Prolonged exposure to methylglyoxal causes disruption of vascular K_ATP channel by mRNA instability

Abstract

Diabetes mellitus is characterized by hyperglycemia and excessive production of intermediary metabolites including methylglyoxal (MGO), a reactive carbonyl species that can lead to cell injuries. Interacting with proteins, lipids, and DNA, excessive MGO can cause dysfunction of various tissues, especially the vascular walls where diabetic complications often take place. However, the potential vascular targets of excessive MGO remain to be fully understood. Here we show that the vascular Kir6.1/SUR2B isoform of ATP-sensitive K(+) (K(ATP)) channels is likely to be disrupted with an exposure to submillimolar MGO. Up to 90% of the Kir6.1/SUR2B currents were suppressed by 1 mM MGO with a time constant of ∼2 h. Consistently, MGO treatment caused a vast reduction of both Kir6.1 and SUR2B mRNAs endogenously expressed in the A10 vascular smooth muscle cells. In the presence of the transcriptional inhibitor actinomycin-D, MGO remained to lower the Kir6.1 and SUR2B mRNAs to the same degree as MGO alone, suggesting that the MGO effect is likely to compromise the mRNA stability. Luciferase reporter assays indicated that the 3'-untranslated regions (UTRs) of the Kir6.1 but not SUR2 mRNA were targeted by MGO. In contrast, the SUR2B mRNAs obtained with in vitro transcription were disrupted by MGO directly, while the Kir6.1 transcripts were unaffected. Consistent with these results, the constriction of mesenteric arterial rings was markedly augmented with an exposure to 1 mM MGO for 2 h, and such an MGO effect was totally eliminated in the presence of glibenclamide. These results therefore suggest that acting on the 3'-UTR of Kir6.1 and the coding region of SUR2B, MGO causes instability of Kir6.1 and SUR2B mRNAs, disruption of vascular K(ATP) channels, and impairment of arterial function.

The SUR2B subunit of rat vascular KATP channel is targeted by miR-9a-3p induced by prolonged exposure to methylglyoxal

“…D42145.1) and mouse SUR2B mRNA isoform (GenBank no. D86038, mRNA isoform NM_011511) were cloned and inserted into pcDNA3.1 (a eukaryotic expression vector), respectively, as previously described (44), which were named with Kir and SUR, respectively.…”

“…This in turn disrupts the signaling network in these cells, triggers structural remodeling of the vascular wall, propagates vascular inflammation, and causes vascular dysfunction (40 -42). Indeed, our recent studies have shown that an important molecular target of MGO in the vasculature is the ATP-sensitive K ϩ (K ATP ) channel (44).…”

“…In diabetic patients, K ATP channeldependent vasodilation is impaired, suggesting that K ATP channel dysfunction occurs under diabetic conditions (14,28). In our recent studies, we have found that a prolonged exposure to MGO brings about inhibition of the K ATP channel and augmentation of vasoconstriction responses (44). However, the underlying mechanisms for the MGO-induced inhibition of K ATP channel are still unclear.…”

“…Most miR binding sites sufficient for the transcript silencing are located in the 3=-untranslated region (3=-UTR), while some are in the coding sequence (CDS). Our previous studies have suggested that MGO acts on the CDS of SUR2B mRNA, impairing their stability as well as K ATP channel activity (44). Thus, it is possible that under diabetic conditions, the reactive carbonyl stress raises the expression of miRs that subsequently target the K ATP channel, and impair the vascular tone regulation.…”

S-Glutathionylation of Ion Channels: Insights into the Regulation of Channel Functions, Thiol Modification Crosstalk, and Mechanosensing

“…(7,27,29,77,84,110,111,136,138). K ATP channels are made of four pore-forming Kir (inward rectifier K + channel) subunits and four regulatory sulphonylurea receptors of the ATP-binding cassette (SUR) subunits (102 (137,138).…”