Browsing by Author "Hens, Basant"

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    Proximity labeling and orthogonal nanobody pulldown (ID-oPD) approaches to map the spinophilin interactome uncover a putative role for spinophilin in protein homeostasis
    (bioRxiv, 2025-01-23) Claeboe, Emily T.; Blake, Keyana L.; Shah, Nikhil R.; Morris, Cameron W.; Hens, Basant; Atwood, Brady K.; Absalon, Sabrina; Mosley, Amber L.; Doud, Emma H.; Baucum, Anthony J., II; Biochemistry and Molecular Biology, School of Medicine
    Spinophilin is a dendritic spine enriched scaffolding and protein phosphatase 1 targeting protein. To detail spinophilin interacting proteins, we created an Ultra-ID and ALFA-tagged spinophilin encoding construct that permits proximity labeling and orthogonal nanobody pulldown (ID-oPD) of spinophilin-associated protein complexes in heterologous cells. We identified 614 specific, and 312 specific and selective, spinophilin interacting proteins in HEK293 cells and validated a subset of these using orthogonal approaches. Many of these proteins are involved in mRNA processing and translation. In the brain, we determined that spinophilin mRNA is highly neuropil localized and that spinophilin may normally function to limit its own expression but promote the expression of other PSD-associated proteins. Overall, our use of an ID-oPD approach uncovers a novel putative role for spinophilin in mRNA translation and synaptic protein expression specifically within dendritic spines.
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    Spinophilin Limits Metabotropic Glutamate Receptor 5 Scaffolding to the Postsynaptic Density and Cell Type Specifically Mediates Excessive Grooming
    (Elsevier, 2023) Morris, Cameron W.; Watkins, Darryl S.; Shah, Nikhil R.; Pennington, Taylor; Hens, Basant; Qi, Guihong; Doud, Emma H.; Mosley, Amber L.; Atwood, Brady K.; Baucum, Anthony J., II; Pharmacology and Toxicology, School of Medicine
    Background: Grooming dysfunction is a hallmark of the obsessive-compulsive spectrum disorder trichotillomania. Numerous preclinical studies have utilized SAPAP3-deficient mice for understanding the neurobiology of repetitive grooming, suggesting that excessive grooming is caused by increased metabotropic glutamate receptor 5 (mGluR5) activity in striatal direct- and indirect-pathway medium spiny neurons (MSNs). However, the MSN subtype-specific signaling mechanisms that mediate mGluR5-dependent adaptations underlying excessive grooming are not fully understood. Here, we investigated the MSN subtype-specific roles of the striatal signaling hub protein spinophilin in mediating repetitive motor dysfunction associated with mGluR5 function. Methods: Quantitative proteomics and immunoblotting were utilized to identify how spinophilin impacts mGluR5 phosphorylation and protein interaction changes. Plasticity and repetitive motor dysfunction associated with mGluR5 action were measured using our novel conditional spinophilin mouse model in which spinophilin was knocked out from striatal direct-pathway MSNs and/or indirect-pathway MSNs. Results: Loss of spinophilin only in indirect-pathway MSNs decreased performance of a novel motor repertoire, but loss of spinophilin in either MSN subtype abrogated striatal plasticity associated with mGluR5 function and prevented excessive grooming caused by SAPAP3 knockout mice or treatment with the mGluR5-specific positive allosteric modulator VU0360172 without impacting locomotion-relevant behavior. Biochemically, we determined that the spinophilin-mGluR5 interaction correlates with grooming behavior and that loss of spinophilin shifts mGluR5 interactions from lipid raft-associated proteins toward postsynaptic density proteins implicated in psychiatric disorders. Conclusions: These results identify spinophilin as a novel striatal signaling hub molecule in MSNs that cell subtype specifically mediates behavioral, functional, and molecular adaptations associated with repetitive motor dysfunction in psychiatric disorders.