Cyclin-dependent kinase 5 is an upstream regulator of mitochondrial fission during neuronal apoptosis

Abstract

Under physiological conditions, mitochondrial morphology dynamically shifts between a punctuate appearance and tubular networks. However, little is known about upstream signal transduction pathways that regulate mitochondrial morphology. We show that mitochondrial fission is a very early and kinetically invariant event during neuronal cell death, which causally contributes to cytochrome c release and neuronal apoptosis. Using a small molecule CDK5 inhibitor, as well as a dominantnegative CDK5 mutant and RNAi knockdown experiments, we identified CDK5 as an upstream signalling kinase that regulates mitochondrial fission during apoptosis of neurons. Vice versa, our study shows that mitochondrial fission is a modulator contributing to CDK5-mediated neurotoxicity. Thereby, we provide a link that allows integration of CDK5 into established neuronal apoptosis pathways. Mitochondria can acquire a broad range of morphologies, from a punctuate shape to a tubular appearance or even extended networks. Several proteins, most of them belonging to the family of large GTPases, have been identified that energy-dependently regulate mitochondrial morphology and subcellular distribution by promoting either fission (Fis1, dynamin-related protein 1 (Drp1), MTP18) or fusion of mitochondria (Mfn1/2 and Opa1). 1-3 As a potential upstream regulatory factor, the actin cytoskeleton has been shown to be a prerequisite for mitochondrial fission, possibly by its influence on Drp1 recruitment to mitochondria. 4 Apoptotic release of the mitochondrial cytochrome c and other proapoptotic mitochondrial proteins results in formation of the so-called apoptosome, which in turn activates downstream effector caspases with death executing function. 5 This intrinsic, mitochondrial death pathway is relevant for most forms of neuronal apoptosis, in contrast to extrinsic, for example death receptor mediated, activation of programmed cell death. 6 However, despite the contribution of mitochondrial dysfunction specifically to the demise of neurons, there is only scarce knowledge about mitochondrial fission during neuronal apoptosis, nor about its functional contribution to neuronal cell death. Furthermore, little is known about upstream regulatory pathways for mitochondrial morphology in general.Cyclin-dependent kinase 5 (CDK5) is a member of the CDK family, but, distinct from other CDKs, it does not participate in cell cycle regulation. 7 Instead, CDK5 is physiologically implicated in cytoskeletal functions, as well as regulation of membrane turnover and morphology, for example cell migration or endocytosis. Moreover, it is involved in neuronspecific functions including, for example, synaptic plasticity, axonal outgrowth or transmitter release. [8][9][10][11] In neurological diseases, deregulated CDK5 turned out to be an apical instigator of neuronal cell death cascades. Amyloid-toxicity was shown to induce calpain-mediated cleavage of the CDK5 activators p35 or p39 to p25 and p29. This leads to redistribution and overactivation of CDK5 after its association...

S-Nitrosylation activates Cdk5 and contributes to synaptic spine loss induced by β-amyloid peptide

“…These findings suggest that SNO-Cdk5 may represent a unique therapeutic target for ameliorating spine damage in AD and other neurodegenerative conditions. Prior reports showed that Cdk5 can trigger excessive mitochondrial fission involving Drp1 and suggested that Drp1 may be a substrate for Cdk5 regulation (43). Such a relationship may link our recent findings that Aβ-induced activation of Drp1 via S-nitrosylation causes excessive mitochondrial fragmentation, with consequent bioenergetic compromise and dendritic spine loss (20,40).…”

“…Therefore, we asked whether SNO-Cdk5 in brains with Alzheimer's disease might possibly contribute to this pathway to synaptic damage via transfer of the NO group to Drp1 (to form SNODrp1) by a reaction mechanism termed transnitrosylation (39,41,42). Previously, it was postulated that Cdk5 could activate Drp1, but the pathway remained obscure because direct phosphorylation of Drp1 by Cdk5 was not observed (43). Here, using the biotin-switch assay, we initially found that SNO-Cdk5 could indeed S-nitrosylate Drp1 in vitro via transnitrosylation, with consequent formation of SNO-Drp1 (Fig.…”

Activated Cyclin-Dependent Kinase 5 Promotes Microglial Phagocytosis of Fibrillar β-Amyloid by Up-regulating Lipoprotein Lipase Expression

“…In AD brains, increased cleavage of p35 to p25 because of A␤ deposition results in formation of the CDK5/ p25 complex and CDK5 overactivation (3), which dysregulates the downstream substrates of CDK5, such as hyperphosphorylated tau proteins (36). Moreover, expression of the CDK5/p25 complex in cultured primary neurons induces cytoskeletal disruption, morphological degeneration, mitochondrial fission (54), and apoptosis (55). In addition, CDK5/p25 increases A␤ production by regulating BACE1 expression, and enhances the A␤-calpain-CDK5/p25 cycle that aggravates the disease in AD brains.…”

“…CDK5 is a critical proline-directed protein kinase in AD pathogenesis, and its activity is tightly regulated by cleavage of the catalytic subunit p35 to p25. CDK5 plays a critical role in neuronal cell migration, neurite growth, and synaptogenesis in the developing brain (53,54). In AD brains, increased cleavage of p35 to p25 because of A␤ deposition results in formation of the CDK5/ p25 complex and CDK5 overactivation (3), which dysregulates the downstream substrates of CDK5, such as hyperphosphorylated tau proteins (36).…”

Dephosphorylation by calcineurin regulates translocation of Drp1 to mitochondria

“…The inner membrane rhomboid protease Parl, which participates in the production of the soluble intermembrane space form of Opa1 that regulates apoptosis (6,7), is specifically phosphorylated on its vertebrate-specific P␤ domain to block its self-cleavage and mitochondrial fragmentation (8). Additionally, cyclin dependent kinase (Cdk) 5 has been reported to promote mitochondrial fragmentation in neurons, albeit the molecular nature of the mediator of its effects on mitochondrial shape is unknown (9). A step forward in our understanding of the integration of mitochondrial shape changes by cellular cues came with the discovery that Drp1-dependent mitochondrial fragmentation is controlled by phosphorylation at two different conserved sites, serine 616 and 637, by Cdk1 and protein kinase A (PKA) (10)(11)(12).…”