Specific and essential but not sufficient roles of LRRC8A in the activity of volume-sensitive outwardly rectifying anion channel (VSOR)

Abstract

The broadly expressed volume-sensitive outwardly rectifying anion channel (VSOR, also called VRAC) plays essential roles in cell survival and death. Recent findings have suggested that LRRC8A is a core component of VSOR in human cells. In the present study, VSOR currents were found to be largely reduced by siRNA against LRRC8A in mouse C127 cells as well. In contrast, LRRC8A knockdown never affected activities of 4 other types of anion channel activated by acid, Ca 2C , patch excision or cAMP. While cisplatin-resistant KCP-4 cells poorly expressed endogenous VSOR activity, molecular expression levels of LRRC8A, LRRC8D and LRRC8E were indistinguishable between VSOR-deficient KCP-4 cells and the parental VSOR-rich KB cells. Furthermore, overexpression of LRRC8A alone or together with LRRC8D or LRRC8E in KCP-4 cells failed to restore VSOR activity. These results show that deficiency of VSOR currents in KCP-4 cells is not due to insufficient expression of the LRRC8A/D/ E gene, suggesting an essential involvement of some other factor(s), and indicate that further study is required to better understand the complexities of the molecular determinants of VSOR, including the precise role of LRRC8 proteins.

ATP Release Channels

“…Considering the various LRRC8 combinations and variable stoichiometry within a channel complex, LRRC8 proteins can potentially form numerous VRACs with both shared and distinct functions. It should also be noted that LRRC8s are essential but not sufficient components for the generation of VRAC currents, and additional factor(s) remain to be identified [ 204 ].…”

Cell Volume-Activated and Volume-Correlated Anion Channels in Mammalian Cells: Their Biophysical, Molecular, and Pharmacological Properties

“…A fifth criterion can also be proposed: channel activity exhibiting all of the phenotypic properties (see section II.B.1) must be reconstituted by incorporating the candidate protein(s) in the lipid bilayer. Although the essential component of VSOR has now been identified as LRRC8A plus LRRC8C/8D/8E (Voss et al, 2014), findings showing that this combinatory expression did not reproduce VSOR activity (Voss et al, 2014;Okada et al, 2017), that charge-modifying mutation of LRRC8A did not markedly alter the pore properties (Voss et al, 2014), and that VSOR-like activity produced by the reconstitution system lacked intracellular ATP dependence and inactivation kinetics (Syeda et al, 2016) fail to meet the above criteria 1, 4, and 5, respectively. Further investigation is thus required before the molecular identities of the main components of VSOR, which form the pore, are precisely determined.…”

Biophysics and Structure-Function Relationships of LRRC8-Formed Volume-Regulated Anion Channels

“…On the other hand, in a cellular context, there must be more than merely a reduction in intracellular ionic strength to activate VRACs. Further limiting factors are likely involved because the overexpression of functional LRRC8 heteromers did not increase swelling-induced chloride currents above wild-type levels ( 4 ), and different cells with similar LRRC8 expression levels showed differences in VRAC activity ( 73 ). In addition, VRACs can be activated under isovolumetric conditions, e.g., by cisplatin or sphingosine-1-phosphate ( 13 , 44 , 74 ), likely by mechanisms not involving changes in ionic strength.…”