Interaction of the plasma membrane Ca2+ pump 4b/CI with the Ca2+/calmodulin-dependent membrane-associated kinase CASK.
Spatial and temporal regulation of intracellular Ca(2+) is a key event in many signaling pathways. Plasma membrane Ca(2+)-ATPases (PMCAs) are major regulators of Ca(2+) homeostasis and bind to PDZ (PSD-95/Dlg/ZO-1) domains via their C termini. Various membrane-associated guanylate kinase family members have been identified as interaction partners of PMCAs. In particular, SAP90/PSD95, PSD93/chapsyn-110, SAP97, and SAP102 all bind to the C-terminal tails of PMCA "b" splice variants. Additionally, it has been demonstrated that PMCA4b interacts with neuronal nitric-oxide synthase and that isoform 2b interacts with Na(+)/H(+) exchanger regulatory factor 2, both via a PDZ domain. CASK (calcium/calmodulin-dependent serine protein kinase) contains a calmodulin-dependent protein kinase-like domain followed by PDZ, SH3, and guanylate kinase-like domains. In adult brain CASK is located at neuronal synapses and interacts with various proteins, e.g. neurexin and Veli/LIN-7. In kidney it is localized to renal epithelia. Surprisingly, interaction with the Tbr-1 transcription factor, nuclear transport, binding to DNA T-elements (in a complex with Tbr-1), and transcriptional competence has been shown. Here we show that the C terminus of PMCA4b binds to CASK and that both proteins co-precipitate from brain and kidney tissue lysates. Immunofluorescence staining revealed co-expression of PMCA, CASK, and calbindin-d-28K in distal tubuli of rat kidney sections. To test if physical interaction of both proteins results in functional consequences we constructed a T-element-dependent reporter vector and investigated luciferase activity in HEK293 lysates, previously co-transfected with PMCA4b expression and control vectors. Expression of wild-type PMCA resulted in an 80% decrease in T-element-dependent transcriptional activity, whereas co-expression of a point-mutated PMCA, with nearly eliminated Ca(2+) pumping activity, had only a small influence on regulation of transcriptional activity. These results provide evidence of a new direct Ca(2+)-dependent link from the plasma membrane to the nucleus.
Schuh
Kai
K
Department of Medicine, University of Wuerzburg, D-97080 Wuerzburg, Germany.
Uldrijan
Stjepan
S
Gambaryan
Stepan
S
Roethlein
Nicola
N
Neyses
Ludwig
L
D000818:Animals; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D000252:Calcium-Transporting ATPases / Q000737:chemistry* / Q000378:metabolism; D027682:Cation Transport Proteins; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors / Q000378:metabolism; D051528:Guanylate Kinases; D006801:Humans; D007150:Immunohistochemistry; D007668:Kidney / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009399:Nephrons / Q000378:metabolism; D009703:Nucleoside-Phosphate Kinase / Q000737:chemistry* / Q000378:metabolism*; D053499:Plasma Membrane Calcium-Transporting ATPases; D011233:Precipitin Tests; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D051381:Rats; D014162:Transfection
D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D020033:Protein Isoforms; D008156:Luciferases; C106868:CASK kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D009703:Nucleoside-Phosphate Kinase; D051528:Guanylate Kinases; D053499:Plasma Membrane Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases