Discriminating the Message
In cardiac electrophysiology, gap junctions are often conceptualized as passive resistors that allow for electrical charge to move between cells. From that standpoint, gap junctions seem like rigid structures that sit idle between cells as small ions traverse across. Yet, although the importance of gap junctions in electrical synchronization is not questioned, it is generally accepted that these structures are more than electrical elements. Indeed, gap junctions are highly regulable molecular complexes present in the vast majority of cells in the body, including many cell types that are electrically nonexcitable. In addition to allowing the passage of ions, gap junctions allow the passage of small molecules as well. Hence, gap junctions provide not only electrical coupling to excitable cells but also metabolic coupling to all cell types where they are present.1 Yet, although the nature of the message that carries electrical information is rather well understood (ions carrying charge), the nature of the molecule(s) providing metabolic coupling is mostly unknown. The answer to the obvious question remains elusive: what goes through gap junctions in living cells?
The answer to this question is complicated by the fact that not all gap junctions are exactly the same. Gap junctions are formed by the oligomerization of a protein called connexin (Figure). Six connexin subunits noncovalently bind to form a hemichannel (or connexon); two connexons, one provided by each cell, dock their extracellular domains and open a hydrophilic pore into the neighboring cytoplasmic spaces, forming a gap junction channel. Twenty connexins have been identified in the human genome.2 Each connexin forms a channel with a signature unitary conductance and voltage dependence.1 Not all connexins act as substrates for the same kinases, and only a few are known to associate with either scaffolding proteins,3 microtubules,4 or other intracellular proteins.5 As such, a gap junction channel formed by one connexin isotype may function quite differently from another, built by a different isotype. In fact, genetic manipulation studies show that connexins are not interchangeable. Indeed, malformations caused by the absence of one connexin are not prevented when the missing connexin is substituted by a different isotype.6,7⇓ Given that individual connexins seem to have such a strong identity, it is reasonable to speculate that the molecular messages that can traverse across a gap junction would be different depending on the type of connexin that is present.
Adding to the complexity is the fact that many cells express more than one connexin isotype. Atrial myocytes, for example, express both Cx40 and Cx43, which oligomerize into the same hemichannel.8 In other words, atrial connexons may be heteromeric, with anywhere between 1 and 5 subunits being of one connexin (eg, Cx40) and the rest from the alternative isotype (Cx43). The question then is whether, in the case of a gap junction channel made out of two different connexins, the properties of the channel are dictated by the dominant behavior of one subunit or by the sum of the individual properties of each connexin in the complex.
The elegant studies of Valiunas et al,9 published in this issue of Circulation Research, bring us a step closer to understanding the functional consequences of connexin diversity. These authors have combined two techniques previously used independently to study gap junctions: patch clamp and dye transfer. Through a series of simple calculations, the authors estimate the number of Lucifer yellow molecules that traverse a gap junction channel depending on the connexin isotype that is present: Cx40, Cx43, or a combination of both. The findings are clear: Lucifer yellow passes with much more ease through a Cx43 channel than a Cx40 channel. Indeed, whereas 1351 molecules of Lucifer yellow can move through a Cx43 channel in one second, only 272 molecules pass when the gap junction is formed by Cx40. Comparing the ratio of potassium to Lucifer yellow permeability gives a similar result: the ratio is more than an order of magnitude higher in the case of Cx43. Interestingly, the numbers for heteromeric Cx40/Cx43 channels fall in between those obtained for the homomeric pairs. These data convincingly support the hypothesis that the ability of a molecule to navigate a gap junction is a function of the molecular composition of that particular channel. The findings are as robust as they are significant. It is tempting to speculate, for example, that variations in connexin expression within a tissue may be a key component of any remodeling process. A case in point may be that of atrial fibrillation, where it has been shown that the ratio of atrial Cx40/Cx43 expression is regionally modified.10 Could that be translated into a different type of intercellular message that may differentially regulate myocyte gene expression and control cardiac remodeling? In general, the results of this study suggest that the diversity of connexins translates into a diverse set of filters that can be placed between cells to control the flux of molecular information as it is proper to that particular cell group, under a particular set of conditions.
Although the nature of the molecules still remains to be determined, this study improves our understanding of the properties that are relevant for a molecule to move (or not) through a cardiac connexin channel. The results also support previous evidence indicating that, at least in the case of Cx40 and Cx43, a mixed channel is not dominated by one isotype. Instead, properties are added (either linearly or not11,12⇓) to bring about a functional unit with unique characteristics. This democratic organization brings to the system enormous flexibility (and complexity), as separate individuals keep their identity but, at the same time, integrate into a unified structure to preserve the function of the cell.
Connexin is not the only family of channel proteins that is composed of multiple isotypes. Yet, because of the peculiarities of the channel, connexins have been actively studied to determine what molecules, other than ions, move through, and which functions (other than the one relevant to electrophysiology) they cover. The data presented in this study and in others (see Reference 1 for review) clearly show that connexin diversity carries multiple functional implications, many of them unrelated to the electrical behavior of the organ. One must wonder if this is a fact exclusive to the connexin family. Take for example the potassium channels. Numerous integral membrane proteins have the ability to form a hydrophilic, potassium selective pore. But is that their only function? Could they also have a different role in cell homeostasis, maybe through their association with other intracellular proteins and/or extracellular molecules? Could that be a partial explanation for the large diversity of potassium channel-forming genes?
In summary, the study by Valiunas et al9 represents a very valuable contribution to the gap junction field and has implications for other ion channels as well. Their data emphasize the fact that connexins may act as selectivity filters for molecular information, and that the composition of such filters can be modulated by the combination of individual subunits in a somewhat democratic fashion. The big questions are still outstanding. But this study moves us one significant step closer to the long-sought answers.
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
- ↵Toyofuku T, Yabuki M, Otsu K, Kuzuya T, Hori M, Tada M. Direct association of the gap junction protein connexin-43 with ZO-1 in cardiac myocytes. J Biol Chem. 1998; 273: 12725–12731.
- ↵Giepmans BN, Hengeveld T, Postma FR, Moolenaar WH. Interaction of c-Src with gap junction protein connexin-43: role in the regulation of cell-cell communication. J Biol Chem. 2001; 276: 8544–8549.
- ↵White TW. Unique and redundant connexin contributions to lens development. Science. 2002; 295: 319–320.
- ↵Valiunas V, Beyer EC, Brink PR. Cardiac gap junction channels show quantitative differences in selectivity. Circ Res. 2002; 91: 104–111.
- ↵van der Velden HM, Ausma J, Rook MB, Hellemons AJ, van Veen TA, Allessie MA, Jongsma HJ. Gap junctional remodeling in relation to stabilization of atrial fibrillation in the goat. Cardiovasc Res. 2000; 46: 476–486.