Preassembled GPCR signaling complexes mediate distinct cellular responses to ultralow ligand concentrations

G protein–coupled receptors (GPCRs) are the largest class of cell surface signaling proteins, participate in nearly all physiological processes, and are the targets of 30% of marketed drugs. Typically, nanomolar to micromolar concentrations of ligand are used to activate GPCRs in experimental systems. We detected GPCR responses to a wide range of ligand concentrations, from attomolar to millimolar, by measuring GPCR-stimulated production of cyclic adenosine monophosphate (cAMP) with high spatial and temporal resolution. Mathematical modeling showed that femtomolar concentrations of ligand activated, on average, 40% of the cells in a population provided that a cell was activated by one to two binding events. Furthermore, activation of the endogenous β2-adrenergic receptor (β2AR) and muscarinic acetylcholine M3 receptor (M3R) by femtomolar concentrations of ligand in cell lines and human cardiac fibroblasts caused sustained increases in nuclear translocation of extracellular signal–regulated kinase (ERK) and cytosolic protein kinase C (PKC) activity, respectively. These responses were spatially and temporally distinct from those that occurred in response to higher concentrations of ligand and resulted in a distinct cellular proteomic profile. This highly sensitive signaling depended on the GPCRs forming preassembled, higher-order signaling complexes at the plasma membrane. Recognizing that GPCRs respond to ultralow concentrations of neurotransmitters and hormones challenges established paradigms of drug action and provides a previously unappreciated aspect of GPCR activation that is quite distinct from that typically observed with higher ligand concentrations

Astro concepts : learning underlying physics principles in conceptual astronomy

Astro Concepts is a project within Swinburne University of Technology developing browser-based software modules on Optical Telescopes, Nebulae and Binary Stars. The modules are designed to enhance students' understanding of basic physics concepts, which underlie introductory-level conceptual astronomy courses. When complete, the Astro Concepts modules will be available for use in university courses in introductory astronomy, introductory physics teaching, secondary teaching and online astronomy education. The strategy outlined here is to obtain a reasonable level of understanding of the necessary physics concepts by presenting them embedded in relevant and interesting astronomy contexts, and by the use of an engaging educational approach requiring active learning by the student

Astro Concepts: Learning Underlying Physics Principles in Conceptual Astronomy

Astro Concepts is a project within Swinburne University of Technology in Melbourne developing browser-based software modules on Optical Telescopes, Nebulae and Binary Stars. The modules are designed to enhance students’ understanding of basic physics concepts, which underlie introductory-level conceptual astronomy courses. When complete, the Astro Concepts modules will be available for use in university courses in introductory astronomy, introductory physics teaching, secondary teaching and online astronomy education. The strategy outlined here is to obtain a reasonable level of understanding of the necessary physics concepts by presenting them embedded in relevant and interesting astronomy contexts, and by the use of an engaging educational approach requiring active learning by the student

Interaction with caveolin-1 modulates G protein coupling of the mouse beta3-adrenoceptor

Caveolins act as scaffold proteins in multiprotein complexes and have been implicated in signaling by G protein-coupled receptors. Studies using knock-out mice suggest that β(3)-adrenoceptor (β(3)-AR) signaling is dependent on caveolin-1; however, it is not known whether caveolin-1 is associated with the β(3)-AR or solely with downstream signaling proteins. We have addressed this question by examining the impact of membrane rafts and caveolin-1 on the differential signaling of mouse β(3a)- and β(3b)-AR isoforms that diverge at the distal C terminus. Only the β(3b)-AR promotes pertussis toxin (PTX)-sensitive cAMP accumulation. When cells expressing the β(3a)-AR were treated with filipin III to disrupt membrane rafts or transfected with caveolin-1 siRNA, the cyclic AMP response to the β(3)-AR agonist CL316243 became PTX-sensitive, suggesting Gα(i/o) coupling. The β(3a)-AR C terminus, SP(384)PLNRF(389)DGY(392)EGARPF(398)PT, resembles a caveolin interaction motif. Mutant β(3a)-ARs (F389A/Y392A/F398A or P384S/F389A) promoted PTX-sensitive cAMP responses, and in situ proximity assays demonstrated an association between caveolin-1 and the wild type β(3a)-AR but not the mutant receptors. In membrane preparations, the β(3b)-AR activated Gα(o) and mediated PTX-sensitive cAMP responses, whereas the β(3a)-AR did not activate Gα(i/o) proteins. The endogenous β(3a)-AR displayed Gα(i/o) coupling in brown adipocytes from caveolin-1 knock-out mice or in wild type adipocytes treated with filipin III. Our studies indicate that interaction of the β(3a)-AR with caveolin inhibits coupling to Gα(i/o) proteins and suggest that signaling is modulated by a raft-enriched complex containing the β(3a)-AR, caveolin-1, Gα(s), and adenylyl cyclase