Plasma membrane targeting is essential for Rem-mediated Ca2+ channel inhibition.
The small GTPase Rem is a potent negative regulator of high voltage-activated Ca(2+) channels and a known interacting partner for Ca(2+) channel accessory beta subunits. The mechanism for Rem-mediated channel inhibition remains controversial, although it has been proposed that Ca(V)beta association is required. Previous work has shown that a C-terminal truncation of Rem (Rem-(1-265)) displays reduced in vivo binding to membrane-localized beta 2a and lacks channel regulatory function. In this paper, we describe a role for the Rem C terminus in plasma membrane localization through association with phosphatidylinositol lipids. Moreover, Rem-(1-265) can associate with beta 2a in vitro and beta 1b in vivo, suggesting that the C terminus does not directly participate in Ca(V)beta association. Despite demonstrated beta 1b binding, Rem-(1-265) was not capable of regulating a Ca(V)1.2-beta 1b channel complex, indicating that beta subunit binding is not sufficient for channel regulation. However, fusion of the CAAX domain from K-Ras4B or H-Ras to the Rem-(1-265) C terminus restored membrane localization and Ca(2+) channel regulation, suggesting that beta binding and membrane localization are independent events required for channel inhibition.
Departments of Molecular and Cellular Biochemistry.
Robert N
RN
Correll
Departments of Molecular and Cellular Biochemistry.
Chunyan
C
Pang
Departments of Molecular and Cellular Biochemistry.
Brian S
BS
Finlin
Departments of Molecular and Cellular Biochemistry.
Alexandria M
AM
Dailey
Departments of Physiology, University of Kentucky College of Medicine, Lexington, Kentucky 40536-0509.
Jonathan
J
Satin
Departments of Molecular and Cellular Biochemistry. Electronic address: dandres@uky.edu.
Douglas A
DA
Andres
D000818:Animals; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism
D020746:Calcium Channels, L-Type; C117307:KRAS protein, human; C419385:L-type calcium channel alpha(1C); D010716:Phosphatidylinositols; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; C108135:Rem protein, mouse; D020559:Monomeric GTP-Binding Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins