BACKGROUND: Calliptamus italicus L. is a major pest in Xinjiang grassland. The diapause overwintering strategy is one of the important reasons for the large population of this pest. This study investigated the function of the genes associated with the release of diapause (DIB, JHE and CAM) in Calliptamus italicus by RNA interference (RNAi) technology to aid in its biological control.
RESULTS: The expression levels of DIB and its downstream-associated genes (EcR and FTZ-F1) in the eggs injected with dsDIB for 12 h decreased by 96.6%, 55.8% and 81.8%, respectively. Diapause began to terminate on day 3, and development was almost complete on day 6. However, the head was significantly smaller. The expression levels of JHE and its downstream-associated genes (JHEH and VgR) at 48 h after dsJHE treatment decreased by 76.5%, 85.6% and 85.9%, respectively. The termination of diapause occured on day 3 of incubation. The development was basically complete on day 6, but the yolk had been incompletely absorbed. The expression of CAM and its downstream-associated genes (CAMK4 and MYL) at 24 h after dsCAM treatment decreased by 42.4%, 95.3% and 82.7%, respectively. Diapause termination was completed on day 4 for incubation, and development was abnormal on day 6. The absorption of yolk was incomplete.
CONCLUSIONS: DIB, JHE and CAM can delay the diapause termination of Calliptamus italicus eggs to different degrees and can be developed as potential target genes for its biological control. © 2024 Society of Chemical Industry.

We exploit single-molecule tracking and optical single channel recording in droplet interface bilayers to resolve the assembly pathway and pore formation of the archetypical cholesterol-dependent cytolysin nanopore, Perfringolysin O. We follow the stoichiometry and diffusion of Perfringolysin O complexes during assembly with 60 ms temporal resolution and 20 nm spatial precision. Our results suggest individual nascent complexes can insert into the lipid membrane where they continue active assembly. Overall, these data support a model of stepwise irreversible assembly dominated by monomer addition, but with infrequent assembly from larger partial complexes.

Droplet interface bilayer (DIB) is a method of fabricating lipid bilayer membrane by contacting two aqueous droplets coated with a monolayer of lipid molecules in oil media. Lipids coat the droplet surface either by vesicles fusing to the water-oil interface from the droplet side or diffusing toward the interface from the oil side, thereby forming a lipid monolayer. With the DIB technique, nanoliter amounts of aqueous solution is needed and one may obtain two different compositions of monolayers to form asymmetric bilayer which is difficult to replicate by other in vitro lipid membrane methods. Here, a DIB-based protocol is reported to fabricate a stable lipid bilayer membrane to perform single-channel electrophysiology on a pore-forming toxin.

The objective of this study was to evaluate the effects of post artificial insemination (AI) treatment with intravaginal progesterone device (P4 device) on conception rate, synchronization of returning estrus and plasma P4 concentration in Japanese Black cows. Nineteen cows were treated with DIB (1.0 g P4) from Day 12 to 19 (Day 0=day of the first AI), 27 cows were treated with a CIDR (1.9 g P4) from Day 12 to 19, and 33 cows were not treated after the first AI (control). Estrous behavior was daily examined between Day 20 and 25, and cows returning to estrus were inseminated (the second AI). On Day 19, plasma P4 concentration was not different among DIB, CIDR and control groups. There was no significant difference in conception rate after the first AI among three groups (DIB: 63.2%, CIDR: 66.7% and control: 72.7%). In non-pregnant cows, there was no significant difference in the proportion of cows showed returning estrus between Day 20 and 25 (DIB: 57.1%, CIDR: 22.2% and control: 44.4%), and day of returning estrus was not synchronized. The overall conception rate after the first and second AI was not different among the groups. In conclusion, post-AI treatment with intravaginal devices containing 1.0 and 1.9 g P4 from Day 12 to 19 neither increased plasma P4 concentration nor improved fertility and synchronization of the returning estrus in Japanese Black cows.

The study of ion channel activity and the screening of possible inhibitor molecules require reliable methods for production of active channel proteins, their insertion into artificial membranes and for the measurement of their activity. Here we report on cell-free expression of soluble and active Kv1.1 and Kv1.3 channels and their efficient insertion into liposomes. Two complementary methods for the determination of the electrical activity of the proteoliposome-embedded channels were compared using Kv1.1 as a model system: (1) single channel recordings in droplet interface bilayers (DIB) and (2) measurement of the membrane voltage potential generated by a potassium ion diffusion potential using the voltage-sensitive fluorescent dye oxonol VI. Single channel recordings in DIBs proved unreliable because of the non-reproducible fusion of proteoliposomes with an artificial membrane. Therefore, the use of the optical indicator oxonol VI was adapted for 96 well microtiter plates using the ionophore valinomycin as a positive control. The activity of Kv1.1 and Kv1.3 channels was then monitored in the absence and presence of different venom toxins, demonstrating that fluorescent dyes can be used very efficiently when screening small molecules for their channel blocking activity.

Hydroxylated seco-analogs of cytotoxic phenanthroindolizidine alkaloids were prepared in good yields from inexpensive 4-hydroxyproline derivatives, in just two steps. Thus, a sequential oxidative radical scission-oxidation was used for the direct conversion of the proline derivative into a 2-(2-aryl-oxoethyl)pyrrolidine with a variety of aryl and heteroaryl groups. The 4R-stereogenic center allowed ready isomer separation, and stereocontrol in the introduction of new chains (interestingly, the 2,4-cis isomers predominated). In the second step, a cyclization reaction afforded alkaloid analogs with an indolizidinone core; a partial isomerization took place but the isomers were readily purified. Then the cytotoxic activity of the bicyclic indolizidinones and the simpler pyrrolidine derivatives was compared against tumorogenic human neuronal SHSY-5Y and breast cancer MCF7 cells. All the biphenyl derivatives displayed a potent activity (one derivative caused >80% cell death in both tumor lines at micromolar dosis), being comparable in the pyrrolidine and indolizidinone series.