Transverse tubules strike back: may the junctophilin-2 be with you

This editorial refers to ‘Junctophilin-2 tethers T-tubules and recruits functional L-type calcium channels to lipid rafts in adult cardiomyocytes’ by C. Poulet et al., pp. 149–161. Twenty years ago, in our galaxy, transverse tubules (t-tubules) interest awaken as not only a structural but also a functional compartment. In mammalian cardiac ventricular myocytes t-tubules are invaginations of the sarcolemma that occur at the Z line as transverse but also as longitudinal elements.1 It is now well admitted that t-tubules are central to excitation–contraction coupling (ECC), with major actors concentrated in this sub-structure; this is not only true for ventricular myocytes but also atrial myocytes.2 In more details, close apposition of L-type calcium channels (LTCCs) at the sarcolemma, to Ryanodine receptors (RyR), at the sarcoplasmic reticulum (SR) membrane, forms junctional membrane complexes (JMCs),3 and this configuration is essential to obtain an very efficient ECC: the cell depolarization during action potential (AP) activates LTCCs which permit a small amount of Ca2+ influx to reach the cytosol.4 Then, calcium increases in the JMCs allow the opening of the RyR to trigger Ca2+ release from the SR, phenomenon called Ca2+-induced Ca2+ release (CICR). Relaxation is brought about by removal of Ca2+ from the cell cytoplasm by two main routes: the SR Ca2+ ATPase (SERCA), pumps Ca2+ back into the SR and while Na+/Ca2+ exchange (NCX) to extrude Ca2+ from the cell.5 Twenty years ago, Cardiovascular Research was the first journal to publish compelling evidences that t-tubules structure is altered in heart failure.6 Since then, there is now a consensus that t-tubules alterations is involved in reduced synchrony of Ca2+ release not only in heart failure but also atrial fibrillation in animal model.6–11 Meanwhile, in the same galaxy, biochemists identified several key proteins as key players in t-tubule scafolding in cardiomyocytes: junctonphinlin-2 (JPH2), Bridging Integrator-(Bin1), and Caveolin-3 (Cav3).12 The article by Poulet et al.13 from the Gorelik lab, takes advantages of cardiomyocytes that losses their t-tubules during culture14 to investigate in close details the role of JPH2 and Cav3 in t-tubules maintenance. They observed that t-tubules loss during cell culture is correlated with down-regulation of the expression of JPH2 and Cav3. In contrary, JPH2 overexpression with Ad-h-JPH2 adenovirus after 4 days in culture helps to prevent JPH2, Cav3, and t-tubule loss and disorganization. Interestingly, t-tubule network organization by JPH2 overexpression is completely abolished by methyl-b-cyclodextrin (MβCD) which removes cholesterol from the membrane and also disrupts lipids rafts, including caveolae. This observation clearly shows that even in presence of JPH2, the absence of lipids rafts does not permit formation of t-tubule by anchoring of, JPH2 at the membrane. Nevertheless, previous experimentations described in the literature failed to demonstrate any interaction between JPH2 and lipids, so the anchoring of JPH2 in the lipids raft seems to be due to an intermediary. Proximity ligation assay indicates a close colocalization between JPH2 and RyR, Cav3, Cav1.2, and Cavβ2 but only colocalization with Cav3 and Cavβ2 is disrupted by MβCD. No interaction motif has been identified between JPH2 and Cav3, this need further experiments. Furthermore, Cav3 overexpression failed to protect against the detrimental effect of cholesterol depletion on t-tubule. Nevertheless, even if the molecular interaction identity is unclear, it is clearly demonstrated that JPH2 anchors in the lipids rafts with cholesterol and Cav3 as patterns. Moreover, JPH2 is essential for t-tubule structure but also directly involved in ECC efficiency. By using the state of the art scanning ion conductance microscopy, Poulet et al.13 were able to generate high resolution 3D mapping of the membrane and to record single-channel activity in patch-clamp at the surface membrane (crest) and inside the t-tubule. Under basal conditions, LTTCs had higher density in the t-tubule rather than the crest.15 However, LTCCs distribution is disrupted after 4 days in culture, resulting in a predominant localization at the crest. JPH2 overexpression counteracts these effects. A comprehensive study investigating JPH2, Bin1, Cav3 in the role of t-tubules is still required (Figure 1). Junctonphinlin-2 promotes t-tubules stability. Junctonphinlin-2 recruits functional L-type Ca channels, composed of pore forming Cav1.2 subunit and accessory Cavβ2 subunit. Cholesterol is involved in the binding of Junctonphinlin-2 to T-tubules via lipid rafts with the support from Caveoline 3. The role of bin1 is unclear and deserve further investigation. Furthermore, it has been established that JPH2 overexpression recruits a greater number of LTTCs functionals in t-tubule and crest. LTCCs recruitment seems to involve the channel trafficking rather than by increased expression of Cav1.2. Interestingly, LTTCs activity is disrupted the dissociation of lipids rafts but not equally between t-tubule and crest. This observation suggests a different regulation of LTTCs activity in these two domains. The findings of Poulet et al. suggest that lipids rafts contain important molecules to modulate LTCCs activity such as accessory LTTCs subunits. Indeed, regulation of LTCCs activity is essential to maintain an effective CICR at the t-tubule. The work by Poulet et al. on JPH2 provides a promising tool for future treatment in cardiac diseases. Is it the NEW HOPE? Conflict of interest: none declared. This work was supported by the French National Funding Agency for Research (ANR) (ANR-10-IAHU04-LIRYC). The opinions expressed in this article are not necessarily those of the Editors of Cardiovascular Research or of the European Society of Cardiology.