Deciphering signaling cascades involved in neuronal maturation and plasticity
Multiple signaling molecules in the brain, including those that participate in the WNT, Ca2+, and mTOR pathways, serve as essential players during neuronal maturation and synapse formation and maintenance. These molecules are capable of interacting with and regulating a host of protein interactions in the cell over the course of neuronal development through their ability to scaffold and/or post-translationally modify targets in order to engage in signal transduction. Oftentimes, they perform distinct functions in different cellular compartments at different developmental times. Notably, human mutations in these genes are apparent in patients with severe neurodevelopmental disorders, thereby affecting a large percentage of the human population. Currently, we are focusing on investigating the molecular and cellular mechanisms of three distinct, incompletely understood, disease-associated protein kinases/scaffolding molecules during neuronal maturation and synaptic maintenance and plasticity – CASK (Calcium/Calmodulin Dependent Serine Protein Kinase), TBCK (TBC1 Domain Containing Kinase), and CaMKII (Calcium/Calmodulin Dependent Protein Kinase-II).
Electrophysiological analysis of CASK KO human neurons. Figure from McSweeney et al., 2022
CASK as a regulator of neuronal maturation and synaptic function
We have recently reported that loss-of-function (LOF) mutations of CASK differentially regulate neuronal maturation and synaptic function in human induced cortical excitatory neurons (McSweeney et al., 2022, iScience). CASK LOF induces neuronal overgrowth in early maturing cells, suggesting that CASK functions to restrict dendritic morphogenesis at this developmental time window. Transcriptomic analysis of CASK LOF immature neurons revealed that these KO cells resemble morphology and gene expression programs associated with hyperactivated WNT signaling pathway. We are currently testing which proteins are being regulated by CASK during neuronal maturation. Post neuronal maturation, neurons lacking CASK show significant impairment in neurotransmitter release and network connectivity. Synaptic signaling is impaired in these cells similar to what has been observed in rodents. From this project, we are learning that protein kinases important for neurodevelopment perform distinct functions during neuronal growth and synapse maturation, which encourages us to understand the developmental timing-dependent functions of these molecules.
Human TBCK protein structure. The three TBCK protein domains are shown, as well as the location of mutations that cause TBCK Syndrome in humans.
Solving the function of “black box” molecule TBCK
In collaboration with Dr. Gerry Downes (UMass Biology), we are establishing dual model systems (zebrafish and human-induced neurons) to investigate the developmental and synaptic roles of a novel kinase TBCK. TBCK is the gene responsible for the TBCK neurodevelopmental syndrome, whose function has yet to be characterized since the first cloning of the gene in 2013. Although preliminary results from patient fibroblasts suggest misregulation of mTOR signaling upon disruption of TBCK, it is still a black box as to what TBCK does, where it functions, and which targets it regulates. Patients with TBCK syndrome suffer from loss of muscle tone, intellectual disability, and drug resistant epilepsy and currently, there are no treatments for this disorder. In this project, my lab is establishing the very first human induced neuronal models of TBCK syndrome by differentiating edited iPSC lines harboring pathogenic mutations found in TBCK syndrome patients. We are probing the molecular and cellular functions of this molecule in human induced neurons and determining which signaling pathways are under the regulatory control of TBCK during neuronal development and synapse maturation. Any information we gain from this study will be tremendously helpful for advancing TBCK biology and identifying potential therapeutic development for these patients.
Understanding the regulatory mechanisms of CaMKII during synaptic plasticity in health and disease states
Through collaboration with the Stratton/Strieter labs at UMass, we are investigating how the memory molecule CAMKII is regulated at the synapse and how pathogenic mutations found in ASD patients impact its function.
Funding sources supporting these projects include:
NIMH R01, Armstrong Science Fund, IALS M2M Midi grant
Highlighted papers:
McSweeney et al., 2022, Iossifov et al., 2014 (PMID 25363768), Bhoj et al., 2016 (PMID 27040691)