Quantitative bioluminescence resonance energy transfer (BRET) analysis was applied to the study of ␤ 1 -and ␤ 2 -adrenergic receptor homo-and heterodimerization. To assess the relative affinity between each of the protomers, BRET saturation experiments were carried out in HEK-293T cells. ␤ 1 -and ␤ 2 -adrenergic receptors were found to have similar propensity to engage in homo-and heterotropic interactions suggesting that, at equivalent expression levels of the two receptor subtypes, an equal proportion of homo-and heterodimers would form. Analysis of the data also revealed that, at equimolar expression levels of energy donor and acceptor, more than 80% of the receptor molecules exist as dimers and that this high incidence of receptor dimerization is insensitive to receptor density for expression levels varying between 1.4 and 26.9 pmol of receptor/mg of membrane protein. Taken together, these results indicate that most of the receptors expressed in cells exist as constitutive dimers and that, at least in undifferentiated fibroblasts, the proportion of homo-and heterodimers between the closely related ␤ 1 -and ␤ 2 -adrenergic receptors is determined by their relative levels of expression. G protein-coupled receptors (GPCRs)1 represent the largest family of transmembrane receptors involved in cell signaling. In the past few years, many studies indicated that GPCR dimerization can occur between two identical receptors (homodimerization), between two different receptor subtypes of the same family, or even between receptors that are only distantly related (heterodimerization) (for a review, see Refs. 1 and 2). In most instances, co-immunoprecipitation was used as the primary experimental evidence supporting the existence of such dimers. More recently, however, light resonance energy transfer techniques such as fluorescence and bioluminescence resonance energy transfer (FRET and BRET) were also used. These "non-invasive" proximity-based assays confirmed that GPCR dimerization does not represent biochemical artifacts due to receptor solubilization and can occur in living cells. They have been used to demonstrate homodimerization of the ␤ 2 -adrenergic (3), the yeast alpha mating factor (4), the SST5 somatostatin (5), the gonadotropin releasing hormone (6), the luteinizing hormone (7), the ␦-opioid (8), the thyrotropin-releasing hormone (9), the cholecystokinin (10), and the melatonin (11) receptors as well as heterodimerization between somatostatin receptor subtypes (5), somatostatin and dopamine receptors (12), melatonin receptor subtypes (11), and opioid receptor subtypes (13).An advantage of BRET and FRET over co-immunoprecipitation approaches lies in the more quantitative nature of the assay. However, relatively few studies exploited this quantitative potential for the study of GPCR dimerization. For the melatonin receptors, Ayoub et al. (11) recently used BRET competition assays to determine that the transfer of energy resulted from the formation of dimers and not of higher order oligomers. They also showed that...

Although homodimerization has been demonstrated for a large number of G protein-coupled receptors (GPCRs), no general role has been attributed to this process. Because it is known that oligomerization plays a key role in the quality control and endoplasmic reticulum (ER) export of many proteins, we sought to determine if homodimerization could play such a role in GPCR biogenesis. Using the ␤2-adrenergic receptor (␤2AR) as a model, cell fractionation studies revealed that receptor homodimerization is an event occurring as early as the ER. Supporting the hypothesis that receptor homodimerization is involved in ER processing, ␤2AR mutants lacking an ER-export motif or harboring a heterologous ER-retention signal dimerized with the wild-type receptor and inhibited its trafficking to the cell surface. Finally, in addition to inhibiting receptor dimerization, disruption of the putative dimerization motif, 276 GXXXGXXXL 284, prevented normal trafficking of the receptor to the plasma membrane. Taken together, these data indicate that ␤2AR homodimerization plays an important role in ER export and cell surface targeting.Recent studies have established that G protein-coupled receptors (GPCRs), 1 much like other membrane receptors, form both homodimers and heterodimers (1, 2). Various functions have been attributed to the dimerization between different receptor subtypes. For instance, heterodimerization has been proposed to promote the formation of receptors with unique pharmacological properties, contributing to the pharmacological diversity of GPCRs (3-5). Changes in G protein coupling specificity (6) as well as altered receptor endocytosis (3, 4, 6 -9) have also been proposed to result from receptor heterodimerization. For the metabotropic GABAb receptor, heterodimerization between GABAb-R1 and GABAb-R2 is an obligatory step for cell surface expression of a functional receptor (10,11). This was shown to result from the masking of an endoplasmic reticulum (ER) retention signal found in the C terminus of GABAb-R1, presumably through the formation of a coiled-coil domain between the C termini of the two receptor subtypes (12). The obligatory nature of this heterodimerization for cell surface trafficking and signaling was considered for a long time as a particularity of the GABAb receptor, which belongs to a GPCR subfamily (family 3) containing only a few members (13). However, a recent study indicated that the heterodimerization between two closely related members of family 1 GPCRs, the adrenergic ␣ 1D and ␣ 1B receptors, may be a prerequisite for the proper cell surface expression of the ␣ 1D subtype in both experimental models and real tissues (14).So far, despite the numerous examples of receptor homodimerization reported in the literature (15, 16), the biological function of this phenomenon has remained elusive, probably because it is extremely difficult to experimentally differentiate the dimer properties of identical receptor subtypes from those of monomers. However, a growing number of observations indicates that GPC...

␤ 1 -and ␤ 2 -adrenergic receptors (␤ 1 AR and ␤ 2 AR) are co-expressed in numerous tissues where they play a central role in the responses of various organs to sympathetic stimulation. Although the two receptor subtypes share some signaling pathways, each has been shown to have specific signaling and regulatory properties. Given the recent recognition that many G protein-coupled receptors can form homo-and heterodimers, the present study was undertaken to determine whether the ␤ 1 AR and ␤ 2 AR can form dimers in cells and, if so, to investigate the potential functional consequences of such heterodimerization. Using co-immunoprecipitation and bioluminescence resonance energy transfer, we show that ␤ 1 AR and ␤ 2 AR can form heterodimers in HEK 293 cells co-expressing the two receptors. Functionally, ␤-adrenergic stimulated adenylyl cyclase activity was found to be identical in cells expressing ␤ 1 AR, ␤ 2 AR, or both receptors at similar levels, indicating that heterodimerization did not affect this signaling pathway. When considering ERK1/2 MAPK activity, a significant agonistpromoted activation was detected in ␤ 2 AR-but not ␤ 1 AR-expressing cells. Similarly to what was observed in cells expressing the ␤ 1 AR alone, no ␤-adrenergic stimulated ERK1/2 phosphorylation was observed in cells co-expressing the two receptors. A similar inhibition of agonist-promoted internalization of the ␤ 2 AR was observed upon co-expression of the ␤ 1 AR, which by itself internalized to a lesser extent. Taken together, our data suggest that heterodimerization between ␤ 1 AR and ␤ 2 AR inhibits the agonist-promoted internalization of the ␤ 2 AR and its ability to activate the ERK1/2 MAPK signaling pathway.

The stability of buoyant-thermocapillary-driven flows in a fluid layer subjected to a horizontal temperature gradient is analysed. Our purpose is the modelization of recent experimental results obtained for a fluid of Prandtl number Pr=7, by Daviaud and Vince [Phys. Rev. E, 4432 (1993)] who observed a transition between traveling waves and stationary rolls when the height of fluid is increased. Our model takes into account several effects which were examined separately in previous studies. The relative importance of buoyancy and thermocapillarity is controlled by the ratio W of Marangoni number to Rayleigh number. The fluid layer is bounded below by a rigid plane whose temperature varies linearly along the direction of the thermal gradient. A Biot number is introduced to describe heat transfer at the top free surface. Our stability analysis establishes the existence of a transition between stationary and oscillatory modes when W exceeds a value W0 which is function of the Biot number. Moreover, two types of oscillatory modes have been identified which differ by the range of variation of their critical parameters (wave number, frequency, angle of propagation) versus W .

We extend our lattice model of gel electrophoresis (and diffusion) to the study of the continuum limit (no lattice effect) for hard spherical particles migrating in periodic and random threedimensional gels. To reach the continuum limit, the mesh size of the system is progressively decreased until a clear extrapolation to the continuum limit (where the size of the lattice parameter is infinitely small in comparison to the size of the gel fibers) can be done. Various types of gel are studied, starting from simple periodic parallel and straight fibers to a representation of a more realistic gel formed of irregular, cross-linked fibers placed along random directions. Our mobility data for pseudospherical particles in the continuum limit are interpreted in terms of the obstruction model of diffusion.