Fig. 3: SLAMR and kinesin KIF5C reciprocally regulate each other. | Nature Communications

Fig. 3: SLAMR and kinesin KIF5C reciprocally regulate each other.

From: Synaptically-targeted long non-coding RNA SLAMR promotes structural plasticity by increasing translation and CaMKII activity

Fig. 3

A Top: Experimental timeline. Bottom: qRT-PCR results show changes in SLAMR RNA levels in total homogenates and synaptic fractions of primary mouse neurons after kinesin silencing (n = 3 biological replicates for each except Hom siKif2a n = 2; *p = 0.0184, **p = 0.0002; One-way ANOVA + Dunnett’s test; mean ± SEM). B Experimental timeline for KIF5C knockdown and MS2-SLAMR:MCP-RFP transport in rat hippocampal neurons. C, D Anterograde and retrograde velocity of MS2-SLAMR in KIF5C-KD or scrambled control neurons (N = 10 neurons each; Two-tailed Student’s t-test; mean ± SEM). E, F The distance from the soma that anterograde (E) and retrograde tracks (F) of MS2-SLAMR begin transport in KIF5C-KD and scrambled control neurons. (N = 10 neurons each; Two-tailed Student’s t-test; mean ± SEM). G The percentage of mobile MS2-SLAMR:MCP-RFP in KIF5C-KD and scrambled control neurons (N = 10 neurons each; Two-tailed Student’s t-test; mean ± SEM). H Normalized fold change of Kif5c mRNA abundance in mouse neuronal cultures transduced by shSLAMR, NC-GFP lentivirus, or treatment with PBS. (PBS and SLAMR-KD n = 3, NC-GFP n = 4 biological replicates; One-way ANOVA + Tukey’s test; mean ± SEM). I Normalized fold change of Kif5c mRNA abundance in mouse neuronal cultures transduced by lentivirus containing SLAMR-OE compared to NC-GFP or treatment with PBS. (PBS and NC-GFP n = 4, SLAMR-OE n = 3; One-way ANOVA + Tukey’s test; mean ± SEM). J Model of the reciprocal regulation of SLAMR and KIF5C. K Diagram illustrating single spine stimulation DIV18-21 rat neurons transfected with RCaMP1.07 (red) and MS2-SLAMR:MCP (white). Individual spines were stimulated with 30 pulses of 2-photon excitation to uncage MNI-Caged glutamate and evaluate transport dynamics of SLAMR to dendritic spines. L Single frames of an unresponsive spine with no increase in volume and a responsive spine that did grow following local glutamate uncaging, prestimulation, and 5 min poststimulation. Red=RCaMP1.07, White=MS2-SLAMR:MCP granules. White asterisk= stimulated spine. M The change in number of MS2-SLAMR:MCP granules prestimulation to 5 min poststimulation in a 5 µm dendritic region of the stimulated spine. (n = 11 neurons each; Two-tailed Student’s t-test; mean ± SEM). N The change in number of MS2-SLAMR:MCP granules moving toward the stimulated spine within 5 min after stimulation in a 25 µm region of the stimulated spine (n = 6 neurons each; Two-tailed Student’s t-test; mean ± SEM).

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