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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic <t>(PSD-95,</t> green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.
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Image Search Results


(A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic (PSD-95, green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.

Journal: The Journal of cell biology

Article Title: Pathogenic KIF1A variants differentially disrupt axonal trafficking and impede synaptic development

doi: 10.1083/jcb.202601034

Figure Lengend Snippet: (A) Experimental schematic and representative immunocytochemistry images of presynaptic synapsin I/II (SYN, magenta), postsynaptic (PSD-95, green), and somatodendritic compartments (MAP2, blue) in WT, KIF1A-C92* , - P305L , and - R350G iNeurons co-cultured with primary rat astrocytes. Insets display representative presynaptic accumulations apposed to postsynaptic puncta (WT, P305L, and R350G) or alone (C92*). (B, C) Syn puncta density (B) and bona fide synapse density consisting of apposed pre/postsynaptic puncta (C) normalized to MAP2 area for WT or KIF1A mutant iNeurons. Pre- and postsynaptic puncta were defined using SynapseJ quantification on z-stack images. Plots display mean ± standard deviation of experimental replicates, n = 30 imaging fields from 3 independent experiments, reported p -values determined using linear mixed effect model. (D) Percentage of SYN puncta apposed to a postsynaptic compartment (magenta) or alone (light pink). Despite differences observed for SYN puncta/synapse density between loss-of-function and gain-of-function KIF1A mutants, each mutation disrupts the ratio of presynaptic puncta aligning with postsynaptic partners. Reported p -values were determined using Chi-Square test of independence.

Article Snippet: The following primary antibodies were used: NF-H (mouse anti-NF-H; BioLegend, 801601; 1:1000), Synapsin I/II (guinea pig anti-Synapsin I/II; Synaptic Systems, 106-004; 1:1000), PSD-95 (rabbit anti-PSD-95; Synaptic Systems, 124-008; 1:500), MAP2 (mouse anti-MAP2; EMD Millipore, MAB3418; 1:200), Synaptophysin (mouse anti-Synaptophysin; Sigma-Aldrich, S5768; 1:200), and Synaptobrevin-2 (rabbit anti-Synaptobrevin-2; Cell Signaling, 13508; 1:250).

Techniques: Immunocytochemistry, Cell Culture, Mutagenesis, Standard Deviation, Imaging