Browsing School, Graduate by Subject "Mitochondrial Dynamics"
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Novel Signaling Mechanisms in the Regulation of Mitochondrial DynamicsMitochondria are dynamic organelles that constantly undergo fission and fusion events (referred to as mitochondrial dynamics) to form highly interconnected networks within cells. These networks allow mitochondria to share resources such as mitochondrial DNA and antioxidant molecules to maintain the health of the network. Because mitochondria are the main source of production of ATP through oxidative phosphorylation, and also regulate cell death through apoptosis, it is critically important to maintain homeostasis in these organelles. Indeed, dysfunction in mitochondrial dynamics has been linked to numerous diseases, including cancer, neurodegenerative, endocrine, and cardiovascular diseases. Therefore, understanding the mechanisms by which mitochondrial dynamics contributes to the overall health of this organelle is of great interest. The primary proteins involved in the regulation of mitochondrial fusion and fission, and the mechanisms by which they act, are generally understood. It is also well accepted that mitochondrial fusion and fission is balanced; however, how these two separate processes communicate and signal to each other is currently unknown. To better understand the crosstalk between mitochondrial fission and fusion, we studied a function of the outer mitochondrial membrane associated E3 ubiquitin ligase, MARCH5. We found that MARCH5 acts as a negative regulator of mitochondrial fission through the ubiquitin-dependent degradation of the fission factor, MiD49. Shedding light on a possible mechanism by which the activities of fission factors are coordinated, we found that the Drp1 receptor, Mff, promotes MiD49 stability by negatively regulating MARCH5 activity, thereby enhancing mitochondrial fission rates. Finally, supporting molecular crosstalk between fission and fusion, we found that Mff also regulates the stability of the outer mitochondrial membrane fusion factors, Mfn1 and Mfn2, and that loss of Mff expression/activity results in reduced mitochondrial fusion rates in those cells. Thus, the studies presented here display novel crosstalk and signaling mechanisms by which fission factors are able to fine-tune mitochondrial fission and fusion rates through modification of the ubiquitin-proteasome system.
The regulation and role of dendritic mitochondrial fission during long-term potentiationNeurons continuously modify their synaptic strength to encode memories and to adapt to experience and the environment. Long-term potentiation (LTP) is a critical cellular mechanism of this adaptation and is the prevailing form of synaptic plasticity. Baseline synaptic function is bioenergetically demanding, and this demand is elevated during episodes of synaptic plasticity. Therefore, mitochondrial functions, such as ATP synthesis and calcium handling, are likely essential for plasticity. Furthermore, mitochondrial functions themselves are regulated by mitochondrial dynamics including fission, fusion, and motility. Although axonal mitochondria have been extensively studied, LTP induction predominantly occurs postsynaptically, where the roles of mitochondria are less well understood. Additionally, mitochondrial fission has recently garnered interest because it is necessary for development and is required for normal mitochondrial function, and because perturbed fission is associated with many neurological and psychiatric diseases. However, whether or how fission in dendrites supports ongoing synaptic transmission and plasticity is still unclear. Furthermore, although the molecular mechanisms underlying mitochondrial fission have been well described in other cell types, little is known about how mitochondrial fission is accomplished in neurons, particularly in dendrites, or how neuronal activity might modulate these mechanisms. Here I tested the hypothesis that dendritic mitochondrial fission is triggered during LTP induction, and is necessary for LTP expression. Mitochondria in dendrites at rest are stationary and rarely undergo fission. However, I found that chemical induction of LTP (cLTP) by NMDAR activation in cultured rat hippocampal neurons prompted a rapid burst of dendritic mitochondrial fission. Mitochondrial fission canonically requires actin nucleation and membrane constriction by the GTPase dynamin-related protein 1 (Drp1). Consistent with this, inhibition of actin polymerization or expression of a dominant negative (DN) mutant or knockdown of Drp1 each suppressed the cLTP fission burst. Furthermore, the GTPase Dynamin 2 (Dyn2) was recently implicated in fission in cell lines, and I found similarly that expressing DN Dyn2 abolished the fission burst. Drp1 function is also known to be regulated by phosphorylation, with CaMKII as a possible activator based on studies of non-neuronal cells. In line with this, I found that fission was triggered by cytosolic calcium elevation via glutamate photolysis at dendritic spines, and also that the fission burst was prevented by acutely inhibiting CaMKII activation or by prohibiting Drp1 phosphorylation. I then tested whether mitochondrial fission is required for LTP expression. Knocking down Drp1 or expressing DN Drp1 suppressed dendritic spine growth and synaptic AMPA receptor trafficking following LTP induction. Furthermore, NMDAR-dependent LTP induction by high-frequency stimulation (HFS) of Schaffer collaterals in acute hippocampal slices decreased dendritic mitochondrial length in area CA1. Remarkably, postsynaptic expression of DN Drp1 prevented HFS LTP at Schaffer collateral-CA1 synapses in slices, with no effect on basal transmission or intrinsic electrophysiological properties of neurons. Furthermore, I found that cLTP stimulation produced transient elevations of dendritic mitochondrial calcium (i.e. mCaTs), and that expression of DN Drp1 suppressed the frequency, amplitude, and duration of evoked mCaTs. These data illustrate a novel pathway whereby synaptic activity controls mitochondrial fission, and show that dynamic control of fission is required for LTP induction perhaps by modulating mitochondrial calcium handling. Impaired synaptic function is implicated in myriad neuropsychiatric diseases, many of which are also associated with mitochondrial dysfunction. Therefore, our findings raise the important question of whether neuronal mitochondrial dysfunction contributes to cognitive impairment in these diseases by perturbing dendritic and/or synaptic plasticity.