Recombinant tandem of pore-domains in a Weakly Inward rectifying K+ channel 2 (TWIK2) forms active lysosomal channels

Abstract

Recombinant TWIK2 channels produce weak basal background K+ currents. Current amplitudes depend on the animal species the channels have been isolated from and on the heterologous system used for their re-expression. Here we show that this variability is due to a unique cellular trafficking. We identified three different sequence signals responsible for the preferential expression of TWIK2 in the Lamp1-positive lysosomal compartment. Sequential inactivation of tyrosine-based (Y308ASIP) and di-leucine-like (E266LILL and D282EDDQVDIL) trafficking motifs progressively abolishes the targeting of TWIK2 to lysosomes, and promotes its functional relocation at the plasma membrane. In addition, TWIK2 contains two N-glycosylation sites (N79AS and N85AS) on its luminal side, and glycosylation is necessary for expression in lysosomes. As shown by electrophysiology and electron microscopy, TWIK2 produces functional background K+ currents in the endolysosomes, and its expression affects the number and mean size of the lysosomes. These results show that TWIK2 is expressed in lysosomes, further expanding the registry of ion channels expressed in these organelles.

Mutation of a single residue promotes gating of vertebrate and invertebrate two-pore domain potassium channels

“…Insets for hTASK1, mTRAAK, mTRESK, and hTWIK1 AA represent wild-type channel currents at a reduced scale. rTWIK2 LY and hTHIK2 5RA harbor additional mutations in intracellular trafficking signals that allow increased surface expression 10,52 . Each point represents the mean ± standard error of the mean, numbers in parentheses represent the number of oocytes tested for each condition.…”

“…10 , and rTWIK2-LY in ref. 52 . For mammalian cell electrophysiology, K2P channel cDNA sequences were inserted into the pIRES2-EGFP vector.…”

The TWIK2 Potassium Efflux Channel in Macrophages Mediates NLRP3 Inflammasome-Induced Inflammation

“…In our model, TWIK2 regulates cell immunity by expression in the plasma membrane as a K + efflux channel responsible for the ATP-induced NLRP3 inflammasome activation and therefore participates in the regulation of inflammation induced by PAMPs. Another recent study has shown that TWIK2 generates background K + currents in endolysosomes and thus regulates the number and size of lysosomes (Bobak et al, 2017). TWIK2 contains sequence signals responsible for the expression of TWIK2 in the Lamp1-positive lysosomal compartment (Bobak et al, 2017), and sequential inactivation of these trafficking motifs abolishes the targeting of TWIK2 to lysosomes, thus promoting functional relocation of the channel to the plasma membrane (Bobak et al, 2017).…”

A Mathematical Model of Lysosomal Ion Homeostasis Points to Differential Effects of Cl− Transport in Ca2+ Dynamics

“…While the transport of Na + is often coupled to that of metabolites and can additionally be mediated by the Na + -conductance of TPCs [40,41], several K + -specific channels have recently been identified on lysosomes. These comprise TMEM175 [42], Slo1/BK channels [24,26] and TWIK2 [43]. While efflux of monovalent cations can support the acidification of lysosomes [39], the presence of TMEM175 is important for pH stability under starving conductions in RAW 264.7 macrophages [42].…”