Biology subjects

Maness, P. F.

Publications and source records attributed to Maness, P. F..

6 recordsLinked to original sources

bioRxiv2025

Structural Interactions of Ankyrin B with NrCAM and β2 Spectrin

Ank2 is a high confidence autism spectrum disorder (ASD) gene encoding the spectrin-actin scaffold protein Ankyrin B (AnkB). The 220 kDal isoform of AnkB has multiple functions including developmental spine pruning through L1 family cell adhesion molecules (L1-CAMs) and class 3 Semaphorins on dendrites of pyramidal neurons to achieve an appropriate excitatory balance in the neocortex. Molecular modeling employing AlphaFold was used to predict the structure and interactions of AnkB with the cytoplasmic domain of Neuron-glial Related L1-CAM (NrCAM), and with {beta}2-Spectrin. The validity of the models was assessed by analyzing protein-protein interactions by co-immunoprecipitation from HEK293 cell lysates after mutating key residues in AnkB predicted to impair these associations. Results revealed a pocket with critical residues in the AnkB membrane-binding domain that engages NrCAM at the conserved cytoplasmic motif FIGQY. Alphafold modeling of the AnkB/{beta}2-Spectrin complex also identified key interactions between the AnkB spectrin-binding domain and {beta}2-Spectrin repeats 14-15. Selected ASD-linked mutations in AnkB predicted to impact binding to NrCAM or {beta}2-Spectrin were then assayed for protein interactions. Maternally inherited ASD missense mutations AnkB A368G located in the NrCAM binding pocket and AnkB R977Q in the Zu51 subdomain disrupted associations with NrCAM and {beta}2-Spectrin, respectively. Moreover, AnkB A368G impaired the neuronal function of 220 kDal AnkB in Semaphorin 3F-induced spine pruning in mouse cortical neuron cultures. These new findings provide structural insights into the L1-CAM/AnkB complex and the molecular basis of ASD etiology associated with AnkB missense mutations.

neuroscience↗

bioRxiv2024

Perisomatic Synaptic Targeting by Cholecystokinin-Basket Interneurons through NrCAM and Ankyrin B

The perisomatic region of cortical pyramidal neurons (PNs) integrates local and long-range inputs and regulates firing. This domain receives GABAergic inputs from cholecystokinin (CCK)- and Parvalbumin (PV)-expressing basket cells (BCs) but how synaptic contacts are established is unclear. Neuron-glial related cell adhesion molecule (NrCAM) is a homophilic transmembrane protein that binds the scaffold protein Ankyrin B. Here we show that NrCAM and Ankyrin B mediate perisomatic synaptic contact between CCK-BCs and PNs in mouse medial prefrontal cortex (mPFC). Immunolabeling of CCK-BC terminals for vesicular glutamate transporter-3 (VGLUT3) or vesicular GABA transporter (VGAT) revealed a significant decrease in CCK-BC synaptic puncta on PN soma in NrCAM-null mice, however no decrease in PV-BC puncta or cell loss. VGLUT3+ CCK-BC puncta were also decreased by Ankyrin B deletion from PNs in Nex1Cre-ERT2:Ank2flox/flox:EGFP mice. A novel CCK-BC reporter mouse expressing tdTomato (tdT) at the Synuclein-{gamma} (Sncg) locus showed NrCAM localized to Sncg+ CCK-BCs, and to postsynaptic PN soma in Nex1Cre-ERT2:Ank2+/+:EGFP mice. Results suggest that NrCAM and Ankyrin B contribute to the establishment of connectivity between CCK-BCs and excitatory neurons of the mPFC.

neuroscience↗

bioRxiv2023

Ankyrin B Promotes Developmental Spine Regulation in the Mouse Prefrontal Cortex

Postnatal regulation of dendritic spine formation and refinement in cortical pyramidal neurons is critical for excitatory/inhibitory balance in neocortical networks. Recent studies have identified a selective spine pruning mechanism in the mouse prefrontal cortex (PFC) mediated by class 3 Semaphorins and the L1-CAM cell adhesion molecules Neuron-glia related CAM (NrCAM), Close Homolog of L1 (CHL1), and L1. L1-CAMs bind Ankyrin B (AnkB), an actin-spectrin adaptor encoded by Ankyrin2 (ANK2), a high confidence gene for autism spectrum disorder (ASD). In a new inducible mouse model (Nex1Cre-ERT2: Ank2flox: RCE), Ank2 deletion in early postnatal pyramidal neurons increased spine density on apical dendrites in PFC layer 2/3 of homozygous and heterozygous Ank2-deficient mice. In contrast, Ank2 deletion in adulthood had no effect on spine density. Sema3F-induced spine pruning was impaired in cortical neuron cultures from AnkB-null mice and was rescued by re-expression of the 220 kDa AnkB isoform but not 440 kDa AnkB. AnkB bound to NrCAM at a cytoplasmic domain motif (FIGQY1231), and mutation to FIGQH inhibited binding, impairing Sema3F-induced spine pruning in neuronal cultures. Identification of a novel function for AnkB in dendritic spine regulation provides insight into cortical circuit development, as well as potential molecular deficiencies in ASD.

neuroscience↗

bioRxiv2022

The L1 Cell Adhesion Molecule Constrains Dendritic Spine Density through Ankyrin Binding in Pyramidal Neurons of the Mouse Cerebral Cortex

A novel function for L1 cell adhesion molecule and its interaction with Ankyrin, an actin-spectrin adaptor protein, was identified in constraining dendritic spine density on pyramidal neurons in the mouse neocortex. In an L1-null mouse mutant increased spine density was observed on apical but not basal dendrites of pyramidal neurons in diverse cortical areas (prefrontal cortex layer 2/3, motor cortex layer 5, visual cortex layer 4).The Ankyrin binding motif (FIGQY) in L1s cytoplasmic domain was critical for spine formation, as demonstrated by increased spine density in the prefrontal cortex of a mouse mutant (L1YH) harboring a tyrosine to histidine mutation in this motif, which disrupts L1-Ankyrin association. This mutation is a known variant in the human L1 syndrome. In both mutants mature mushroom spines rather than immature spines were predominant. L1 was detected in spines and dendrites of wild-type prefrontal cortical neurons by immmunostaining. L1 coimmunoprecipitated with Ankyrin B (220 kDa) from cortical lysates of wild-type but not L1YH mice. Spine pruning assays in cortical neuron cultures from wild-type and L1YH mutant mice showed that the L1-Ankyrin interaction mediated spine retraction in response to the class 3 Semaphorins, Sema3F and to a lesser extent Sema3B. These ligands also induce spine pruning through other L1 family adhesion molecules, NrCAM and Close Homolog of L1 (CHL1), respectively. This study provides insight into the molecular mechanism of spine regulation and underscore the potential for this adhesion molecule to regulate cognitive and other L1-related functions that are abnormal in the L1 syndrome.

neuroscience↗

bioRxiv2022

Doublecortin-like kinase 1 (DCLK1) Facilitates Dendritic Spine Growth of Pyramidal Neurons in Mouse Prefrontal Cortex

The L1 cell adhesion molecule NrCAM (Neuron-glia related cell adhesion molecule) functions as a co-receptor for secreted class 3 Semaphorins to prune subpopulations of dendritic spines on apical dendrites of pyramidal neurons in the developing mouse neocortex. The developing spine cytoskeleton is enriched in actin filaments but a small number of microtubules have been shown to enter the spine apparently trafficking vesicles to the membrane. Doublecortin-like kinase 1 (DCLK1) is a member of the Doublecortin (DCX) family of microtubule-binding proteins with serine/threonine kinase activity. To determine if DCLK1 plays a role in spine remodeling, we generated a tamoxifen-inducible mouse line (Nex1Cre-ERT2: DCLK1flox/flox : RCE) to delete microtubule binding isoforms of DCLK1 from pyramidal neurons during postnatal stages of spine development. Homozygous DCLK1 conditional mutant mice exhibited decreased spine density on apical dendrites of pyramidal neurons in the prefrontal cortex (layer 2/3). Mature mushroom spines were selectively decreased upon DCLK1 deletion but dendritic arborization was unaltered. Mutagenesis and binding studies revealed that DCLK1 bound NrCAM at the conserved FIGQY1231 motif in the NrCAM cytoplasmic domain, a known interaction site for the actin-spectrin adaptor Ankyrin. These findings demonstrate that DCLK1 facilitates spine growth and maturation on cortical pyramidal neurons in the mouse prefrontal cortex potentially through microtubule and NrCAM interactions.

neuroscience↗

bioRxiv2021

Semaphorin3F Drives Dendritic Spine Pruning through Rho-GTPase Signaling

Dendritic spines of cortical pyramidal neurons are initially overproduced then remodeled substantially in the adolescent brain to achieve appropriate excitatory balance in mature circuits. Here we investigated the molecular mechanism of developmental spine pruning by Semaphorin 3F (Sema3F) and its holoreceptor complex, which consists of immunoglobulin-class adhesion molecule NrCAM, Neuropilin-2 (Npn2), and PlexinA3 (PlexA3) signaling subunits. Structure-function studies of the NrCAM-Npn2 interface showed that NrCAM stabilizes binding between Npn2 and PlexA3 necessary for Sema3F-induced spine pruning. Using a mouse neuronal culture system, we identified a dual signaling pathway for Sema3F-induced pruning, which involves activation of Tiam1-Rac1-PAK1-3 -LIMK1/2-Cofilin1 and RhoA-ROCK1/2-Myosin II in dendritic spines. Inhibitors of actin remodeling impaired spine collapse in the cortical neurons. Elucidation of these pathways expands our understanding of critical events that sculpt neuronal networks and may provide insight into how interruptions to these pathways could lead to spine dysgenesis in diseases such as autism, bipolar disorder, and schizophrenia.

neuroscience↗