Functional analysis of a miRNA‐like small RNA derived from Bombyx mori cytoplasmic polyhedrosis virus

Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) is a major pathogen of the economic insect silkworm, Bombyx mori. Virus‐encoded microRNAs (miRNAs) have been proven to play important roles in host–pathogen interactions. In this study we identified a BmCPV‐derived miRNA‐like 21 nt small RNA, BmCPV‐miR‐1, from the small RNA deep sequencing of BmCPV‐infected silkworm larvae by stem‐loop quantitative real‐time PCR (qPCR) and investigated its functions with qPCR and lentiviral expression systems. Bombyx mori inhibitor of apoptosis protein (BmIAP) gene was predicted by both target prediction software miRanda and Targetscan to be one of its target genes with a binding site for BmCPV‐miR‐1 at the 5′ untranslated region. It was found that the expression of BmCPV‐miR‐1 and its target gene BmIAP were both up‐regulated in BmCPV‐infected larvae. At the same time, it was confirmed that BmCPV‐miR‐1 could up‐regulate the expression of BmIAP gene in HEK293T cells with lentiviral expression systems and in BmN cells by transfecting mimics. Furthermore, BmCPV‐miR‐1 mimics could up‐regulate the expression level of BmIAP gene in midgut and fat body in the silkworm. In the midgut of BmCPV‐infected larvae, BmCPV‐miR‐1 mimics could be further up‐regulated and inhibitors could lower the virus‐mediated expression of BmIAP gene. With the viral genomic RNA segments S1 and S10 as indicators, BmCPV‐miR‐1 mimics could up‐regulate and inhibitors down‐regulate their replication in the infected silkworm. These results implied that BmCPV‐miR‐1 could inhibit cell apoptosis in the infected silkworm through up‐regulating BmIAP expression, providing the virus with a better cell circumstance for its replication.     

Functional RelBE-Family Toxin-Antitoxin Pairs Affect Biofilm Maturation and Intestine Colonization in Vibrio cholerae

Toxin–antitoxin (TA) systems are small genetic elements that typically encode a stable toxin and its labile antitoxin. These cognate pairs are abundant in prokaryotes and have been shown to regulate various cellular functions. Vibrio cholerae, a human pathogen that is the causative agent of cholera, harbors at least thirteen TA loci. While functional HigBA, ParDE have been shown to stabilize plasmids and Phd/Doc to mediate cell death in V. cholerae, the function of seven RelBE-family TA systems is not understood. In this study we investigated the function of the RelBE TA systems in V. cholerae physiology and found that six of the seven relBE loci encoded functional toxins in E. coli. Deletion analyses of each relBE locus indicate that RelBE systems are involved in biofilm formation and reactive oxygen species (ROS) resistance. Interestingly, all seven relBE loci are induced under the standard virulence induction conditions and two of the relBE mutants displayed a colonization defect, which was not due to an effect on virulence gene expression. Although further studies are needed to characterize the mechanism of action, our study reveals that RelBE systems are important for V. cholerae physiology.     

A comparative analysis of photosynthetic characteristics of hulless barley at two altitudes on the Tibetan Plateau

To determine the photosynthetic characteristics of C₃ plants and their sensitivity to CO₂ at different altitudes on the Tibetan Plateau, hulless barley (Hordeum vulgare L. ssp. vulgare) was grown at altitudes of 4,333 m and 3,688 m. Using gas-exchange measurements, photosynthetic parameters were simulated, including the maximum net photosynthesis (P _max) and the apparent quantum efficiency (α). Plants growing at higher altitude had higher net photosynthetic rates (P _N), photosynthesis parameters (P _max and α) and sensitivities to CO₂ enhancement than plants growing at lower altitude on the Tibetan Plateau. The enhancements of P _N, P _max, and α for plants growing at higher altitude, corresponding with 10 μmol(CO₂) mol⁻¹ increments, were approximately 0.20∼0.45%, 0.05∼0.20% and 0.12∼0.36% greater, respectively, than for plants growing at lower altitude, respectively, where CO₂ levels rose from 10 to 170 μmol(CO₂) mol⁻¹. Therefore, on the Tibetan Plateau, the changes in the photosynthetic capacities and the photosynthetic sensitivities to CO₂ observed in the C₃ plants grown above 3,688 m are likely to increase with altitude despite the decreasing CO₂ partial pressure.     

Ovine fetal electrocortical activity and regional cerebral blood flow

Fetuses of 12 near-term sheep were prepared for microsphere determination of cerebral blood flow. Experiments were performed 5 days postsurgery. The regional blood flows were measured in successive high (HV), low (LV) and high voltage electrocorticographic states. Comparisons were made between the observations made in the LV and averaged flanking HV cycles. Total cerebral blood flow was 95 +/- 8, 119 +/- 11 and 100 +/- 9 ml/min/100 g in HV, LV and HV, respectively. Low voltage electrocortical activity increased average cerebral blood flow by 22% (P less than 0.01). Significant changes were seen in all regions except the occipital cortex. The maximum change was observed in the thalamus in which the flows were 152 +/- 23, 243 +/- 35 and 138 +/- 20 ml/min/per 100 g tissue, respectively. The increase was 68% (P less than 0.001). The percent changes seen in the cerebrum are as follows: Frontal grey + 18%, frontal white + 22%, parietal white + 22%, temporal + 18%. A + 17% change was seen in the cord (P less than 0.03). It is concluded that in low voltage electrocortical activity all of the brain, except the occipital region, shows an increase in cerebral blood flow. This is probably secondary to a variance in cerebral activity. This preparation may be useful in localizing function in the fetal brain.     

Interaction and combination of separate A and B chains of insulin effects of D-and L-tryptophan in position A1

The S-thiomethyl derivatives of insulin A chain with A1-Gly replaced by D- or L-Trp have been prepared and their respective interaction and combination with the S-thiomethyl B chain studied. The UV difference spectra of the mixed against the separated [Trp1]A chains with the B chain at pH 10.8 are similar to those obtained for the unmodified chains except that the 295-nm-negative peak for ionized Tyr residue appears to be less marked. Fluorescence studies show very little environmental changes at the A1-Trp residues when mixed with the B chain. The intact hormone with A1-Gly replaced by D-Trp is known to be considerably more active than the analog with L-Trp replacement. However, for both derivatives the resynthesis of the whole molecules correctly joined by disulfide bridges starting from the separated reduced chains, gives similar low yields as shown by HPLC analysis and by receptor-binding assay. The replacement of A1-Gly by D-Trp appears to affect the separated A chain more than the intact hormone and replacements at A1 by both D- and L-Trp probably lead to significant conformational changes of the A chain so as to prevent its correct pairing with the B chain.     

Comparison of different methods of immobilization for lignin peroxidase production by Phanerochaete chrysosporium

Summary Phanerochaete chrysosporium was immobilized in agar, agarose and -carrageenan gel beads, nylon web, and polyurethane foam, and used for the production of lignin peroxidase in shake cultures on a carbon-limited medium. Nylon was found to be the best carrier, with the maximum lignin peroxidase activity (340 U/l) reached on the 7th day. The enzyme production rate was significantly lower with freely suspended mycelial pellets. Both nylon and polyurethane based biocatalysts were active for at least 38 days after the addition of veratryl alcohol. Best results were obtained when a spore inoculum was used instead of day-old pellets. -Carrageenan was found unsuitable as a carrier for lignin peroxidase production.     

Resonance electroconformational coupling: A proposed mechanism for energy and signal transductions by membrane proteins

Recent experiments show that membrane ATPases are capable of absorbing free energy from an applied oscillating electric field and converting it to chemical bond energy of ATP or chemical potential energy of concentration gradients. Presumably these enzymes would also respond to endogenous transmembrane electric fields of similar intensity and waveform. A mechanism is proposed in which energy coupling is achieved via Coulombic interaction of an electric field and the conformational equilibria of an ATPase. Analysis indicates that only an oscillating or fluctuating electric field can be used by an enzyme to drive a chemical reaction away from equilibrium.In vivo, the stationary transmembrane potential of a cell must be modulated to become locally oscillatory if it is to derive energy and signal transduction processes.