A Pore Segment in DEG/ENaC Na+ Channels Snyder, Peter M.; Olson, Diane R.; Bucher, Daniel B.
The Journal of biological chemistry,
10/1999, Volume:
274, Issue:
40
Journal Article
Peer reviewed
Open access
DEG/ENaC Na+ channels have diverse functions, including Na+ absorption, neurotransmission, and sensory transduction. The ability of these channels to discriminate between different ions is critical ...for their normal function. Several findings suggest that DEG/ENaC channels have a pore structure similar to K+ channels. To test this hypothesis, we examined the accessibility of native and introduced cysteines in the putative P loop of ENaC. We identified residues that span a barrier that excludes amiloride as well as anionic and large methanethiosulfonate reagents from the pore. This segment contains a structural element ((S/G)CS) involved in selectivity of ENaC. The results are not consistent with predictions from the K+channel pore, suggesting that DEG/ENaC Na+ channels have a novel pore structure.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The epithelial Na + channel (ENaC) absorbs Na + across the apical membrane of epithelia. The activity of ENaC is controlled by its interaction with Nedd4; mutations that
disrupt this interaction ...increase Na + absorption, causing an inherited form of hypertension (Liddle's syndrome). Nedd4 contains an N-terminal C2 domain, a C-terminal
ubiquitin ligase domain, and multiple WW domains. The C2 domain is thought to be involved in the Ca 2+ -dependent localization of Nedd4 at the cell surface. However, we found that the C2 domain was not required for human Nedd4
(hNedd4) to inhibit ENaC in both Xenopus oocytes and Fischer rat thyroid epithelia. Rather, hNedd4 lacking the C2 domain inhibited ENaC more potently than wild-type
hNedd4. Earlier work indicated that the WW domains bind to PY motifs in the C terminus of ENaC. However, it is not known which
WW domains mediate this interaction. Glutathione S -transferase-fusion proteins of WW domains 2â4 each bound to α, β, and γENaC in vitro . The interactions were abolished by mutation of two residues. WW domain 3 (but not the other WW domains) was both necessary
and sufficient for the binding of hNedd4 to αENaC. WW domain 3 was also required for the inhibition of ENaC by hNedd4; inhibition
was nearly abolished when WW domain 3 was mutated. However, the interaction between ENaC and WW domain 3 alone was not sufficient
for inhibition. Moreover, inhibition was decreased by mutation of WW domain 2 or WW domain 4. Thus, WW domains 2â4 each participate
in the functional interaction between hNedd4 and ENaC in intact cells.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
We show here, using locust wholemount ganglia as an example, that scaling artifacts in three-dimensional reconstructions from confocal microscopic images due to refractive index mismatch in the light ...path and tissue shrinking, can account for dramatic errors in measurements of morphometric values. Refractive index mismatch leads to considerable alteration of the axial dimension, and true dimensions must be restored by rescaling the
Z-axis of the image stack. The appropriate scaling factor depends on the refractive indices of the media in the light path and the numerical aperture of the objective used and can be determined by numerical simulations, as we show here. In addition, different histochemical procedures were tested in regard to their effect on tissue dimensions. Reconstructions of scans at different stages of these protocols show that shrinking can be avoided prior to clearing when dehydrating ethanol series are carefully applied. Fixation and mismatching buffer osmolarity have no effect. We demonstrate procedures to reduce artifacts during mounting and clearing in methyl salicylate, such that only isometric shrinkage occurs, which can easily be corrected by rescaling the image dimensions. Glycerol-based clearing agents produced severe anisometric and nonlinear shrinkage and we could not find a way to overcome this.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Central pattern generators (CPGs) are circuits that generate organized and repetitive motor patterns, such as those underlying feeding, locomotion and respiration. We summarize recent work on ...invertebrate CPGs which has provided new insights into how rhythmic motor patterns are produced and how they are controlled by higher-order command and modulatory interneurons.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Reaching new depths in nuclear structure investigations requires new experimental equipment and new techniques of data analysis. The modern γ-ray spectrometers, like AGATA and GRETINA are now built ...of new-generation segmented germanium detectors. These most advanced detectors are able to reconstruct the trajectory of a γ-ray inside the detector. These are powerful detectors, but they need careful characterization, since their output signals are more complex. For instance for each γ-ray interaction that occurs in a segment of such a detector additional output signals (called proportional crosstalk), falsely appearing as an independent (often negative) energy depositions, are registered on the non-interacting segments. A failure to implement crosstalk correction results in incorrectly measured energies on the segments for two- and higher-fold events. It affects all experiments which rely on the recorded segment energies. Furthermore incorrectly recorded energies on the segments cause a failure to reconstruct the γ-ray trajectories using Compton scattering analysis. The proportional crosstalk for the iThemba LABS segmented clover was measured and a crosstalk correction was successfully implemented. The measured crosstalk-corrected energies show good agreement with the true γ-ray energies independent on the number of hit segments and an improved energy resolution for the segment sum energy was obtained.
The measurements of direct photons in 158 AGeV Pb+Pb collisions by the WA98 experiment at the CERN SPS are discussed. The methods used to determine the inclusive photon and the π
0 yields are ...discussed. The results of investigation of the systematic errors are presented.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
The epithelial Na(+) channel (ENaC) is comprised of three homologous subunits (alpha, beta, and gamma). The channel forms the pathway for Na(+) absorption in the kidney, and mutations cause disorders ...of Na(+) homeostasis. However, little is known about the mechanisms that control the gating of ENaC. We investigated the gating mechanism by introducing bulky side chains at a position adjacent to the extracellular end of the second membrane spanning segment (549, 520, and 529 in alpha, beta, and gammaENaC, respectively). Equivalent "DEG" mutations in related DEG/ENaC channels in Caenorhabditis elegans cause swelling neurodegeneration, presumably by increasing channel activity. We found that the Na(+) current was increased by mutagenesis or chemical modification of this residue and adjacent residues in alpha, beta, and gammaENaC. This resulted from a change in the gating of ENaC; modification of a cysteine at position 520 in betaENaC increased the open state probability from 0. 12 to 0.96. Accessibility to this side chain from the extracellular side was state-dependent; modification occurred only when the channel was in the open conformation. Single-channel conductance decreased when the side chain contained a positive, but not a negative charge. However, alterations in the side chain did not alter the selectivity of ENaC. This is consistent with a location for the DEG residue in the outer vestibule. The results suggest that channel gating involves a conformational change in the outer vestibule of ENaC. Disruption of this mechanism could be important clinically since one of the mutations that increased Na(+) current (gamma(N530K)) was identified in a patient with renal disease.
Which features of network output are well preserved during growth of the nervous system and across different preparations of the same size? To address this issue, we characterized the pyloric rhythms ...generated by the stomatogastric nervous systems of 99 adult and 12 juvenile lobsters (Homarus americanus). Anatomical studies of single pyloric network neurons and of the whole stomatogastric ganglion (STG) showed that the STG and its neurons grow considerably from juvenile to adult. Despite these changes in size, intracellularly recorded membrane potential waveforms of pyloric network neurons and the phase relationships in the pyloric rhythm were very similar between juvenile and adult preparations. Across adult preparations, the cycle period and number of spikes per burst were not tightly maintained, but the mean phase relationships were independent of the period of the rhythm and relatively tightly maintained across preparations. We interpret this as evidence for homeostatic regulation of network activity.
New collective structures in the 163Yb nucleus Sithole, M. A.; Sharpey Schafer, J. F.; Majola, S. N. T. ...
The European physical journal. A, Hadrons and nuclei,
2019/10, Volume:
55, Issue:
10
Journal Article
Peer reviewed
.
The
152
Sm(
16
O, 5n)
163
Yb reaction at a beam energy of 93 MeV was used to study the excited states of
163
Yb with the AFRODITE
γ
-ray spectrometer at iThemba LABS. The level scheme of
163
Yb has ...been extended and new rotational bands established. The band based on the ground-state has been extended from a spin of 11/2
-
to spin 43/2
-
. A high-
K
band based on the neutron 50511/2
-
Nilsson orbital has been observed and is reported for the first time in this work. Additional new states in
163
Yb were observed which all decay to the yrast band. Some of these states are placed in a sequence which is conjectured to be a
γ
band involving a coupling with the
i
13/2
6425/2
+
neutron orbital. The band structures are discussed with reference to Cranked Shell Model (CSM) calculations and a systematic comparison with the neighbouring nuclei.
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DOBA, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, SIK, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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