Efficacy of intercropping and harvest time modification for reducing field infestation of maize by Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae)

Most management practices of Sitophilus zeamais Motschulsky, a field-to-post-harvest insect pest of cereals, have focused on post harvest control methods. This experiment was designed to investigate the potential of cropping system and modification of time of harvest to control S. zeamais. Intercropping and harvest time modification had significant (P < 0.05) effect on the number of S. zeamais emerging 42 days post-harvest. For the early harvest (15 weeks after planting (WAP)), the mean number of S. zeamais recorded from a maize monoculture (7.39) was significantly (P < 0.05) higher than the mean numbers of weevils emerging from a maize–soybean intercrop (2.31), but not significantly higher than the number recorded in maize–groundnut (3.87) intercrop. For the late harvest (18 WAP), the mean number of emerged adult observed in the maize–soybean intercrop (6.13) was significantly lower than the mean number of adult emerging from the monocrop maize (13.24). Maize–groundnut intercrop did not significantly reduce field infestation of S. zeamais compared with monocrop maize. Percentage weight loss observed in early harvested maize was significantly (P < 0.0001) lower than what was observed in late-harvested maize. Percentage weight loss was highest in stored maize harvested from monocrop maize plots for the early harvest, whereas intercropping maize with soybean reduced percentage weight loss when harvest was delayed.     

Regulation of cell proliferation and migration in gallbladder cancer by zinc finger X-chromosomal protein

Gallbladder carcinoma (GBC) is one of the mostly aggressive and fatal malignancies. However, little is known about the oncogenic genes that contributed to the development of GBC. Zinc finger X-chromosomal protein (ZFX) was a novel member of the Krueppel C2H2-type zinc-finger protein family and its down-regulation led to impaired cell growth in human laryngeal squamous cell carcinoma. Here, we aim to investigate the function of ZFX in GBC cell proliferation and migration. Loss of function analysis was performed on GBC cell line (GBC-SD) using lentivirus-mediated siRNA against ZFX. The proliferation, in vitro tumorigenesis (colony-formation) ability as well as cell migration was significantly suppressed after GBC-SD cells which were infected with ZFX-siRNA-expressing lentivirus (Lv-shZFX). Our finding suggested that ZFX promoted the growth and migration of GBC cells and could present a potential molecular target for gene therapy of GBC.     

THE EFFECT OF 40 HOURS OF CONSTANT WAKEFULNESS ON NUMBER COMPARISON PERFORMANCE

We investigated the effects of sleep loss and circadian rhythm on number comparison performance. Magnitude comparison of single-digits is robustly characterized by a distance effect: Close numbers (e.g., 5 versus 6) produce longer reaction times than numbers further apart (e.g., 2 versus 8). This distance effect is assumed to reflect the difficulty of a comparison process based on an analogous representation of general magnitude. Twelve male participants were required to stay awake for 40?h in a quasi-constant-routine protocol. Response speed and accuracy deteriorated between 00:00 and 06:00?h but recovered afterwards during the next day, indicating a circadian rhythm of elementary cognitive function (i.e., attention and speed of mental processing). The symbolic distance effect, however, did not increase during the nighttime, indicating that neither cumulative sleep loss nor the circadian clock prolongs numerical comparison processes. The present findings provide first evidence for a relative insensitivity of symbolic magnitude processing against the temporal variation in energy state. (Author correspondence: michael.steinborn@uni-tuebingen.de)     

Fibroblast Circadian Rhythms of PER2 Expression Depend on Membrane Potential and Intracellular Calcium

The suprachiasmatic nucleus (SCN) of the hypothalamus synchronizes circadian rhythms of cells and tissues throughout the body. In SCN neurons, rhythms of clock gene expression are suppressed by manipulations that hyperpolarize the plasma membrane or lower intracellular Ca²⁺. However, whether clocks in other cells also depend on membrane potential and calcium is unknown. In this study, the authors investigate the effects of membrane potential and intracellular calcium on circadian rhythms in mouse primary fibroblasts. Rhythms of clock gene expression were monitored using a PER2::LUC knockin reporter. Rhythms were lost or delayed at lower (hyperpolarizing) K⁺ concentrations. Bioluminescence imaging revealed that this loss of rhythmicity in cultures was due to loss of rhythmicity of single cells rather than loss of synchrony among cells. In lower Ca²⁺ concentrations, rhythms were advanced or had shorter periods. Buffering intracellular Ca²⁺ by the calcium chelator 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis acetoxymethyl ester (BAPTA-AM) or manipulation of inositol triphosphate (IP₃)-sensitive intracellular calcium stores by thapsigargin delayed rhythms. These results suggest that the circadian clock in fibroblasts, as in SCN neurons, is regulated by membrane potential and Ca²⁺. Changes in intracellular Ca²⁺ may mediate the effects of membrane potential observed in this study. (Author correspondence: welshdk@ucsd.edu)     

Shift Work and Inter‐Individual Differences in Sleep and Sleepiness

Inter‐individual differences in tolerance for shift work have been studied primarily in terms of external factors affecting alertness on the job or the ability to rest and sleep while at home. However, there is increasing evidence that neurobiological factors play a role as well, particularly the major processes involved in the regulation of sleep and wakefulness. These include a sleep homeostatic process seeking to balance wakefulness and sleep and a circadian process seeking to promote wakefulness during the day and sleep during the night. Shift work is associated with a temporal misalignment of these two endogenous processes. During nightwork, this misalignment makes it difficult to stay awake during the nightshift and sleep during the day. However, inter‐individual variability in the processes involved in sleep/wake regulation is substantial. Recent studies have demonstrated the existence of inter‐individual differences in vulnerability to cognitive deficits from sleep loss. Moreover, these inter‐individual differences were shown to constitute a trait. Interestingly, self‐evaluations of sleepiness did not correspond well with the trait inter‐individual variability in objective levels of performance impairment during sleep deprivation. Perhaps because of this discrepancy, in operational settings, the inter‐individual differences in vulnerability to sleep loss do not appear to be limited due to self‐selection mechanisms. Indeed, even among a highly select group of active‐duty jet fighter pilots flying a series of simulated night missions, systematic inter‐individual differences in performance impairment from sleep loss were still observed. There are significant personal and economic consequences to human error and accidents caused by performance deficits due to sleep loss. It is important, therefore, to study the inter‐individual differences in the regulation of sleep and wakefulness in the work environment so that cognitive impairment during shift work may be better anticipated and prevented.     

Facilitated Hyperpolarization Signaling in Vascular Smooth Muscle-overexpressing TRIC-A Channels

The TRIC channel subtypes, namely TRIC-A and TRIC-B, are intracellular monovalent cation-specific channels and likely mediate counterion movements to support efficient Ca²⁺ release from the sarco/endoplasmic reticulum. Vascular smooth muscle cells (VSMCs) contain both TRIC subtypes and two Ca²⁺ release mechanisms; incidental opening of ryanodine receptors (RyRs) generates local Ca²⁺ sparks to induce hyperpolarization and relaxation, whereas agonist-induced activation of inositol trisphosphate receptors produces global Ca²⁺ transients causing contraction. Tric-a knock-out mice develop hypertension due to insufficient RyR-mediated Ca²⁺ sparks in VSMCs. Here we describe transgenic mice overexpressing TRIC-A channels under the control of a smooth muscle cell-specific promoter. The transgenic mice developed congenital hypotension. In Tric-a-overexpressing VSMCs from the transgenic mice, the resting membrane potential decreased because RyR-mediated Ca²⁺ sparks were facilitated and cell surface Ca²⁺-dependent K⁺ channels were hyperactivated. Under such hyperpolarized conditions, L-type Ca²⁺ channels were inactivated, and thus, the resting intracellular Ca²⁺ levels were reduced in Tric-a-overexpressing VSMCs. Moreover, Tric-a overexpression impaired inositol trisphosphate-sensitive stores to diminish agonist-induced Ca²⁺ signaling in VSMCs. These altered features likely reduced vascular tonus leading to the hypotensive phenotype. Our Tric-a-transgenic mice together with Tric-a knock-out mice indicate that TRIC-A channel density in VSMCs is responsible for controlling basal blood pressure at the whole-animal level.     

Energetics and Location of Phosphoinositide Binding in Human Kir2.1 Channels

Kir2.1 channels are uniquely activated by phosphoinositide 4,5-bisphosphate (PI(4,5)P₂) and can be inhibited by other phosphoinositides (PIPs). Using biochemical and computational approaches, we assess PIP-channel interactions and distinguish residues that are energetically critical for binding from those that alter PIP sensitivity by shifting the open-closed equilibrium. Intriguingly, binding of each PIP is disrupted by a different subset of mutations. In silico ligand docking indicates that PIPs bind to two sites. The second minor site may correspond to the secondary anionic phospholipid site required for channel activation. However, 96–99% of PIP binding localizes to the first cluster, which corresponds to the general PI(4,5)P₂ binding location in recent Kir crystal structures. PIPs can encompass multiple orientations; each di- and triphosphorylated species binds with comparable energies and is favored over monophosphorylated PIPs. The data suggest that selective activation by PI(4,5)P₂ involves orientational specificity and that other PIPs inhibit this activation through direct competition.     

Auxiliary KChIP4a Suppresses A-type K+ Current through Endoplasmic Reticulum (ER) Retention and Promoting Closed-state Inactivation of Kv4 Channels

In the brain and heart, auxiliary Kv channel-interacting proteins (KChIPs) co-assemble with pore-forming Kv4 α-subunits to form a native K⁺ channel complex and regulate the expression and gating properties of Kv4 currents. Among the KChIP1–4 members, KChIP4a exhibits a unique N terminus that is known to suppress Kv4 function, but the underlying mechanism of Kv4 inhibition remains unknown. Using a combination of confocal imaging, surface biotinylation, and electrophysiological recordings, we identified a novel endoplasmic reticulum (ER) retention motif, consisting of six hydrophobic and aliphatic residues, 12–17 (LIVIVL), within the KChIP4a N-terminal KID, that functions to reduce surface expression of Kv4-KChIP complexes. This ER retention capacity is transferable and depends on its flanking location. In addition, adjacent to the ER retention motif, the residues 19–21 (VKL motif) directly promote closed-state inactivation of Kv4.3, thus leading to an inhibition of channel current. Taken together, our findings demonstrate that KChIP4a suppresses A-type Kv4 current via ER retention and enhancement of Kv4 closed-state inactivation.     

Up-regulation of GABAB Receptor Signaling by Constitutive Assembly with the K+ Channel Tetramerization Domain-containing Protein 12 (KCTD12)

GABA_B receptors are the G-protein coupled receptors (GPCRs) for GABA, the main inhibitory neurotransmitter in the central nervous system. Native GABA_B receptors comprise principle and auxiliary subunits that regulate receptor properties in distinct ways. The principle subunits GABA_B1a, GABA_B1b, and GABA_B2 form fully functional heteromeric GABA_B(1a,2) and GABA_B(1b,2) receptors. Principal subunits regulate forward trafficking of the receptors from the endoplasmic reticulum to the plasma membrane and control receptor distribution to axons and dendrites. The auxiliary subunits KCTD8, -12, -12b, and -16 are cytosolic proteins that influence agonist potency and G-protein signaling of GABA_B(1a,2) and GABA_B(1b,2) receptors. Here, we used transfected cells to study assembly, surface trafficking, and internalization of GABA_B receptors in the presence of the KCTD12 subunit. Using bimolecular fluorescence complementation and metabolic labeling, we show that GABA_B receptors associate with KCTD12 while they reside in the endoplasmic reticulum. Glycosylation experiments support that association with KCTD12 does not influence maturation of the receptor complex. Immunoprecipitation and bioluminescence resonance energy transfer experiments demonstrate that KCTD12 remains associated with the receptor during receptor activity and receptor internalization from the cell surface. We further show that KCTD12 reduces constitutive receptor internalization and thereby increases the magnitude of receptor signaling at the cell surface. Accordingly, knock-out or knockdown of KCTD12 in cultured hippocampal neurons reduces the magnitude of the GABA_B receptor-mediated K⁺ current response. In summary, our experiments support that the up-regulation of functional GABA_B receptors at the neuronal plasma membrane is an additional physiological role of the auxiliary subunit KCTD12.     

Caveolin-1 Facilitates the Direct Coupling between Large Conductance Ca2+-activated K+ (BKCa) and Cav1.2 Ca2+ Channels and Their Clustering to Regulate Membrane Excitability in Vascular Myocytes

L-type voltage-dependent Ca²⁺ channels (LVDCC) and large conductance Ca²⁺-activated K⁺ channels (BK_Ca) are the major factors defining membrane excitability in vascular smooth muscle cells (VSMCs). The Ca²⁺ release from sarcoplasmic reticulum through ryanodine receptor significantly contributes to BK_Ca activation in VSMCs. In this study direct coupling between LVDCC (Cav1.2) and BK_Ca and the role of caveoline-1 on their interaction in mouse mesenteric artery SMCs were examined. The direct activation of BK_Ca by Ca²⁺ influx through coupling LVDCC was demonstrated by patch clamp recordings in freshly isolated VSMCs. Using total internal reflection fluorescence microscopy, it was found that a large part of yellow fluorescent protein-tagged BK_Ca co-localized with the cyan fluorescent protein-tagged Cav1.2 expressed in the plasma membrane of primary cultured mouse VSMCs and that the two molecules often exhibited FRET. It is notable that each BKα subunit of a tetramer in BK_Ca can directly interact with Cav1.2 and promotes Cav1.2 cluster in the molecular complex. Furthermore, caveolin-1 deficiency in knock-out (KO) mice significantly reduced not only the direct coupling between BK_Ca and Cav1.2 but also the functional coupling between BK_Ca and ryanodine receptor in VSMCs. The measurement of single cell shortening by 40 mm K⁺ revealed enhanced contractility in VSMCs from KO mice than wild type. Taken together, caveolin-1 facilitates the accumulation/clustering of BK_Ca-LVDCC complex in caveolae, which effectively regulates spatiotemporal Ca²⁺ dynamics including the negative feedback, to control the arterial excitability and contractility.