3 research outputs found
Presynaptic α2δ subunits are key organizers of glutamatergic synapses
In nerve cells the genes encoding for α2δ subunits of voltage-gated calcium channels have been linked to synaptic functions and neurological disease. Here we show that α2δ subunits are essential for the formation and organization of glutamatergic synapses. Using a cellular α2δ subunit triple-knockout/knockdown model, we demonstrate a failure in presynaptic differentiation evidenced by defective presynaptic calcium channel clustering and calcium influx, smaller presynaptic active zones, and a strongly reduced accumulation of presynaptic vesicle-associated proteins (synapsin and vGLUT). The presynaptic defect is associated with the downscaling of postsynaptic AMPA receptors and the postsynaptic density. The role of α2δ isoforms as synaptic organizers is highly redundant, as each individual α2δ isoform can rescue presynaptic calcium channel trafficking and expression of synaptic proteins. Moreover, α2δ-2 and α2δ-3 with mutated metal ion-dependent adhesion sites can fully rescue presynaptic synapsin expression but only partially calcium channel trafficking, suggesting that the regulatory role of α2δ subunits is independent from its role as a calcium channel subunit. Our findings influence the current view on excitatory synapse formation. First, our study suggests that postsynaptic differentiation is secondary to presynaptic differentiation. Second, the dependence of presynaptic differentiation on α2δ implicates α2δ subunits as potential nucleation points for the organization of synapses. Finally, our results suggest that α2δ subunits act as transsynaptic organizers of glutamatergic synapses, thereby aligning the synaptic active zone with the postsynaptic density
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α2δ-4 and Cachd1 proteins are regulators of presynaptic functions
The α2δ auxiliary subunits of voltage-gated calcium channels (VGCC) were traditionally regarded as modulators of biophysical channel properties. In recent years, channel-independent functions of these subunits, such as involvement in synapse formation, have been identified. In the central nervous system, α2δ isoforms 1, 2, and 3 are strongly expressed, regulating glutamatergic synapse formation by a presynaptic mechanism. Although the α2δ-4 isoform is predominantly found in the retina with very little expression in the brain, it was recently linked to brain functions. In contrast, Cachd1, a novel α2δ-like protein, shows strong expression in brain, but its function in neurons is not yet known. Therefore, we aimed to investigate the presynaptic functions of α2δ-4 and Cachd1 by expressing individual proteins in cultured hippocampal neurons. Both α2δ-4 and Cachd1 are expressed in the presynaptic membrane and could rescue a severe synaptic defect present in triple knockout/knockdown neurons that lacked the α2δ-1-3 isoforms (α2δ TKO/KD). This observation suggests that presynaptic localization and the regulation of synapse formation in glutamatergic neurons is a general feature of α2δ proteins. In contrast to this redundant presynaptic function, α2δ-4 and Cachd1 differentially regulate the abundance of presynaptic calcium channels and the amplitude of presynaptic calcium transients. These functional differences may be caused by subtle isoform-specific differences in α1 -α2 δ protein–protein interactions, as revealed by structural homology modelling. Taken together, our study identifies both α2δ-4 and Cachd1 as presynaptic regulators of synapse formation, differentiation, and calcium channel functions that can at least partially compensate for the loss of α2δ-1-3. Moreover, we show that regulating glutamatergic synapse formation and differentiation is a critical and surprisingly redundant function of α2δ and Cachd1
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The presynaptic α2δ protein family and their therapeutic potential
α2δ proteins are well-established modulators of membrane expression 5 and biophysical properties of voltage-gated calcium channels. Moreover, they are 6 critical regulators of synapse formation and function and key players in transsynap- 7 tic signalling. The α2δ isoforms are highly glycosylated membrane-anchored pro- 8 teins with distinct structural features, some of which are also observed in a human 9 α2δ-like protein termed CACHD1. Accumulating evidence has underpinned the 10 involvement of α2δ proteins in neurological and neurodevelopmental disorders, 11 making them attractive novel therapeutic targets. Also, CACHD1, through its mod- 12 ulation of T-type currents, is an emerging potential drug target, particularly for epi- 13 lepsy and pain. Furthermore, α2δ proteins are targets of the widely prescribed 14 gabapentinoids. Among these, gabapentin and pregabalin, which have a high bind- 15 ing affinity for α2δ-1 and α2δ-2, have been administered particularly in the treatment 16 of neuropathic pain conditions, epilepsy, and restless leg syndrome. Extensive 17 efforts have been and are being made to understand the structure and functions of 18 α2δ proteins and how they interact with synaptic proteins. This is ultimately helping 19 to understand the contribution of the α2δ protein family to neurological and neuro- 20 developmental disorders and provides insight into potential novel treatment options