Pannexins are mammalian orthologs of the invertebrate gap junction proteins innexins and thus have been proposed to play a role in gap junctional intercellular communication. Localization of exogenously expressed pannexin 1 (Panx1) and pannexin 3 (Panx3), together with pharmacological studies, revealed a cell surface distribution profile and life cycle dynamics that were distinct from connexin 43 (Cx43, encoded by Gja1). Furthermore, N-glycosidase treatment showed that both Panx1 (~41-48 kD species) and Panx3 (~43 kD) were glycosylated, whereas N-linked glycosylation-defective mutants exhibited a decreased ability to be transported to the cell surface. Tissue surveys revealed the expression of Panx1 in several murine tissues -including in cartilage, skin, spleen and brain -whereas Panx3 expression was prevalent in skin and cartilage with a second higher-molecular-weight species present in a broad range of tissues. Tissue-specific localization patterns of Panx1 and Panx3 ranging from distinct cell surface clusters to intracellular profiles were revealed by immunostaining of skin and spleen sections. Finally, functional assays in cultured cells transiently expressing Panx1 and Panx3 were incapable of forming intercellular channels, but assembled into functional cell surface channels. Collectively, these studies show that Panx1 and Panx3 have many characteristics that are distinct from Cx43 and that these proteins probably play an important biological role as single membrane channels.

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Background: Pannexins functions in skeletal myogenesis are unknown.Results: Panx1 and Panx3 species are co-expressed in skeletal muscle. Their levels are modulated during myoblast differentiation regulating either myoblast proliferation and/or differentiation status.Conclusion: Panx1 and Panx3 channels are novel regulators of skeletal muscle myoblast differentiation and proliferation.Significance: Skeletal muscle development and health depend on functional pannexin channels.

Connexin43 (Cx43) has been reported to interact with caveolin (Cav)-1, but the role of this association and whether other members of the caveolin family bind Cx43 had yet to be established. In this study, we show that Cx43 coimmunoprecipitates and colocalizes with Cav-1 and Cav-2 in rat epidermal keratinocytes. The colocalization of Cx43 with Cav-1 was confirmed in keratinocytes from human epidermis in vivo. Our mutation and Far Western analyses revealed that the C-terminal tail of Cx43 is required for its association with Cavs and that the Cx43/Cav-1 interaction is direct. Our results indicate that newly synthesized Cx43 interacts with Cavs in the Golgi apparatus and that the Cx43/Cavs complex also exists at the plasma membrane in lipid rafts. Using overexpression and small interfering RNA approaches, we demonstrated that caveolins regulate gap junctional intercellular communication (GJIC) and that the presence of Cx43 in lipid raft domains may contribute to the mechanism modulating GJIC. Our results suggest that the Cx43/Cavs association occurs during exocytic transport, and they clearly indicate that caveolin regulates GJIC. INTRODUCTIONGap junctions are specialized intercellular membrane channels that allow direct transfer of molecules of Ͻ1000 Dalton to selectively pass from one cell to another (Sohl and Willecke, 2004;Laird, 2005). Gap junctional intercellular communication (GJIC) plays a critical role in the coordination of development, tissue function, and cell homeostasis (Segretain and Falk, 2004;Wei et al., 2004). Gap junction channels are composed of two hemichannels, termed connexons, each provided by one of the two contacting cells and each connexon is composed of six transmembrane proteins, called connexins (Cxs) (Laird, 2006). The large gene family encoding Cx comprises 21 members in human (Sohl and Willecke, 2004;Wei et al., 2004). Cx43 is the most abundant gap junction protein in a wide spectrum of tissues, including the epidermis (Kretz et al., 2004;Laird, 2006). Cx43 is most likely assembled into connexons in the trans-Golgi network and then traffics along microtubules to the plasma membrane where they diffuse laterally and aggregate into gap junction plaques (Saez et al., 2003;Martin and Evans, 2004;Segretain and Falk, 2004;Laird, 2006;Lauf et al. 2002), although one report suggests that Cx43 containing vesicles may be directly targeted to adherens junctions adjacent to gap junctions (Shaw et al., 2007). The function of existing gap junction channels can be controlled by their opening and closing, whereas the regulation of their delivery, assembly, and removal constitutes additional mechanisms to control GJIC (Saez et al., 2003;Segretain and Falk, 2004;Laird, 2006). The extent of intercellular coupling is thus finely regulated by the control of connexin channels present at the plasma membrane, their functional state, and their permeability. All of these properties are determined by the constituent proteins of the connexon and their interaction with other protein partners and lipids (Cascio, ...