Novel MECP2 gene therapy is effective in a multicenter study using two mouse models of Rett syndrome and is safe in non-human primates.

The AAV9 gene therapy vector presented in this study is safe in mice and non-human primates and highly efficacious without causing overexpression toxicity, a major challenge for clinical translation of Rett syndrome gene therapy vectors to date. Our team designed a new truncated methyl-CpG-binding protein 2 (MECP2) promoter allowing widespread expression of MECP2 in mice and non-human primates after a single injection into the cerebrospinal fluid without causing overexpression symptoms up to 18 months after injection. Additionally, this new vector is highly efficacious at lower doses compared with previous constructs as demonstrated in extensive efficacy studies performed by two independent laboratories in two different Rett syndrome mouse models carrying either a knockout or one of the most frequent human mutations of Mecp2. Overall, data from this multicenter study highlight the efficacy and safety of this gene therapy construct, making it a promising candidate for first-in-human studies to treat Rett syndrome.

Radically truncated MeCP2 rescues Rett syndrome-like neurological defects.

Heterozygous mutations in the X-linked MECP2 gene cause the neurological disorder Rett syndrome. The methyl-CpG-binding protein 2 (MeCP2) protein is an epigenetic reader whose binding to chromatin primarily depends on 5-methylcytosine. Functionally, MeCP2 has been implicated in several cellular processes on the basis of its reported interaction with more than 40 binding partners, including transcriptional co-repressors (for example, the NCoR/SMRT complex), transcriptional activators, RNA, chromatin remodellers, microRNA-processing proteins and splicing factors. Accordingly, MeCP2 has been cast as a multi-functional hub that integrates diverse processes that are essential in mature neurons. At odds with the concept of broad functionality, missense mutations that cause Rett syndrome are concentrated in two discrete clusters coinciding with interaction sites for partner macromolecules: the methyl-CpG binding domain and the NCoR/SMRT interaction domain. Here we test the hypothesis that the single dominant function of MeCP2 is to physically connect DNA with the NCoR/SMRT complex, by removing almost all amino-acid sequences except the methyl-CpG binding and NCoR/SMRT interaction domains. We find that mice expressing truncated MeCP2 lacking both the N- and C-terminal regions (approximately half of the native protein) are phenotypically near-normal; and those expressing a minimal MeCP2 additionally lacking a central domain survive for over one year with only mild symptoms. This minimal protein is able to prevent or reverse neurological symptoms when introduced into MeCP2-deficient mice by genetic activation or virus-mediated delivery to the brain. Thus, despite evolutionary conservation of the entire MeCP2 protein sequence, the DNA and co-repressor binding domains alone are sufficient to avoid Rett syndrome-like defects and may therefore have therapeutic utility.

Neurophysiological Signatures of Motor Impairment in Patients with Rett Syndrome.

OBJECTIVE: Rett syndrome (RTT) is an X-linked dominant neurodevelopmental disorder due to pathogenic mutations in the MECP2 gene. Motor impairment constitutes the core diagnostic feature of RTT. Preclinical studies have consistently demonstrated alteration of excitation/inhibition (E/I) balance and aberrant synaptic plasticity at the cortical level. We aimed to understand neurobiological mechanisms underlying motor deficit by assessing in vivo synaptic plasticity and E/I balance in the primary motor cortex (M1). METHODS: In 14 patients with typical RTT, 9 epilepsy control patients, and 11 healthy controls, we applied paired-pulse transcranial magnetic stimulation (TMS) protocols to evaluate the excitation index, a biomarker reflecting the contribution of inhibitory and facilitatory circuits in M1. Intermittent TMS-theta burst stimulation was used to probe long-term potentiation (LTP)-like plasticity in M1. Motor impairment, assessed by ad hoc clinical scales, was correlated with neurophysiological metrics. RESULTS: RTT patients displayed a significant increase of the excitation index (p = 0.003), as demonstrated by the reduction of short-interval intracortical inhibition and increase of intracortical facilitation, suggesting a shift toward cortical excitation likely due to GABAergic dysfunction. Impairment of inhibitory circuits was also confirmed by the reduction of long-interval intracortical inhibition (p = 0.002). LTP-like plasticity in M1 was abolished (p = 0.008) and scaled with motor disability (all p = 0.003). INTERPRETATION: TMS is a method that can be used to assess cortical motor function in RTT patients. Our findings support the introduction of TMS measures in clinical and research settings to monitor the progression of motor deficit and response to treatment. ANN NEUROL 2020;87:763-773.

Inhibition of Piezo1 attenuates demyelination in the central nervous system.

Piezo1 is a mechanosensitive ion channel that facilitates the translation of extracellular mechanical cues to intracellular molecular signaling cascades through a process termed, mechanotransduction. In the central nervous system (CNS), mechanically gated ion channels are important regulators of neurodevelopmental processes such as axon guidance, neural stem cell differentiation, and myelination of axons by oligodendrocytes. Here, we present evidence that pharmacologically mediated overactivation of Piezo1 channels negatively regulates CNS myelination. Moreover, we found that the peptide GsMTx4, an antagonist of mechanosensitive cation channels such as Piezo1, is neuroprotective and prevents chemically induced demyelination. In contrast, the positive modulator of Piezo1 channel opening, Yoda-1, induces demyelination and neuronal damage. Using an ex vivo murine-derived organotypic cerebellar slice culture model, we demonstrate that GsMTx4 attenuates demyelination induced by the cytotoxic lipid, psychosine. Importantly, we confirmed the potential therapeutic effects of GsMTx4 peptide in vivo by co-administering it with lysophosphatidylcholine (LPC), via stereotactic injection, into the cerebral cortex of adult mice. GsMTx4 prevented both demyelination and neuronal damage usually caused by the intracortical injection of LPC in vivo; a well-characterized model of focal demyelination. GsMTx4 also attenuated both LPC-induced astrocyte toxicity and microglial reactivity within the lesion core. Overall, our data suggest that pharmacological activation of Piezo1 channels induces demyelination and that inhibition of mechanosensitive channels, using GsMTx4, may alleviate the secondary progressive neurodegeneration often present in the latter stages of demyelinating diseases.

Differential Dependence of GABAergic and Glutamatergic Neurons on Glia for the Establishment of Synaptic Transmission.

In the mammalian cortex, GABAergic and glutamatergic neurons represent 2 major neuronal classes, which establish inhibitory and excitatory synapses, respectively. Despite differences in their anatomy, physiology and developmental origin, both cell types require support from glial cells, particularly astrocytes, for their growth and survival. Recent experiments indicate that glutamatergic neurons also depend on astrocytes for synapse formation. However, it is not clear if the same holds true for GABAergic neurons. By studying highly pure GABAergic cell cultures, established through fluorescent activated cell sorting, we find that purified GABAergic neurons are smaller and have reduced survival, nevertheless they establish robust synaptic transmission in the absence of glia. Support from glial cells reverses morphological and survival deficits, but does little to alter synaptic transmission. In contrast, in cultures of purified glutamatergic neurons, morphological development, survival and synaptic transmission are collectively dependent on glial support. Thus, our results demonstrate a fundamental difference in the way GABAergic and glutamatergic neurons depend on glia for the establishment of synaptic transmission, a finding that has important implications for our understanding of how neuronal networks develop.

Exclusive expression of MeCP2 in the nervous system distinguishes between brain and peripheral Rett syndrome-like phenotypes.

Rett syndrome (RTT) is a severe genetic disorder resulting from mutations in the X-linked MECP2 gene. MeCP2 protein is highly expressed in the nervous system and deficiency in the mouse central nervous system alone recapitulates many features of the disorder. This suggests that RTT is primarily a neurological disorder, although the protein is reportedly widely expressed throughout the body. To determine whether aspects of the RTT phenotype that originate in non-neuronal tissues might have been overlooked, we generated mice in which Mecp2 remains at near normal levels in the nervous system, but is severely depleted elsewhere. Comparison of these mice with wild type and globally MeCP2-deficient mice showed that the majority of RTT-associated behavioural, sensorimotor, gait and autonomic (respiratory and cardiac) phenotypes are absent. Specific peripheral phenotypes were observed, however, most notably hypo-activity, exercise fatigue and bone abnormalities. Our results confirm that the brain should be the primary target for potential RTT therapies, but also strongly suggest that some less extreme but clinically significant aspects of the disorder arise independently of defects in the nervous system.

A computational study on plasticity during theta cycles at Schaffer collateral synapses on CA1 pyramidal cells in the hippocampus.

Cellular activity in the CA1 area of the hippocampus waxes and wanes at theta frequency (4-8 Hz) during exploratory behavior of rats. Perisomatic inhibition onto pyramidal cells tends to be strongest out of phase with pyramidal cell activity, whereas dendritic inhibition is strongest in phase with pyramidal cell activity. Synaptic plasticity also varies across the theta cycle, from strong long-term potentiation (LTP) to long-term depression (LTD), putatively corresponding to encoding and retrieval phases for information patterns encoded by pyramidal cell activity (Hasselmo et al. (2002a) Neural Comput 14:793-817). The mechanisms underpinning the phasic changes in plasticity are not clear, but it is likely that inhibition plays a role by affecting levels of electrical activity and calcium concentration at synapses. We explore the properties of synaptic plasticity during theta at Schaffer collateral synapses on CA1 pyramidal neurons and the influence of spatially and temporally targeted inhibition using a detailed multicompartmental model of the CA1 pyramidal neuron microcircuit and a phenomenological model of synaptic plasticity. The results suggest CA3-CA1 synapses are potentiated on one phase of theta due to high calcium levels provided by paired weak CA3 and layer III entorhinal cortex (EC) inputs even when somatic spiking is inhibited by perisomatic interneuron activity. Weak CA3 inputs alone induce lower calcium transients and result in depression of the CA3-CA1 synapses. These synapses are depressed if activated in phase with dendritic inhibition as strong CA3 inputs alone are not able to cause high calcium in this theta phase even though the CA1 pyramidal neuron shows somatic spiking. Dendritic inhibition acts as a switch that prevents LTP and promotes LTD during the retrieval phases of the theta rhythm in CA1 pyramidal cell. This may be important for not overly reinforcing recalled memories and in forgetting no longer relevant memories.

Label-free imaging of thick tissue at 1550 nm using a femtosecond optical parametric generator.

We have developed a simple wavelength-tunable optical parametric generator (OPG), emitting broadband ultrashort pulses with peak wavelengths at 1530-1790 nm, for nonlinear label-free microscopy. The OPG consists of a periodically poled lithium niobate crystal, pumped at 1064 nm by a ultrafast Yb:fiber laser with high pulse energy. We demonstrate that this OPG can be used for label-free imaging, by third-harmonic generation, of nuclei of brain cells and blood vessels in a >150 µm thick brain tissue section, with very little decay of intensity with imaging depth and no visible damage to the tissue at an incident average power of 15 mW.

Spine head calcium as a measure of summed postsynaptic activity for driving synaptic plasticity.

We use a computational model of a hippocampal CA1 pyramidal cell to demonstrate that spine head calcium provides an instantaneous readout at each synapse of the postsynaptic weighted sum of all presynaptic activity impinging on the cell. The form of the readout is equivalent to the functions of weighted, summed inputs used in neural network learning rules. Within a dendritic layer, peak spine head calcium levels are either a linear or sigmoidal function of the number of coactive synapses, with nonlinearity depending on the ability of voltage spread in the dendrites to reach calcium spike threshold. This is strongly controlled by the potassium A-type current, with calcium spikes and the consequent sigmoidal increase in peak spine head calcium present only when the A-channel density is low. Other membrane characteristics influence the gain of the relationship between peak calcium and the number of active synapses. In particular, increasing spine neck resistance increases the gain due to increased voltage responses to synaptic input in spine heads. Colocation of stimulated synapses on a single dendritic branch also increases the gain of the response. Input pathways cooperate: CA3 inputs to the proximal apical dendrites can strongly amplify peak calcium levels due to weak EC input to the distal dendrites, but not so strongly vice versa. CA3 inputs to the basal dendrites can boost calcium levels in the proximal apical dendrites, but the relative electrical compactness of the basal dendrites results in the reverse effect being less significant. These results give pointers as to how to better describe the contributions of pre- and postsynaptic activity in the learning "rules" that apply in these cells. The calcium signal is closer in form to the activity measures used in traditional neural network learning rules than to the spike times used in spike-timing-dependent plasticity.

Improved survival and reduced phenotypic severity following AAV9/MECP2 gene transfer to neonatal and juvenile male Mecp2 knockout mice.

Typical Rett syndrome (RTT) is a pediatric disorder caused by loss-of-function mutations in the methyl-CpG binding protein 2 (MECP2) gene. The demonstrated reversibility of RTT-like phenotypes in mice suggests that MECP2 gene replacement is a potential therapeutic option in patients. We report improvements in survival and phenotypic severity in Mecp2-null male mice after neonatal intracranial delivery of a single-stranded (ss) AAV9/chicken ß-actin (CBA)-MECP2 vector. Median survival was 16.6 weeks for MECP2-treated versus 9.3 weeks for green fluorescent protein (GFP)-treated mice. ssAAV9/CBA-MECP2-treated mice also showed significant improvement in the phenotype severity score, in locomotor function, and in exploratory activity, as well as a normalization of neuronal nuclear volume in transduced cells. Wild-type (WT) mice receiving neonatal injections of the same ssAAV9/CBA-MECP2 vector did not show any significant deficits, suggesting a tolerance for modest MeCP2 overexpression. To test a MECP2 gene replacement approach in a manner more relevant for human translation, a self-complementary (sc) adeno-associated virus (AAV) vector designed to drive MeCP2 expression from a fragment of the Mecp2 promoter was injected intravenously (IV) into juvenile (4-5 weeks old) Mecp2-null mice. While the brain transduction efficiency in juvenile mice was low (~2-4% of neurons), modest improvements in survival were still observed. These results support the concept of MECP2 gene therapy for RTT.

Morphological and functional reversal of phenotypes in a mouse model of Rett syndrome.

Rett syndrome is a neurological disorder caused by mutation of the X-linked MECP2 gene. Mice lacking functional Mecp2 display a spectrum of Rett syndrome-like signs, including disturbances in motor function and abnormal patterns of breathing, accompanied by structural defects in central motor areas and the brainstem. Although routinely classified as a neurodevelopmental disorder, many aspects of the mouse phenotype can be effectively reversed by activation of a quiescent Mecp2 gene in adults. This suggests that absence of Mecp2 during brain development does not irreversibly compromise brain function. It is conceivable, however, that deep-seated neurological defects persist in mice rescued by late activation of Mecp2. To test this possibility, we have quantitatively analysed structural and functional plasticity of the rescued adult male mouse brain. Activation of Mecp2 in ∼70% of neurons reversed many morphological defects in the motor cortex, including neuronal size and dendritic complexity. Restoration of Mecp2 expression was also accompanied by a significant improvement in respiratory and sensory-motor functions, including breathing pattern, grip strength, balance beam and rotarod performance. Our findings sustain the view that MeCP2 does not play a pivotal role in brain development, but may instead be required to maintain full neurological function once development is complete.

Long-term muscle-specific overexpression of DOK7 in mice using AAV9-tMCK-DOK7.

Neuromuscular junction (NMJ) dysfunction underlies several diseases, including congenital myasthenic syndromes (CMSs) and motor neuron disease (MND). Molecular pathways governing NMJ stability are therefore of interest from both biological and therapeutic perspectives. Muscle-specific kinase (MuSK) is necessary for the formation and maintenance of post-synaptic elements of the NMJ, and downstream of tyrosine kinases 7 (DOK7) is crucial for activation of the MuSK pathway. Overexpression of DOK7 using AAV9 has been shown to ameliorate neuromuscular pathology in pre-clinical disease models of CMS and MND. However, long-term consequences of DOK7 expression have been sparsely investigated and targeted overexpression of DOK7 in skeletal muscle yet to be established. Here, we developed and characterized a novel AAV9-DOK7 facilitating forced expression of DOK7 under a skeletal muscle-specific promoter. AAV9-tMCK-DOK7 was systemically delivered to newborn mice that were monitored over 6 months. DOK7 overexpression was restricted to skeletal muscles. Body weight, blood biochemistry, and histopathological assessments were unaffected by AAV9-tMCK-DOK7 treatment. In contrast, forced expression of DOK7 resulted in enlargement of both the pre- and post-synaptic components of the NMJ, without causing denervation. We conclude that muscle-specific DOK7 overexpression can be achieved in a safe manner, with the capacity to target NMJs in vivo.

MeCP2 and Rett syndrome: reversibility and potential avenues for therapy.

Mutations in the X-linked gene MECP2 (methyl CpG-binding protein 2) are the primary cause of the neurodevelopmental disorder RTT (Rett syndrome), and are also implicated in other neurological conditions. The expression product of this gene, MeCP2, is a widely expressed nuclear protein, especially abundant in mature neurons of the CNS (central nervous system). The major recognized consequences of MECP2 mutation occur in the CNS, but there is growing awareness of peripheral effects contributing to the full RTT phenotype. MeCP2 is classically considered to act as a DNA methylation-dependent transcriptional repressor, but may have additional roles in regulating gene expression and chromatin structure. Knocking out Mecp2 function in mice recapitulates many of the overt neurological features seen in RTT patients, and the characteristic postnatally delayed onset of symptoms is accompanied by aberrant neuronal morphology and deficits in synaptic physiology. Evidence that reactivation of endogenous Mecp2 in mutant mice, even at adult stages, can reverse aspects of RTT-like pathology and result in apparently functionally mature neurons has provided renewed hope for patients, but has also provoked discussion about traditional boundaries between neurodevelopmental disorders and those involving dysfunction at later stages. In the present paper we review the neurobiology of MeCP2 and consider the various genetic (including gene therapy), pharmacological and environmental interventions that have been, and could be, developed to attempt phenotypic rescue in RTT. Such approaches are already providing valuable insights into the potential tractability of RTT and related conditions, and are useful pointers for the development of future therapeutic strategies.

Convergent cerebrospinal fluid proteomes and metabolic ontologies in humans and animal models of Rett syndrome.

MECP2 loss-of-function mutations cause Rett syndrome, a neurodevelopmental disorder resulting from a disrupted brain transcriptome. How these transcriptional defects are decoded into a disease proteome remains unknown. We studied the proteome of Rett cerebrospinal fluid (CSF) to identify consensus Rett proteome and ontologies shared across three species. Rett CSF proteomes enriched proteins annotated to HDL lipoproteins, complement, mitochondria, citrate/pyruvate metabolism, synapse compartments, and the neurosecretory protein VGF. We used shared Rett ontologies to select analytes for orthogonal quantification and functional validation. VGF and ontologically selected CSF proteins had genotypic discriminatory capacity as determined by receiver operating characteristic analysis in Mecp2 -/y and Mecp2 -/+ . Differentially expressed CSF proteins distinguished Rett from a related neurodevelopmental disorder, CDKL5 deficiency disorder. We propose that Mecp2 mutant CSF proteomes and ontologies inform putative mechanisms and biomarkers of disease. We suggest that Rett syndrome results from synapse and metabolism dysfunction.

A role for RhoB in synaptic plasticity and the regulation of neuronal morphology.

Actin-rich dendritic spines are the locus of excitatory synaptic transmission and plastic events such as long-term potentiation (LTP). Morphological plasticity of spines accompanies activity-dependent changes in synaptic strength. Several Rho GTPase family members are implicated in regulating neuronal and, in particular, spine structure via actin and the actin-binding protein cofilin. However, despite expression in hippocampus and cortex, its ability to modulate actin-regulatory proteins, and its induction during aging, RhoB has been relatively neglected. We previously demonstrated that LTP is associated with specific RhoB activation. Here, we further examined its role in synaptic function using mice with genetic deletion of the RhoB GTPase (RhoB(-/-) mice). Normal basal synaptic transmission accompanied reduced paired-pulse facilitation and post-tetanic potentiation in the hippocampus of RhoB(-/-) mice. Early phase LTP was significantly reduced in RhoB(-/-) animals, whereas the later phase was unaffected. In wild-type mice (RhoB(+/+)), Western blot analysis of potentiated hippocampus showed significant increases in phosphorylated cofilin relative to nonpotentiated slices, which were dramatically impaired in RhoB(-/-) slices. There was also a deficit in phosphorylated Lim kinase levels in the hippocampus from RhoB(-/-) mice. Morphological analysis suggested that lack of RhoB resulted in increased dendritic branching and decreased spine number. Furthermore, an increase in the proportion of stubby relative to thin spines was observed. Moreover, spines demonstrated increased length along with increased head and neck widths. These data implicate RhoB in cofilin regulation and dendritic and spine morphology, highlighting its importance in synaptic plasticity at a structural and functional level.

Altered apoptotic responses in neurons lacking RhoB GTPase.

Caspase 3 activation has been linked to the acute neurotoxic effects of central nervous system damage, as in traumatic brain injury or cerebral ischaemia, and also to the early events leading to long-term neurodegeneration, as in Alzheimer's disease. However, the precise mechanisms activating caspase 3 in neuronal injury are unclear. RhoB is a member of the Rho GTPase family that is dramatically induced by cerebral ischaemia or neurotrauma, both in preclinical models and clinically. In the current study, we tested the hypothesis that RhoB might directly modulate caspase 3 activity and apoptotic or necrotic responses in neurons. Over-expression of RhoB in the NG108-15 neuronal cell line or in cultured corticohippocampal neurons elevated caspase 3 activity without inducing overt toxicity. Cultured corticohippocampal neurons from RhoB knockout mice did not show any differences in sensitivity to a necrotic stimulus - acute calcium ionophore exposure - compared with neurons from wild-type mice. However, corticohippocampal neurons lacking RhoB exhibited a reduction in the degree of DNA fragmentation and caspase 3 activation induced by the apoptotic agent staurosporine, in parallel with increased neuronal survival. Staurosporine induction of caspase 9 activity was also suppressed. RhoB knockout mice showed reduced basal levels of caspase 3 activity in the adult brain. These data directly implicate neuronal RhoB in caspase 3 activation and the initial stages of programmed cell death, and suggest that RhoB may represent an attractive target for therapeutic intervention in conditions involving elevated caspase 3 activity in the central nervous system.

Encoding and retrieval in a model of the hippocampal CA1 microcircuit.

It has been proposed that the hippocampal theta rhythm (4-7 Hz) can contribute to memory formation by separating encoding (storage) and retrieval of memories into different functional half-cycles (Hasselmo et al. (2002) Neural Comput 14:793-817). We investigate, via computer simulations, the biophysical mechanisms by which storage and recall of spatio-temporal input patterns are achieved by the CA1 microcircuitry. A model of the CA1 microcircuit is presented that uses biophysical representations of the major cell types, including pyramidal (P) cells and four types of inhibitory interneurons: basket (B) cells, axo-axonic (AA) cells, bistratified (BS) cells and oriens lacunosum-moleculare (OLM) cells. Inputs to the network come from the entorhinal cortex (EC), the CA3 Schaffer collaterals and medial septum. The EC input provides the sensory information, whereas all other inputs provide context and timing information. Septal input provides timing information for phasing storage and recall. Storage is accomplished via a local STDP mediated hetero-association of the EC input pattern and the incoming CA3 input pattern on the CA1 pyramidal cell target synapses. The model simulates the timing of firing of different hippocampal cell types relative to the theta rhythm in anesthetized animals and proposes experimentally confirmed functional roles for the different classes of inhibitory interneurons in the storage and recall cycles (Klausberger et al., (2003, 2004) Nature 421:844-848, Nat Neurosci 7:41-47). Measures of recall performance of new and previously stored input patterns in the presence or absence of various inhibitory interneurons are employed to quantitatively test the performance of our model. Finally, the mean recall quality of the CA1 microcircuit is tested as the number of stored patterns is increased.

Reversibility of functional deficits in experimental models of Rett syndrome.

Mutations in the X-linked MECP2 gene are the primary cause of the severe autism spectrum disorder RTT (Rett syndrome). Deletion of Mecp2 in mice recapitulates many of the overt neurological features seen in humans, and the delayed onset of symptoms is accompanied by deficits in neuronal morphology and synaptic physiology. Recent evidence suggests that reactivation of endogenous Mecp2 in young and adult mice can reverse aspects of RTT-like pathology. In the current perspective, we discuss these findings as well as other genetic, pharmacological and environmental interventions that attempt phenotypic rescue in RTT. We believe these studies provide valuable insights into the tractability of RTT and related conditions and are useful pointers for the development of future therapeutic strategies.

A kinome-wide screen identifies a CDKL5-SOX9 regulatory axis in epithelial cell death and kidney injury.

Renal tubular epithelial cells (RTECs) perform the essential function of maintaining the constancy of body fluid composition and volume. Toxic, inflammatory, or hypoxic-insults to RTECs can cause systemic fluid imbalance, electrolyte abnormalities and metabolic waste accumulation- manifesting as acute kidney injury (AKI), a common disorder associated with adverse long-term sequelae and high mortality. Here we report the results of a kinome-wide RNAi screen for cellular pathways involved in AKI-associated RTEC-dysfunction and cell death. Our screen and validation studies reveal an essential role of Cdkl5-kinase in RTEC cell death. In mouse models, genetic or pharmacological Cdkl5 inhibition mitigates nephrotoxic and ischemia-associated AKI. We propose that Cdkl5 is a stress-responsive kinase that promotes renal injury in part through phosphorylation-dependent suppression of pro-survival transcription regulator Sox9. These findings reveal a surprising non-neuronal function of Cdkl5, identify a pathogenic Cdkl5-Sox9 axis in epithelial cell-death, and support CDKL5 antagonism as a therapeutic approach for AKI.