title: Intracellular calmodulin availability accessed with two-photon cross-correlation.
abstract: The availability and interactions of signaling proteins are tightly regulated in time and space to produce specific and localized effects. For calmodulin (CaM), a key transducer of intracellular Ca(2+) signaling, binding to its variety of targets initiates signaling cascades and regulates its subcellular localization, thereby making it unavailable for subsequent binding interactions. Among CaM's numerous targets, Ca(2+)/CaM-dependent protein kinase II is one of the most striking due to its unique ability to increase its affinity for CaM by autophosphorylation and to translocate when bound to Ca(2+)/CaM. Two-photon fluorescence correlation spectroscopy and cross-correlation spectroscopy were utilized to compare mobility and molecular interactions between CaM and Ca(2+)/CaM-dependent protein kinase II in solution and in living cells. These techniques revealed that CaM availability in cells could be altered by a change in intracellular conditions. Two-photon fluorescence cross-correlation spectroscopy exemplifies a generally applicable approach for studying protein-protein interactions in living cells that allows access to the behavior of signaling molecules within their native environment to probe for heterogeneities in signaling pathways in different cellular compartments.
authors.affiliation: Department of Neurobiology and Anatomy, University of Texas Health Science Center, Houston, TX 77030-1501, USA.
authors.forename: Sally A
authors.initials: SA
authors.lastname: Kim
authors.forename: Katrin G
authors.initials: KG
authors.lastname: Heinze
authors.forename: M Neal
authors.initials: MN
authors.lastname: Waxham
authors.forename: Petra
authors.initials: P
authors.lastname: Schwille
mesh_terms: D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D020013:Calcium Signaling; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017785:Photons; D011485:Protein Binding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence
chemical_list: D002147:Calmodulin; D011993:Recombinant Fusion Proteins; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases