Journal: Psychiatry Investigation

Article Title: Memory Decline and Aberration of Synaptic Proteins in X-Linked Moesin Knockout Male Mice

doi: 10.30773/pi.2024.0186

Figure Lengend Snippet: Moesin-KO decreased the expression of synaptic proteins and phosphorylation of signaling proteins in the adult brain. A and B: The expression or activation level of synaptic vesicle proteins such as Syntaxin 1A, Synaptophysin, CDK5, and radixin were confirmed by immunoblotting in the cortex and hippocampus to examine the downstream target of moesin. Relative expression levels of each protein are presented in bar graphs. C and D: The expression and phosphorylation level of synapsin I were examined in the cortex and hippocampus of moesin-KO and presented in bar graphs. E and F: The activation levels of signaling molecules such as ERK and CREB were checked with phosphorylationspecific antibodies. The expression and phosphorylation levels of MUNC18 and PSD95 were examined in the cortex, and hippocampus of moesin-KO, and their relative expression is presented in graphs. All data were analyzed by t-test and are presented as the mean±SEM of three replicates. **p<0.01; ***p<0.001. KO, knockout mice; WT, wild-type mice; CDK5, Cyclin dependent kinase 5; ERK, extracellular signal-regulated kinase; CREB, cAMP response element-binding protein; SEM, standard error of the mean.

Article Snippet: The tissue lysates (30 μg) were electrophoresed and transferred to 0.45 μm nitrocellulose membranes, which were incubated with anti-GFAP (ab7260, 1:1,000, Abcam, Cambridge, MA, USA), anti-MAP2 (ab5392, 1:1,000, Abcam), anti-Sox2 (#4900, CST, 1:1,000), anti-DCX (#91954, CST, 1:1,000), anti-soluble NSF adapter protein 25 (SNAP25) (#5308, CST, 1:1,000), anti-postsynaptic density protein 95 (PSD95) (#3409, CST, 1:1,000), anti-phospho-PSD95 (#45737, CST, 1:1,000), or anti-moesin (#3146, CST, 1:1,000), anti-syntaxin1A (#18572, CST, 1:1,000), anti-synaptophysin (#5467, CST, 1:1,000), anti-CDK5 (#14145, CST, 1:1,000), anti-radixin (#2636, CST, 1:1,000), anti-synapsin 1 (#2312, CST, 1:1,000), anti-phospho-synapsin 1 (Ser9) (#2311, CST, 1:1,000), anti-phospho-synapsin 1 (Ser605) (#88246, CST, 1:1,000), anti-phospho-synapsin 1 (Ser549) (#PA1-4697, 1:1,000; Thermo Fisher Scientific, Waltham, MA, USA), anti-ERK (#4695, CST, 1:1,000), anti-phospho-ERK (#4370, CST, 1:1,000), anti-CREB (#9197, CST, 1:1,000), anti-phospho-CREB (#9190, CST, 1:1,000), anti-mammalian uncoordinated18 (MUNC18) (#13414, CST, 1:1,000), anti-phospho-MUNC18 (ab183484, 1:1,000, Abcam) anti-β-actin (#4967, CST, 1:1,000) for 16 hours at 4°C.

Techniques: Expressing, Phospho-proteomics, Activation Assay, Western Blot, Knock-Out, Binding Assay