP-1, unstimulated PBMC). Furthermore, in normal human peripheral blood mononuclear cells, the expression of ADF mRNA was enhanced by mitogens or phorbol myristate acetate, suggesting a possible involvement of ADF in the lymphocyte activation. Homology analysis revealed an unexpected relationship between ADF and dithiol-reducing enzyme, thioredoxin, involved in many important biological reactions such as the conversion of ribonucleotides into deoxyribonucleotides, or the stabilization of glucocorticoid receptors. The biological significance of the generation of a redox potential in lymphocyte activation, and the possible involvement of dithiol reduction in the induction of IL-2R/Tac are discussed.     

The potential applications of cyanobacterial photosynthesis for clean technologies

Natural photosynthesis may be adapted to advantage in the development of clean energy technologies. Efficient biocatalysts that can be used in solar energy conversion technologies are the cyanobacteria. Photobioreactors incorporating cyanobacteria have been used to demonstrate (a) the production of hydrogen gas, (b) the assimilation of CO₂ with the production of algal biomass, (c) the excretion of ammonium, and (d) the removal of nitrate and phosphate from contaminated waters.Abbreviations Chl chlorophyll - DW dry weight - MSX L-methionine-D-L-sulphoximine - PAR photosynthetically active radiation - PU polyurethane - PV polyvinyl - PVC polyvinylchloride     

Fragments Ba and Bb derived from guinea pig factor B of the properdin system: purification, characterization, and biologic activities.

Functionally active guinea pig factor B was purified by a combination of chromatographic steps including Sephadex G-25, QAE A25, QAE A50, CM C50, and Sepharose 4B coupled with purified cobra venom factor. Purified factor B had a m.w. of 106,000 daltons and a single subunit structure. It was heat labile. After cleavage of native B with cobra venom factor coupled to Sepharose 4B in the presence of D, the resulting two fragments, the larger one (Bb) and the smaller one (Ba), were further purified. The m.w. of Bb and Ba was determined as 64,000 and 53,000 daltons, respectively, by SDS-PAGE. Neither of the fragments evoked a contraction of guinea pig ileum or histamine release from rat mast cells. Only the smaller fragment Ba (at a concentration of 120 nM) stimulated guinea pig peritoneal polymorphonuclear leukocytes to respond with increased movement. This activity as well as the antigenicity of Ba were heat stable, but were sensitive to trypsin digestion, whereas the antigenicity of Bb was heat labile.     

Development of a System to Monitor Laryngeal Movement during Swallowing Using a Bend Sensor

Background

Swallowing dysfunction (also known as dysphagia), which results in a deterioration of nutritional intake, slows rehabilitation and causes aspiration pneumonia, is very common following neurological impairments. Although videofluorographic (VF) examination is widely used for detecting aspiration, an objective and non-invasive method for assessing swallowing function has yet to be established because of a lack of adequate devices and protocols. In this paper, a bend sensor whose resistance is altered by bending was introduced to monitor swallowing-related laryngeal movement.

Methods

Six healthy male volunteers were recruited in the present study. Specific time points on the signal waveform produced by the bend sensor were defined to describe laryngeal movement by differential analysis. Additionally, the physiological significance of the obtained waveform was confirmed by analyzing the sequential correlations between the signal waveform from the bend sensor and hyoid bone kinetics simultaneously recorded by VF.

Results

Seven time points were successfully defined on the signal waveform to reference laryngeal movement. Each time point was well correlated with certain VF events, with evidence of no significant time lags, and there were positive correlations between waveform time points and matched VF events. Furthermore, obvious similarities were noticed between the duration of each phase on the signal waveform and the duration of the matched hyoid bone activity.

Conclusions

The present monitoring system using a bend sensor might be useful for observing the temporal aspects of laryngeal movement during swallowing, and it was well coordinated with hyoid bone movement.     

Characterization of the complex formed between a potent neutralizing ovine-derived polyclonal anti-TNFα Fab fragment and human TNFα

TNFα (tumour necrosis factor α) is an early mediator in the systemic inflammatory response to infection and is therefore a therapeutic target in sepsis. AZD9773 is an ovine-derived, polyclonal anti-TNFα Fab fragment derived from a pool of serum and currently being developed as a treatment for severe sepsis and septic shock. In the present study, we show that although AZD9773 has a modest affinity for TNFα in a binding assay, the K_i in a cell-based assay is approximately four orders of magnitude lower. We show using SEC (size exclusion chromatography) that the maximum size of the complex between AZD9773 and TNFα is consistent with approximately 12 Fabs binding to one TNFα trimer. A number of approaches were taken to map the epitopes recognized by AZD9773. These revealed that a number of different regions on TNFα are involved in binding to the polyclonal Fab. The data suggest that there are probably three epitopes per monomer that are responsible for most of the inhibition by AZD9773 and that all three can be occupied at the same time in the complex. We conclude that AZD9773 is clearly demonstrated to bind to multiple epitopes on TNFα and suggest that the polyclonal nature may account, at least in part, for the very high potency observed in cell-based assays.     

Specific Midgut Region Controlling the Symbiont Population in an Insect-Microbe Gut Symbiotic Association

Many insects possess symbiotic bacteria that affect the biology of the host. The level of the symbiont population in the host is a pivotal factor that modulates the biological outcome of the symbiotic association. Hence, the symbiont population should be maintained at a proper level by the host''s control mechanisms. Several mechanisms for controlling intracellular symbionts of insects have been reported, while mechanisms for controlling extracellular gut symbionts of insects are poorly understood. The bean bug Riptortus pedestris harbors a betaproteobacterial extracellular symbiont of the genus Burkholderia in the midgut symbiotic organ designated the M4 region. We found that the M4B region, which is directly connected to the M4 region, also harbors Burkholderia symbiont cells, but the symbionts therein are mostly dead. A series of experiments demonstrated that the M4B region exhibits antimicrobial activity, and the antimicrobial activity is specifically potent against the Burkholderia symbiont but not the cultured Burkholderia and other bacteria. The antimicrobial activity of the M4B region was detected in symbiotic host insects, reaching its highest point at the fifth instar, but not in aposymbiotic host insects, which suggests the possibility of symbiont-mediated induction of the antimicrobial activity. This antimicrobial activity was not associated with upregulation of antimicrobial peptides of the host. Based on these results, we propose that the M4B region is a specialized gut region of R. pedestris that plays a critical role in controlling the population of the Burkholderia gut symbiont. The molecular basis of the antimicrobial activity is of great interest and deserves future study.     

Carboxy terminus of glucose transporter 3 contains an apical membrane targeting domain

We previously demonstrated that distinct facilitative glucose transporter isoforms display differential sorting in polarized epithelial cells. In Madin-Darby canine kidney (MDCK) cells, glucose transporter 1 and 2 (GLUT1 and GLUT2) are localized to the basolateral cell surface whereas GLUTs 3 and 5 are targeted to the apical membrane. To explore the molecular mechanisms underlying this asymmetric distribution, we analyzed the targeting of chimeric glucose transporter proteins in MDCK cells. Replacement of the carboxy-terminal cytosolic tail of GLUT1, GLUT2, or GLUT4 with that from GLUT3 resulted in apical targeting. Conversely, a GLUT3 chimera containing the cytosolic carboxy terminus of GLUT2 was sorted to the basolateral membrane. These findings are not attributable to the presence of a basolateral signal in the tails of GLUTs 1, 2, and 4 because the basolateral targeting of GLUT1 was retained in a GLUT1 chimera containing the carboxy terminus of GLUT5. In addition, we were unable to demonstrate the presence of an autonomous basolateral sorting signal in the GLUT1 tail using the low-density lipoprotein receptor as a reporter. By examining the targeting of a series of more defined GLUT1/3 chimeras, we found evidence of an apical targeting signal involving residues 473-484 (DRSGKDGVMEMN) in the carboxy tail. We conclude that the targeting of GLUT3 to the apical cell surface in MDCK cells is regulated by a unique cytosolic sorting motif.     

Genetic variations at urotensin II and urotensin II receptor genes and risk of type 2 diabetes mellitus in Japanese

Urotensin II is among the most potent vasoactive hormones known and the urotensin II (UTS2) gene is localized to 1p36-p32, one of the regions reported to show possible linkage with type 2 diabetes in Japanese. When we surveyed genetic polymorphisms in the UTS2 and urotensin II receptor (GPR14) gene, we identified two SNPs with amino acid substitutions (designated T21M and S89N and an SNP in the promotor region (-605G>A) of the UTS2 gene, and two SNPs in the non-coding region of the GPR14 gene. We then studied these three SNPs in the UTS2 gene and two SNPs in the GPR14 gene in 152 Japanese subjects with type 2 diabetes mellitus and two control Japanese populations. The allele frequency of 89N was significantly higher in type 2 diabetic patients than in both elderly normal subjects (P = 0.0018) and subjects with normal glucose tolerance (P = 0.0011), whereas the allele frequency of T21M and -605G>A in the UTS2 gene and those of two SNPs in the GPR14 gene were essentially identical in these three groups. Furthermore, in the subjects with normal glucose tolerance, 89N was associated with significantly higher insulin levels on oral glucose tolerance test, suggesting reduced insulin sensitivity in subjects with 89N. These results strongly suggest that subjects with S89N in the UTS2 gene are more insulin-resistant and thus more susceptible to type 2 diabetes mellitus development.     

Dissecting interdomain communication within cAPK regulatory subunit type IIbeta using enhanced amide hydrogen/deuterium exchange mass spectrometry (DXMS)

cAMP-dependent protein kinase (cAPK) is a heterotetramer containing a regulatory (R) subunit dimer bound to two catalytic (C) subunits and is involved in numerous cell signaling pathways. The C-subunit is activated allosterically when two cAMP molecules bind sequentially to the cAMP-binding domains, designated A and B (cAB-A and cAB-B, respectively). Each cAMP-binding domain contains a conserved Arg residue that is critical for high-affinity cAMP binding. Replacement of this Arg with Lys affects cAMP affinity, the structural integrity of the cAMP-binding domains, and cAPK activation. To better understand the local and long-range effects that the Arg-to-Lys mutation has on the dynamic properties of the R-subunit, the amide hydrogen/deuterium exchange in the RIIbeta subunit was probed by electrospray mass spectrometry. Mutant proteins containing the Arg-to-Lys substitution in either cAMP-binding domain were deuterated for various times and then, prior to mass spectrometry analysis, subjected to pepsin digestion to localize the deuterium incorporation. Mutation of this Arg in cAB-A (Arg230) causes an increase in amide hydrogen exchange throughout the mutated domain that is beyond the modest and localized effects of cAMP removal and is indicative of the importance of this Arg in domain organization. Mutation of Arg359 (cAB-B) leads to increased exchange in the adjacent cAB-A domain, particularly in the cAB-A domain C-helix that lies on top of the cAB-B domain and is believed to be functionally linked to the cAB-B domain. This interdomain communication appears to be a unidirectional pathway, as mutation of Arg230 in cAB-A does not effect dynamics of the cAB-B domain.