A sustained increase in the intracellular Ca²âº concentration induces proteolytic cleavage of EAG2 channel.

Voltage-gated EAG2 channel is abundant in the brain and enhances cancer cell growth by controlling cell volume. The channel contains a cyclic nucleotide-binding homology (CNBH) domain and multiple calmodulin-binding motifs. Here we show that a raised intracellular Ca(2+) concentration causes proteolytic digestion of heterologously expressed and native EAG2 channels. A treatment of EAG2-expressing cells with the Ca(2+) ionophore A23187 for 1h reduces the full-length protein by ∼80% with a concomitant appearance of 30-35-kDa peptides. Similarly, a treatment with the Ca(2+)-ATPase inhibitor thapsigargin for 3h removes 30-35-kDa peptides from ∼1/3 of the channel protein. Moreover, an incubation of the isolated rat brain membrane with CaCl2 leads to the generation of fragments with similar sizes. This Ca(2+)-induced digestion is not seen with EAG1. Mutations in a C-terminal calmodulin-binding motif alter the degrees and positions of the cleavage. Truncated channels that mimic the digested proteins exhibit a reduced current density and altered channel gating. In particular, these shorter channels lack a rapid activation typical in EAG channels with more than 20-mV positive shifts in voltage dependence of activation. The truncation also eliminates the ability of EAG2 channel to reduce cell volume. These results suggest that a sustained increase in the intracellular Ca(2+) concentration leads to proteolytic cleavage at the C-terminal cytosolic region following the CNBH domain by altering its interaction with calmodulin. The observed Ca(2+)-induced proteolytic cleavage of EAG2 channel may act as an adaptive response under physiological and/or pathological conditions.

Differential contribution of Kv4-containing channels to A-type, voltage-gated potassium currents in somatic and visceral dorsal root ganglion neurons.

Little is known about electrophysiological differences of A-type transient K(+) (KA) currents in nociceptive afferent neurons that innervate somatic and visceral tissues. Staining with isolectin B4 (IB4)-FITC classifies L6-S1 dorsal root ganglion (DRG) neurons into three populations with distinct staining intensities: negative to weak, moderate, and intense fluorescence signals. All IB4 intensely stained cells are negative for a fluorescent dye, Fast Blue (FB), injected into the bladder wall, whereas a fraction of somatic neurons labeled by FB, injected to the external urethral dermis, is intensely stained with IB4. In whole-cell, patch-clamp recordings, phrixotoxin 2 (PaTx2), a voltage-gated K(+) (Kv)4 channel blocker, exhibits voltage-independent inhibition of the KA current in IB4 intensely stained cells but not the one in bladder-innervating cells. The toxin also shows voltage-independent inhibition of heterologously expressed Kv4.1 current, whereas its inhibition of Kv4.2 and Kv4.3 currents is voltage dependent. The swapping of four amino acids at the carboxyl portion of the S3 region between Kv4.1 and Kv4.2 transfers this characteristic. RT-PCRs detected Kv4.1 and the long isoform of Kv4.3 mRNAs without significant Kv4.2 mRNA in L6-S1 DRGs. Kv4.1 and Kv4.3 mRNA levels were higher in laser-captured, IB4-stained neurons than in bladder afferent neurons. These results indicate that PaTx2 acts differently on channels in the Kv4 family and that Kv4.1 and possibly Kv4.3 subunits functionally participate in the formation of KA channels in a subpopulation of somatic C-fiber neurons but not in visceral C-fiber neurons innervating the bladder.

AtRbohF contributes to non-host resistance to Magnaporthe oryzae in Arabidopsis.

The infection hyphae in pen2 atrbohF plants were significantly longer than those in pen2 plants. The proportion of branched hyphae development in the pen2 atrbohF plants was significantly higher than in the pen2 plants. These results suggest that AtRbohF is probably involved in post-penetration resistance to Magnaporthe oryzae in Arabidopsis.

Non-host resistance to penetration and hyphal growth of Magnaporthe oryzae in Arabidopsis.

Rice blast caused by Magnaporthe oryzae is a devastating disease of rice. Mechanisms of rice resistance to blast have been studied extensively, and the rice-M. oryzae pathosystem has become a model for plant-microbe interaction studies. However, the mechanisms of non-host resistance (NHR) to rice blast in other plants remain poorly understood. We found that penetration resistance to M. oryzae in multiple mutants, including pen2 NahG pmr5 agb1 and pen2 NahG pmr5 mlo2 plants, was severely compromised and that fungal growth was permitted in penetrated epidermal cells. Furthermore, rice Pi21 enhanced movement of infection hyphae from penetrated Arabidopsis epidermal cells to adjacent mesophyll cells. These results indicate that PEN2, PMR5, AGB1, and MLO2 function in both penetration and post-penetration resistance to M. oryzae in Arabidopsis, and suggest that the absence of rice Pi21 contributed to Arabidopsis NHR to M. oryzae.

Effects of bladder outlet obstruction on properties of Ca2+-activated K+ channels in rat bladder.

In this study, we investigated the effects of bladder outlet obstruction (BOO) on the expression and function of large conductance (BK) and small conductance (SK) Ca(2+)-activated K(+) channels in detrusor smooth muscle. The bladder from adult female Sprague-Dawley rats with 6-wk BOO were used. The mRNA expression of the BK channel alpha-subunit, beta1-, beta2-, and beta4-subunits and SK1, SK2, and SK3 channels were investigated using real-time RT-PCR. All subunits except for the BK-beta2, SK2, and SK3 channels were predominantly expressed in the detrusor smooth muscle rather than in the mucosa. The mRNA expression of the BK channel alpha-subunit was not significantly changed in obstructed bladders. However, the expression of the BK channel beta1-subunit and the SK3 channel was remarkably increased in obstructed bladders. On the other hand, the expression of the BK channel beta4-subunit was decreased as the severity of BOO-induced bladder overactivity progressed. In detrusor smooth muscle strips from obstructed bladders, blockade of BK channels by iberiotoxin (IbTx) or charybdotoxin (CTx) and blockade of SK channels by apamin increased the amplitude of spontaneous contractions. These blockers also increased the contractility and affinity of these strips for carbachol during cumulative applications. The facilitatory effects elicited by these K(+) channel blockers were larger in the strips from obstructed bladders compared with control bladders. These results suggest that long-term exposure to BOO for 6 wk enhances the function of both BK and SK types of Ca(2+)-activated K(+) channels in the detrusor smooth muscle to induce an inhibition of bladder contractility, which might be a compensatory mechanism to reduce BOO-induced bladder overactivity.

Intestinal parasitic infections and socioeconomic status in Prek Russey Commune, Cambodia.

OBJECTIVE: Possible relationships were explored between socioeconomic status (SES) and intestinal parasitic infections among residents of Prek Russey, a commune in a rural area near Phnom Penh. METHODS: An epidemiological study was conducted in Prek Russey Commune, consisting of the following three villages: Kroppeu Ha (KH), Prek Russey (PR), and Prek AngChanh (PA), with respective populations of 3,401, 1,350, and 1,044. Examinations of intestinal parasitic infections and the interview survey were performed. RESULTS: Among the residents of KH, PR, and PA, respectively, the prevalence rates of three major species of parasites were as follows: 17%, 27%, and 34% had hookworm; 14%, 17%, and 19% had Ascaris lumbricoides; and 13%, 17%, and 18% had Strongyloides stercoralis. The prevalence of each parasitic group tended to increase in the order of KH, PR, and PA. The greatest differences between the villages were in the prevalence of hookworm infection. Our epidemiological survey revealed three SES-related factors whose proportions increased or decreased in the order of KH, PR, and PA. First, the percentages of households purchasing relatively safe drinking water were 96%, 63%, and 2%, respectively. Second, whereas 82% and 78% of houses in KH and PR were equipped with a latrine, only 41% of houses in PA were so equipped. Third, the proportion of households whose main income was from farming increased in the order of KH, PR, and PA. CONCLUSION: The present study thus suggests that greater latrine use and less dependence on farming activity are related to a lower prevalence of intestinal parasitic infections, although the results are not conclusive due to the ecological nature of the study.