Document text
Principal Investigator: Gary Thomas
Organization: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
Fiscal Year: 2024
Award: $564,186
Funding agency: National Institute of Neurological Disorders and Stroke
Project Summary
PACS2 syndrome is a recently identified neurodevelopmental disorder caused by a recurrent de novo missense
mutation in PACS2 (p.Glu209Lys (PACS2E209K)). Patients carrying this missense mutation suffer from
developmental and epileptic encephalopathy-66 (DEE66), and share several deficits, including neonatal seizures,
global developmental delay, hypotonia, autism, and cerebellar dysgenesis. The mechanism by which
PACS2E209K causes PACS2 syndrome is unknown, and no curative treatment is available. PACS2 is a
multifunctional sorting protein that is essential for formation of ER-mitochondria contacts (MAMs), and localizes
calcium signaling molecules to the ER, including the calcium-permeable channel PKD2. MAMs are dynamic
quasi-synaptic structures that localize mTORC2/Akt, which phosphorylates PACS2 to modulate calcium transfer
and lipid metabolism. MAM-localized PACS2 acts as a phosphorylation state-dependent metabolic switch that
coordinates SIRT1/PGC-1α-dependent oxidative metabolism with MAM integrity and function. The E209K
substitution is located in a critical autoregulatory domain that controls binding of PACS2 to its client proteins.
Preliminary studies suggest PACS2 interacts with PKD2 to mediate ER-mitochondria calcium transfer and that
the E209K substitution reduces MAM contacts and diverts calcium into the cytosol. Consequently, PACS2E209K
increases aerobic glycolysis in both patient cells and the cortex of Pacs2E209K/+ mice. Consistent with these
findings, preliminary electrophysiology studies reveal PACS2E209K increases glutamatergic synaptic inputs in L2/3
cortical neurons. Together, these findings suggest PACS2E209K disturbs MAM function, impacting neuronal
metabolism, neurotransmission, and behavior. Our long-term goal is to understand how PACS2E209K causes
disease and to use this information to develop effective therapies. The objective of this particular application is
to determine how PACS2E209K dysregulates ER-mitochondrial calcium transfer to disturb CNS metabolism and
behavior. We hypothesize that PACS2E209K disturbs PKD2 function to dysregulate MAM integrity and calcium
diffusion dynamics, which consequently switches the brain to glycolytic metabolism and dysregulates
neurotransmission resulting in behavioral deficits. Guided by strong preliminary data, we will test our hypothesis
by pursuing three specific aims: 1) Determine how PACS2E209K alters ER-mitochondria calcium coupling in
isolated neurons and ex vivo L2/3 slice cultures, 2) Determine the impact of PACS2E209K on fuel handling by
measuring glucose and lactate dynamics, mitochondrial metabolism and gene expression, and 3) Determine
how PACS2E209K alters synaptic activity, learning and behavior. The approach is innovative because we will
combine advanced live-cell imaging with metabolic profiling and behavioral assessments of the first mouse
model for PACS2 syndrome to elucidate the mechanism by which the recurrent E209K substitution causes
neuronal dysfunction. This research is significant because it will advance our mechanistic understanding of a
critical cellular process that might be targeted to treat this debilitating disorder.
Terms: <2-photon microscopy><ASD><Acute><Antisense Agent><Antisense Oligonucleotides><Autism><Autistic Disorder><Behavior><Behavior assessment><Behavioral><Binding><Brain><Brain Nervous System><Buffers><Calcium><Calcium Ion Signaling><Calcium Signaling><Cell Body><Cell Function><Cell Physiology><Cell Process><Cell Respiration><Cells><Cellular Function><Cellular Physiology><Cellular Process><Cellular Respiration><Cellular biology><Client><Communication><Complex><Coupling><Cytoplasm><Cytosol><D-Glucose><Data><Decreased Muscle Tone><Defect><Development><Dextrose><Diffusion><Disease><Disorder><EEG><Early Infantile Autism><Electroencephalogram><Electroencephalography><Electrophysiology><Electrophysiology (science)><Encephalon><Epilepsy><Epileptic Seizures><Epileptics><Frequencies><Gene Alteration><Gene Expression><Gene Mutation><Genetic Alteration><Genetic Change><Genetic defect><Genus Hippocampus><Glucose><Glutamates><Goals><Hypomyotonia><Hypotonia><Impairment><Infantile Autism><Intellectual disability><Intellectual functioning disability><Intellectual limitation><Intermediary Metabolism><Intervention><Intervention Strategies><Kanner's Syndrome><L-Glutamate><Learning><Measures><Mediating><Membrane Protein Traffic><Membrane Traffic><Metabolic><Metabolic Processes><Metabolism><Mice><Mice Mammals><Missense Mutation><Mission><Mitochondria><Modeling><Molecular><Molecular Interaction><Murine><Mus><Muscle Hypotony><Muscle Tone Poor><Muscle hypotonia><Muscular Hypotonia><Mutation><NADH><NIH><National Institutes of Health><Nerve Cells><Nerve Impulse Transmission><Nerve Transmission><Nerve Unit><Neural Cell><Neural Transmission><Neurocyte><Neurodevelopmental Disorder><Neurological Development Disorder><Neuronal Dysfunction><Neuronal Transmission><Neurons><Neurophysiology / Electrophysiology><Nuclear><PKD2 protein><Pathway interactions><Patients><Permeability><Phosphorylation><Protein Phosphorylation><Protein Sortings><Proteins><Public Health><Pyramidal neuron><RNA Seq><RNA based therapeutics><RNA based therapy><RNA sequencing><RNA therapy><RNAseq><Recurrence><Recurrent><Research><Rest><SIRT1><SIRT1 gene><Seahorse><Seizure Disorder><Signaling Molecule><Sirtuin 1><Slice><Source><Structure><Subcellular Process><Synapses><Synaptic><Synaptic Transmission><Syndrome><Testing><Therapeutic><Transfection><United States National Institutes of Health><aerobic glycolysis><aerobic metabolism><aerobic respiration><antisense oligo><autism spectral disorder><autism spectrum disorder><autistic spectrum disorder><axon signaling><axon-glial signaling><axonal signaling><base><bases><behavior study><behavior test><behavioral assessment><behavioral study><behavioral test><cell biology><curative intervention><curative therapeutic><curative therapy><curative treatments><developmental><diffused><diffuses><diffusing><diffusions><effective therapy><effective treatment><electrophysiological><epilepsia><epileptic encephalopathies><epileptogenic><fat metabolism><fundamental research><genome mutation><glia signaling><glial signaling><global developmental delay><glutamatergic><hippocampal pyramidal neuron><inhibitor drug><inhibitor therapeutic><inhibitor therapy><innovate><innovation><innovative><intellectual and developmental disability><interventional strategy><limited intellectual functioning><lipid metabolism><live cell image><live cell imaging><live cellular image><live cellular imaging><metabolism measurement><metabolomics><metabonomics><mitochondrial><mitochondrial metabolism><mouse model><murine model><neonatal seizure><nerve cell metabolism><nerve signaling><neural dysfunction><neural signaling><neurodevelopmental disease><neuron cell metabolism><neuron metabolism><neuronal><neuronal cell metabolism><neuronal metabolism><neuronal signaling><neurotransmission><newborn seizure><oxidative metabolism><pathway><pharmacologic><polycystic kidney disease 2 protein><polycystin 2><postsynaptic><synapse><therapeutic RNA><tool><transcriptome sequencing><transcriptomic sequencing><translational framework><two photon excitation microscopy><two photon microscopy>