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Pumilio-2 regulates translation of Nav1.6 to mediate homeostasis of membrane excitability.

ABSTRACT: The ability to regulate intrinsic membrane excitability, to maintain consistency of action potential firing, is critical for stable neural circuit activity. Without such mechanisms, Hebbian-based synaptic plasticity could push circuits toward activity saturation or, alternatively, quiescence. Although now well documented, the underlying molecular components of these homeostatic mechanisms remain poorly understood. Recent work in the fruit fly, Drosophila melanogaster, has identified Pumilio (Pum), a translational repressor, as an essential component of one such mechanism. In response to changing synaptic excitation, Pum regulates the translation of the voltage-gated sodium conductance, leading to a concomitant adjustment in action potential firing. Although similar homeostatic mechanisms are operational in mammalian neurons, it is unknown whether Pum is similarly involved. In this study, we report that Pum2 is indeed central to the homeostatic mechanism regulating membrane excitability in rat visual cortical pyramidal neurons. Using RNA interference, we observed that loss of Pum2 leads to increased sodium current (I(Na)) and action potential firing, mimicking the response by these neurons to being deprived of synaptic depolarization. In contrast, increased synaptic depolarization results in increased Pum2 expression and subsequent reduction in INa and membrane excitability. We further show that Pum2 is able to directly bind the predominant voltage-gated sodium channel transcript (NaV1.6) expressed in these neurons and, through doing so, regulates translation of this key determinant of membrane excitability. Together, our results show that Pum2 forms part of a homeostatic mechanism that matches membrane excitability to synaptic depolarization in mammalian neurons.

SUBMITTER: Driscoll HE

PROVIDER: S-EPMC3678506 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:RationaleCardiac function is dependent on the coordinate activities of membrane ion channels, transporters, pumps, and hormone receptors to tune the membrane electrochemical gradient dynamically in response to acute and chronic stress. Although our knowledge of membrane proteins has rapidly advanced during the past decade, our understanding of the subcellular pathways governing the trafficking and localization of integral membrane proteins is limited and essentially unstudied in vivo. In the heart, to our knowledge, there are no in vivo mechanistic studies that directly link endosome-based machinery with cardiac physiology.ObjectiveTo define the in vivo roles of endosome-based cellular machinery for cardiac membrane protein trafficking, myocyte excitability, and cardiac physiology.Methods and resultsWe identify the endosome-based Eps15 homology domain 3 (EHD3) pathway as essential for cardiac physiology. EHD3-deficient hearts display structural and functional defects including bradycardia and rate variability, conduction block, and blunted response to adrenergic stimulation. Mechanistically, EHD3 is critical for membrane protein trafficking, because EHD3-deficient myocytes display reduced expression/localization of Na/Ca exchanger and L-type Ca channel type 1.2 with a parallel reduction in Na/Ca exchanger-mediated membrane current and Cav1.2-mediated membrane current. Functionally, EHD3-deficient myocytes show increased sarcoplasmic reticulum [Ca], increased spark frequency, and reduced expression/localization of ankyrin-B, a binding partner for EHD3 and Na/Ca exchanger. Finally, we show that in vivo EHD3-deficient defects are attributable to cardiac-specific roles of EHD3 because mice with cardiac-selective EHD3 deficiency demonstrate both structural and electric phenotypes.ConclusionsThese data provide new insight into the critical role of endosome-based pathways in membrane protein targeting and cardiac physiology. EHD3 is a critical component of protein trafficking in heart and is essential for the proper membrane targeting of select cellular proteins that maintain excitability.

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Pumilio-2 regulates translation of Nav1.6 to mediate homeostasis of membrane excitability.

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