Enhanced Shear Force Responsiveness of Epithelial Na(+) Channel's (ENaC) δ Subunit Following the Insertion of N-Glycosylation Motifs Relies on the Extracellular Matrix.
Enhanced Shear Force Responsiveness of Epithelial Na(+) Channel's (ENaC) δ Subunit Following the Insertion of N-Glycosylation Motifs Relies on the Extracellular Matrix.
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DOI：
10.3390/ijms22052500
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发表时间：
2021-03-02
影响因子：
5.6
通讯作者：
Fronius M
中科院分区：
文献类型：
--
作者：
Barth D;Knoepp F;Fronius M
变性蛋白/上皮Na+通道（ENaC）蛋白家族的成员和细胞外细胞基质（ECM）形成机械敏感复合物。这种复合体的一个核心特征是连接通道与ECM的系链，然而，关于这些系链的性质的知识很少。α ENaC的N-聚糖最近被确定为潜在的系链，但N-聚糖是否作为机械感觉过程的普遍特征仍未得到解决。本研究的目的是揭示向对剪切力（SF）反应较低的δ ENaC添加N-聚糖是否会增加其SF反应性，以及这是否依赖于与ECM的连接。因此，通过定点突变引入N-糖基化基序，在非洲爪蟾卵母细胞中表达β和γ ENaC蛋白，并通过双电极电压钳测量SF激活电流。N-糖基化基序的插入增加了δ ENaC的SF反应性。在位置487处包含糖基化天冬酰胺（N）确实增加了分子量并提供了通道，其SF响应在ECM通过透明质酸酶降解后被消除。这表明N-聚糖的添加改善了SF-响应性，并且这种作用依赖于完整的ECM。这些发现进一步支持N-聚糖作为机械转导的系链的作用。
Members of the Degenerin/epithelial Na+ channel (ENaC) protein family and the extracellular cell matrix (ECM) form a mechanosensitive complex. A core feature of this complex are tethers, which connect the channel with the ECM, however, knowledge about the nature of these tethers is scarce. N-glycans of α ENaC were recently identified as potential tethers but whether N-glycans serve as a ubiquitous feature for mechanosensation processes remains unresolved. The purpose of this study was to reveal whether the addition of N-glycans to δ ENaC—which is less responsive to shear force (SF)—increases its SF-responsiveness and whether this relies on a linkage to the ECM. Therefore, N-glycosylation motifs were introduced via site-directed mutagenesis, the resulting proteins expressed with β and γ ENaC in Xenopus oocytes, and SF-activated currents measured by two-electrode voltage-clamp. The insertion of N-glycosylation motifs increases δ ENaC’s SF responsiveness. The inclusion of a glycosylated asparagine (N) at position 487 did increase the molecular mass and provided a channel whose SF response was abolished following ECM degradation via hyaluronidase. This indicates that the addition of N-glycans improves SF-responsiveness and that this effect relies on an intact ECM. These findings further support the role of N-glycans as tethers for mechanotransduction.
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影响因子：
4.8
作者：
Carattino, MD;Sheng, SH;Kleyman, TR
通讯作者：
Kleyman, TR
DOI：
10.1073/pnas.1911243117
发表时间：
2020-01-07
影响因子：
11.1
作者：
Knoepp, Fenja;Ashley, Zoe;Fronius, Martin
通讯作者：
Fronius, Martin
DOI：
10.1152/ajprenal.00195.2017
发表时间：
2018-03-01
影响因子：
4.2
作者：
Kashlan, Ossama B.;Kinlough, Carol L.;Kleyman, Thomas R.
通讯作者：
Kleyman, Thomas R.
影响因子：
3.4
作者：
COREY, DP;HUDSPETH, AJ
通讯作者：
HUDSPETH, AJ
DOI：
10.1002/neu.480241013
发表时间：
1993-10-01
期刊：
JOURNAL OF NEUROBIOLOGY
影响因子：
--
作者：
CHALFIE, M
通讯作者：
CHALFIE, M