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doi:10.1016/j.pb

Progress in Biophysics and Molecular Biology 94 (2007) 1–4

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Editorial

Gap junction channels: From protein genes to diseases

In animal tissues, most cells are connected via intercellular cytoplasmic channels clustered in plasmamembrane spatial microdomains termed gap junctions, which allow cells to directly exchange ions and smallmolecules. Each channel results from the docking of two half channels, which are hexameric torus ofjunctional proteins around an aqueous pore. All junctional channels have a similar overall structure but,unlike many other membrane channels, different gene families encode the membrane proteins that form themin different animal phyla. Gap junction structure and functions were for a long time mainly investigated in thevertebrates, where they were thought to be formed solely by connexins (Cxs). Then, in different invertebratesthat have no Cx genes, gap junctions were shown to be composed of another gene family, the innexins (Inxs,invertebrate analogues of Cxs), which have no sequence homology to Cxs. These proteins were at firstconsidered to be specific to invertebrate gap junctions but sequences with low similarity to the Inxs were lateralso identified in vertebrate chordates, leading some authors to suggest to rename this protein family‘‘pannexins’’(Panx) (from the Greek ‘‘pan’’, meaning all, entire, and nexus, connection). The present issue ofProgress in Biophysics and Molecular Biology, for which I have the honour to be Guest Editor, is designed tosummarise some of the new information on the characteristics, properties and roles of gap junction channel-forming proteins and on some consequences of their dysfunctions.

Cx, Inx and Panx proteins display the same overall topology, with four a-helical transmembrane domainsconnected by two extracellular loops and a single-cytoplasmic loop, both N- and C-termini being intracellular,but Cxs on the one hand, Inxs and Panxs on the other hand share virtually no similarity in the primary aminoacid sequence. It is not yet clear whether Inxs and Panxs are members of the same superfamily. Yen and Saier(2007) have used statistical, topological and conserved sequence motif analyses to examine this hypothesis andconclude that Inxs and Panxs arose from a single-precursor peptide. They suggest to use the term‘‘superfamily’’ to describe the homologues of Inxs because of the large number of constituent members, andbecause of their extensive sequence divergence.

A unique aspect of gap junction channels is that they are composed of the end-to-end docking, in thenarrow intercellular ‘‘gap’’, of two hemichannels called connexons, each formed by six Cx subunits that arecomposed of four hydrophobic transmembrane segments designated M1–M4 from the N- to the C-terminus.The expression of multiple connexins in the same cell type, the multiplicity of isoforms, and the ability to formhomomeric and heteromeric connexons, as well as homotypic and heterotypic channels, likely providesexquisite ‘‘functional tuning’’ of this family of membrane channels. Kovacs et al. (2007) have surveyed the gapjunction literature of data obtained by mutagenesis, biochemical, dye-transfer and electrophysiologicalstudies, combined with computational analysis, to provide an up to date depiction of the molecular structureof the gap junction channel.

It is now obvious that gap junction proteins do not operate as free-floating entities in the plasma membranebut interact with specific cytoplasmic proteins that link them to the cytoskeleton and to intracellular signal-transduction pathways. Such coassembly into multiprotein complexes is likely to be important forimmobilization and clustering of the cell-to-cell channels, for correct targeting of channels to specificsubcellular sites, for the ability of channels to funnel ions and small molecules, and for modulation of thechannel functions by kinases, phosphatases, and other regulatory proteins. Conversely, gap junction proteinsmay also, through their different partners, be involved in cell functions very different from their classical

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cell-to-cell tunnel-forming functions. Herve et al. (2007) present the diversity of interactions of gap junctionalproteins with protein partners and their potential importance.

Given their architecture, gap junction channels are subject to the influence of two types of voltage, thatbetween the intracellular and extracellular media (i.e. the membrane potential or Vm), and that between thetwo cell interiors (termed transjunctional voltage or Vj), which influence both pore permeation and channelgating. Gonzalez et al. (2007) provide an up to date and precise picture of the molecular and structural aspectsof how Vj and Vm are sensed, and how they therefore control channel opening and closing. Mutagenicstrategies coupled with structural, biochemical and electrophysical approaches are providing significantinsights into how distinct forms of voltage dependence are brought about, showing that gap junction channelscan undergo transitions between multiple conductance states driven by distinct voltage-gating mechanisms.

Phosphorylation, a widespread post-translational modification of proteins, is a primary means of mediatingsignal-transduction events that control numerous cellular processes via a highly regulated dynamic interplayof protein kinases and protein phosphatases. Cxs have been intensely studied, and most of their isoforms areknown to be phosphorylable, leading to modifications in tyrosine, serine, and threonine residues, which havebeen reported to affect the regulation of gap junctional communication at several stages of the Cx lifecycle,including intracellular Cx trafficking, connexon assembly and disassembly, Cx degradation as well as thegating of gap junction channels, but the underlying mechanisms remain poorly understood. Moreno and Lau(2007) cover some of the current, relevant research that attempt to explain how phosphorylation triggers and/or modulates gap junction channel gating.

Cx channels form pores sufficiently wide to be permeable to a wide variety of cytoplasmic molecules; there isevidence for permeability through at least some types of Cx channels of virtually all soluble secondmessengers, amino acids, nucleotides, calcium ions, and glucose and its metabolites. It was for a long timeassumed that limiting pore diameter and perhaps a slight charge preference would be the determinants ofmolecular permeability before it appeared that the exchange of inorganic ions and small metabolites betweenadjacent cells was in fact dynamically regulated. Such regulation is a critical feature that ensures coordinatedcellular activity and tissue homeostasis during both development and adult life in multicellular organisms.Harris (2007) describes and evaluates the various methods used to compare the permeabilities of a range ofcytoplasmic molecules through a variety of Cx channels, presents an annotated compilation of the results, anddiscusses the findings in the context of what can be reasonably inferred about mechanisms of selectivity.

Gap junctions play a pivotal role for the velocity and the safety of impulse propagation in cardiac tissues, thatcan be regarded as three-dimensional networks of coupled excitable elements. In such networks, the velocity andsafety of the spread of excitation is dependent on both active and passive properties of the individual elementsand on the connectivity of the network. Among passive properties, gap junctions play a pivotal role because theyultimately determine how much depolarizing current passes from excited to non-excited regions of the network.The distribution of gap junctions influences velocity and safety of the conduction of the electrical impulsethrough the myocardium and is responsible for its anisotropic properties. Over the last decades, anisotropy hasbeen increasingly recognised as a potential substrate for abnormal rhythms and reentry, both in normal as wellas pathological conditions. Valderrabano overviews the mechanisms of anisotropy, its structural and functionaldeterminants, and its roles in normal and abnormal propagation.

Transcriptome represents the very small percentage of the genetic code that is transcribed into RNA molecules;as each gene may produce many different types of mRNA molecules, the transcriptome is much more complexthan the genome that encodes it and can vary with external environmental conditions. The transcriptome analysisallows to determine when and where a gene is turned on or off in various types of cells and tissues. More than halfof the approximately 20 known mammalian Cxs are expressed in the central nervous system either duringdevelopment or in the mature brain, and ablation of Cx43, the most abundant gap junction protein, had been seento significantly affect the expression level and variability of an enormous number of functionally diverse genes.Iacobas et al. (2007) present evidence that the brain transcriptome contains Cx-dependent regulomes thatencompass genes in multiple cohorts, where the regulome is defined as the set of genes significantly regulated in agiven pathological or experimental sample with respect to the physiological/control sample.

Given the rich potential for regulation of junctional permeability, and directional and molecular gating, gapjunctional communication plays a crucial role in the regulation of information flow that takes place inembryonic events such as the patterning of the embryonic left–right axis, as well as the morphogenesis of the

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heart and limb, for example. During pre-implantation development, several Cxs are already expressed butlocated within the cytoplasm, then moved to the plasma membrane for assembly into gap junctions atcompaction, in the eight-cell stage. The communication pathways through junctional channels appearimportant for post-implantation development but we are only beginning to glimpse the elusive, subtle, yetcrucial molecular cross-talk between the interior machinery of cells. Levin (2007) summarizes the currentknowledge of the involvement of gap junctional communication in the patterning of both vertebrate andinvertebrate systems and discusses in detail several morphogenetic systems in which the properties of thissignalling pathway have been molecularly characterized.

The direct intercellular communication networks that link, connect and coordinate attached cells are foundnot only in compact tissues and organs but also in the various cell types of the immune system comprising thelymphoid organs as well as peripheral lymphocytes migrating through the blood and lymphatic networks.Multiple cell types of the immune system have indeed been shown to express Cxs and evidence has built upthat direct intercellular cross-talk via gap junction channels occurs in the haemopoietic and immune systems,implying important physiological consequences. Cx43-formed channels were recently shown to allow theintercellular diffusion of linear peptides with a molecular weight of up to 1800, the range of manyimmunologically relevant peptides. Such peptide transfer from infected or malignant cells to neighbouringcells may for example cause recognition of viral peptides or tumor-specific antigens by cytotoxic Tlymphocytes, the killers for the immune system. Neijssen et al. (2007) describe how innocent cells can acquirepeptides from infected neighbours, a process called ‘cross-presentation’ and its functional importance.

Direct communication between cells via gap junctional channels is recognised as an indispensablemechanism in the maintenance of cellular homeostasis. Junctional channels have been proposed to propagatecell death and survival-modulating signals, and hemichannels to serve as paracrine conduits to spread factorsthat modulate the fate of the surrounding cells, but the role of gap junctions in cell proliferation and cell deathnow appears to exceed their primary function as intercellular conveyers of essential homeostasis regulators.Cxs have indeed be shown to be present in cell structures other than the plasma membrane, including the cellnucleus, where Cx43 has been suggested to influence cell growth and differentiation, and mitochondria, whereits presence would be important for certain forms of cardioprotection. Antonio Rodrıguez-Sinovas et al.(2007) review the different ways by which Cx43 may influence cell death and survival.

The different channels, receptors and transporters present in the membranes frequently influence each othereither directly (e.g. through direct protein–protein interactions or via an intermediate partner protein) orindirectly (one of them may for example mediate the transport of a molecule regulating the activity of the secondor influence the membrane insertion of the second). Such mutual interactions may have important functionalconsequences on cellular homeostasis and functions. Chanson et al. (2007) summarise the evidence obtained sofar on such interactions between gap junction proteins and membrane channels/transporters and discuss possiblefunctional implications as well as what is known about the molecular basis of these interactions.

Besides their canonical function of intercellular tunnel-forming structures and their influence on membranechannels and transporters, connexins or their fragments (either alone or as components of a nexus complex)appear to be also involved in other processes, including growth control and development. Kardami et al.(2007) provide a brief overview of current thinking on the role of Cxs (particularly of Cx43) in growthregulation, more particularly on their ability to exert extensive effects on gene expression, especially on theexpression of growth-affecting genes. Up to now, at least two distinct mechanisms appear to be involved, oneby cell-to-cell transfer of permeable metabolites carrying genetic information, the second through the possiblebinding (and the subsequent sequestration) of molecules with transcriptional activity by Cxs.

I wish to thank all authors and co-authors for their commitment and the anonymous reviewers whocontributed by their critical constructive remarks to the excellence of this issue.

References

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Gonzalez, D., Gomez-Hernandez, J.M., Barrio, L.C., 2007. Molecular basis of voltage dependence of connexin channels: an integrative

appraisal. Prog. Biophys. Mol. Biol. 94, 65–105.

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Harris, A.L., 2007. Connexin channel permeability to cytoplasmic molecules. Prog. Biophys. Mol. Biol. 94, 119–142.

Herve, J.C., Bourmeyster, N., Sarrouilhe, D., Duffy, H.S., 2007. Gap junctional complexes: from partners to functions. Prog. Biophys.

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5–14.

Jean-Claude HerveEquipe ‘‘Interactions et Communications Cellulaires,’’ IPBC UMR 6187 CNRS-Universite de Poitiers, PBS, 40

Avenue du R. Pineau, 86022 Poitiers Cedex, France

E-mail address: Jean.Claude.Herve@univ-poitiers.fr