LIAISON: Molecular mechanisms and pathophysiological importance of a novel interaction between Kv channels across families

电压门控K(Kv)通道通常是由四个相关的家族成员组成的四聚体组装而成，以允许质膜上的K通量响应于膜电位的去极化。Kv5、Kv6、Kv8和Kv9家族是特殊的，因为当它们单独表达时，它们是电沉默的(称为无声Kv，“KVS”)。然而，它们与Kv2亚基共组装成具有不同于同构体Kv2通道的性质的异构体。这种Kv2-KvS共组装构成了目前已知的唯一跨Kv家族的亚基异构化的例子。我们认识到KV7亚基与KVS的同源性高于所有其他家族，并想知道KV7通道是否也可能受到KVS的调制。在广泛的先导实验中，我们确实发现KVS以亚基特异性的方式调节重组KV7亚型的活性、生物物理性质和质膜表达。我们还获得了KVS和KV7亚基在一个共同的蛋白质复合体中共定位的证据，这可能使这些亚基直接相互作用，甚至可能发生异构化。我们进一步发现，KV7和KV8(唯一与疾病相关的KVS)在海马神经元和视网膜光感受器中共表达。由于KV7和KV8亚基的突变似乎导致相似的病理(例如癫痫和感觉障碍)，这些发现表明KV7-KVS相互作用具有潜在的(病理)生理学相关性。事实上，致病的Kv8突变以某种方式减弱了Kv7电流，这可能有助于Kv8相关癫痫和视网膜病变的发展。我们建议由三个具有互补专业知识的实验室联合进行研究，以揭示KV7和KVS亚基之间意外相互作用的机制和影响。我们将结合生化、电生理和成像技术来阐明表达系统中相互作用的分子机制和原理，并确定允许KVS依赖的KV7电流调节的蛋白质基序。我们还将探索重组系统中确定的原理是否适用于天然组织，从而揭示KV7-KVS复合体在海马区和视网膜中的生理相关性。最终，我们将通过改变KV7通道激动剂的用途和探索基因治疗策略来建立KV8相关疾病的治疗方案。总之，联络将对不同Kv通道家族成员之间新的相互作用背后的分子原理提供重要的见解，从而挑战关于功能性Kv通道形成的中心定理。我们的项目将为天然Kv通道复合体的异质性提供新的见解，并为治疗依赖Kv的通道病提供新的治疗视角。

Voltage-gated K+ (Kv) channels usually are formed by tetrameric assembly of four related family members to allow for K+ flux over the plasma membrane in response to depolarization of the membrane potential. The Kv5, Kv6, Kv8 and Kv9 families are particular, as they are electrically silent (termed silent Kv, “KvS”), when expressed alone. However, they co assemble with Kv2 subunits into heteromers with properties differing from those of homomeric Kv2 channels. This Kv2-KvS co-assembly constitutes the only yet known example for heteromerization of subunits across Kv families. We recognised that Kv7 subunits exhibit higher homology to KvS than to all other families and wondered whether – in analogy to Kv2 – Kv7 channels might also be modulated by KvS. In extensive pilot experiments, we indeed found that KvS modulated activity, biophysical properties, and plasma membrane expression of recombinant Kv7 isoforms in a subunit-specific manner. We also acquired evidence for co-localization of KvS and Kv7 subunits in a common protein complex, which may enable direct interaction and potentially even heteromerization of these subunits. We furthermore found that Kv7 and Kv8 (the only KvS with established disease relevance) are co-expressed in hippocampal neurons and retinal photoreceptors. As mutations in Kv7 and Kv8 subunits appear to cause similar pathologies (e.g. epilepsy and sensory impairment), these findings indicate potential (patho)physiological relevance of the Kv7-KvS interaction. Indeed, pathogenic Kv8 mutations attenuated Kv7 currents in a way that may contribute to development of Kv8-related epilepsy and a retinopathy. We propose LIAISON, a joint research effort of three laboratories with complementary expertise to unravel the mechanisms and implications of the unexpected interaction between Kv7 and KvS subunits. We will utilize a combination of biochemical, electrophysiological and imaging techniques to elucidate the molecular mechanisms and principles of the interaction in expression systems, and we will identify the protein motifs that allow for KvS-dependent modulation of Kv7 currents. We will also explore whether the principles identified in recombinant systems apply in native tissue and thereby unravel the physiological relevance of Kv7-KvS complexes in hippocampus and retina. Eventually, we will establish treatment options for a Kv8-related disorder by repurposing Kv7 channel agonists and by exploring gene-therapeutic strategies. In summary, LIAISON will deliver significant insight into the molecular principles underlying a novel interaction between members of different Kv channel families, thereby challenging a central theorem on the formation of functional Kv channels. Our project will provide new insight into the heterogeneity of native Kv channel complexes and offers novel treatment perspectives against Kv-dependent channelopathies.