Journal: Neurochemical Research

Article Title: Morin Improves Cognitive Deficits in an in Vivo Model of Vascular Dementia by Modulating the N-methyl-D-aspartate Receptor Signaling Pathways

doi: 10.1007/s11064-026-04717-7

Figure Lengend Snippet: Morin modulated the expression of NMDA receptors in the hippocampus of VaD rats. a the expression levels of NR1 ; b the expression levels of NR2A ; c the expression levels of NR2B ; d the expression levels of NR1 protein; e the expression levels of NR2A protein; f the expression levels of NR2B protein; g protein levels of p-CREB; h protein levels of p-CAMK2A; i protein levels of p-CAMK2D. Protein levels of NR1, NR2A, and NR2B were quantified by ELISA. Data are presented as mean ± SD ( n = 8 per group). Statistical analysis was performed by one-way ANOVA with Tukey’s post-hoc test (data met assumptions of normality and homoscedasticity)/Kruskal-Wallis with Dunn’s test (data did not meet assumptions). * indicates a significant difference from the Sham group; # indicates a significant difference 2VO group; *# indicates a significant difference from both the Sham and 2VO groups, with a p-value of less than 0.05 considered statistically significant

Article Snippet: Moreover, phosphorylation levels of calcium/calmodulin-dependent protein kinase II isoforms CAMK2A and CAMK2D at Thr286 (p-CAMK2A, p-CAMK2D) were quantified using the Colorimetric Cell-Based ELISA Kit (CAMK2A/CAMK2D (Phospho-Thr286), Boster Bio, #EKC2366).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Pharmacology

Article Title: Farnesyl Transferase Inhibitor Lonafarnib Enhances α7nAChR Expression Through Inhibiting DNA Methylation of CHRNA7 and Increases α7nAChR Membrane Trafficking

doi: 10.3389/fphar.2020.589780

Figure Lengend Snippet: Role of DNA methylation in the lonafarnib affected activity, total expression, membrane expression, and phosphorylation of a7nAChR. (A) Levels of a7nAChR total proteins in the hippocampus of control and lonafarnib-treated mice treated with vehicle, DNMT inhibitor RG108 or anisomycin. ** P < 0.01 vs. control mice, ## P < 0.01 vs. lonafarnib-treated mice (two-way ANOVA, followed by Tukey’s multiple comparison test). (B) Evoked I ACh by ACh (3 mM) in the slices of control and lonafarnib-treated mice treated with vehicle, RG108, or anisomycin, ** P < 0.01 vs. control mice; # P < 0.05 vs. lonafarnib-treated mice (two-way ANOVA, followed by Tukey’s multiple comparison test). (C) Levels of biotinylated a7nAChR (membrane surface) protein in the hippocampus of control and lonafarnib-treated mice treated with vehicle, RG108, or anisomycin. Surface a7nAChR was normalized by surface GluR2 protein, which was again normalized by vehicle-treated control group. ** P < 0.01 vs. control mice; # P < 0.05 vs. lonafarnib-treated mice (two-way ANOVA, followed by Tukey’s multiple comparison test). (D) Levels of phospho-a7nAChR in the hippocampus of control and lonafarnib-treated mice treated with vehicle, RG108, or anisomycin. The expression of protein and the amplitude of evoked I ACh were normalized by the values of control group with vehicle.

Article Snippet: CaMKII inhibitor KN93 and DNMT inhibitor RG108 were purchased from MCE.

Techniques: DNA Methylation Assay, Activity Assay, Expressing, Membrane, Phospho-proteomics, Control, Comparison

Journal: Frontiers in Pharmacology

Article Title: Farnesyl Transferase Inhibitor Lonafarnib Enhances α7nAChR Expression Through Inhibiting DNA Methylation of CHRNA7 and Increases α7nAChR Membrane Trafficking

doi: 10.3389/fphar.2020.589780

Figure Lengend Snippet: Lonafarnib administration affects CaMKII signaling pathways, partially modulating the membrane expression of a7nAChR. (A and B) Levels of phospho-PKC and phospho-PKA in the hippocampus of control and lonafarnib-treated mice treated with vehicle, RG108, or anisomycin. (C) Levels of phospho-CaMKII in the hippocampus of control and lonafarnib-treated mice treated with vehicle, RG108, or anisomycin. ** P < 0.01 vs. control mice (two-way ANOVA, followed by Tukey’s multiple comparison test). (D) Levels of biotinylated a7nAChR (membrane) protein in the hippocampus of control and lonafarnib-treated mice treated with vehicle or CaMKII pathway blocker KN93. Surface a7nAChR was normalized by surface GluR2 protein, which was again normalized by vehicle-treated control group. ** P < 0.01 vs. control mice; # P < 0.05 vs. lonafarnib-treated mice (two-way ANOVA, followed by Tukey’s multiple comparison test). (E) Levels of a7nAChR total proteins in the hippocampus of control and lonafarnib-treated mice treated with vehicle and KN93, and total a7nAChR was normalized by GAPDH, which was again normalized by vehicle-treated group. ** P < 0.01 vs. control mice (two-way ANOVA, followed by Tukey’s multiple comparison test). The expression of protein was normalized by the values of control group with vehicle.

Article Snippet: CaMKII inhibitor KN93 and DNMT inhibitor RG108 were purchased from MCE.

Techniques: Protein-Protein interactions, Membrane, Expressing, Control, Comparison

Journal: bioRxiv

Article Title: Neuron type-specific mRNA translation programs provide a gateway for memory consolidation

doi: 10.1101/2025.01.13.632784

Figure Lengend Snippet: ( a ) Schematics of the experimental approach used to study cell type-specific dynamic changes in the translatome induced by threat conditioning. ( b ) Contextual threat conditioning experiment. White = control mice placed in the experimental box, but not shocked; Black = mice that were shocked in the experimental box. Left panel = Training session; Right panel = Testing session, 24h after training session. Unpaired t test with Welch’s correction. p = 0.0090. N = 4-5 mice/group; Mean ± SEM; F = 124.8 ( c ) Immunofluorescence confirmation of cell type-specific expression of RPL10a-EGFP. Green = GFP; Blue = DAPI; ( d ) Enrichment of cell type-specific markers in the purified translatome of each cell type. Camk2a and Slc17a7 = excitatory cell markers; Gad1, Pvalb, Sst = Inhibitory cell markers; Gfap, Cnp, Cx3cr1 = Glial markers. The upper and lower whiskers represent the largest and smallest value within 1.5 times above or below the interquartile range, respectively. ( e ) PCA of baseline translatomes. Light-blue = Camk2a + ; dark green = Pvalb + ; Light-green = Sst + . N = 6-9 mice/group. ( f ) Venn diagram of the mRNAs identified in each cell type-specific purification. Color intensity illustrates the size of the set. ( g ) Spearman correlation of Log 2 FC of Camk2a + resting translatome versus Pvalb + (x axis) or Sst + (y axis). Spearman Rho = 0.8. ( h ) Gene set enrichment analysis of resting cell type-specific translatomes. Yellow means enriched in the first cell type in the comparison, magenta means enriched in the second cell type in the comparison. NES = normalized enrichment score.

Article Snippet: Camk2a.Cre ERT2 /GADD34 flfl mice were injected with AAV9.CAG.FLEX.RPL10a::eGFP (Addgene).

Techniques: Control, Immunofluorescence, Expressing, Purification, Comparison

Journal: bioRxiv

Article Title: Neuron type-specific mRNA translation programs provide a gateway for memory consolidation

doi: 10.1101/2025.01.13.632784

Figure Lengend Snippet: ( a ) Pearson’s correlation of time-dependent mRNA translation in shock (left panels) or box conditions (right panels). Light colors = mRNAs differentially regulated exclusively at the 15 min time point; bright colors = mRNAs that are differentially regulated in both time points; dark colors = mRNAs that are differentially regulated in 60 min time point. Grey = mRNAs not significantly regulated. Top left corner = R represents Pearson’s correlation coefficient; p is the significance of the correlation. ( b ) GSEA per dataset. NES = normalized enrichment score. Size of circles is mapped to -log 10 (FDR). Yellow = Upregulated terms; Magenta = Downregulated terms. ( c ) Representation of WGCNA network and the modules identified. Colors of modules are mapped to the network. Insets represent best overrepresented gene ontologies for each of the modules identified. ( d ) Map of the conditioning-induced fold in the gene network obtained through WGCNA. Red = upregulated (FC > 0.64), blue = downregulated (FC < -0.64). Top left = Camk2a + neurons 15 minutes after conditioning. Bottom left = Camk2a + neurons 60 minutes after conditioning. Top middle = Pvalb + neurons 15 minutes after conditioning. Bottom left = Pvalb + neurons 60 minutes after conditioning. Top left = Sst + neurons 15 minutes after conditioning. Bottom left = Sst + neurons 60 minutes after conditioning. ( e ) Heatmap of fold changes of all mRNAs clustered in the blue module. Magenta = downregulated genes; Gold = upregulated genes. Selected mRNAs with symbols highlighted are encompassed in GOs “ Proteasomal protein catabolic process ”, “ Endoplasmic reticulum membrane” and “Unfolded protein binding ”. Number of mice per group in the dataset used is listed in Methods section. Differential expression analysis and GSEA results were corrected for multiple comparisons using Benjamini-Hochberger post-hoc test.

Article Snippet: Camk2a.Cre ERT2 /GADD34 flfl mice were injected with AAV9.CAG.FLEX.RPL10a::eGFP (Addgene).

Techniques: Membrane, Protein Binding, Expressing

Journal: bioRxiv

Article Title: Neuron type-specific mRNA translation programs provide a gateway for memory consolidation

doi: 10.1101/2025.01.13.632784

Figure Lengend Snippet: ( a ) Rationale for eIF2α dephosphorylation-mediated increase in de novo translation. ( b ) Log 2 FC of Ppp1r15a (mRNA encoding GADD34) and Ppp1r15b (CReP) in the translatome of Camk2a + , Pvalb + or Sst + neurons of mice that underwent contextual threat conditioning against home cage baseline . ** is p = 0.0054; * is p = 0.0076 ; *** is p = 0.00009. Number of mice per group in the dataset used is listed in Methods section. Mean ± SEM. ( c ) Log 2 FC of Ppp1r15a and Ppp1r15b in the box control against home cage baseline. Number of mice per group in the dataset used is listed in Methods section. Mean ± SEM. ( d ) Schematics of contextual threat conditioning paradigm to test the cell type-specific relevance of GADD34 in long-term memory formation. ( e ) contextual threat conditioning of Camk2a.Cre ERT2 /GADD34 flfl . Left panel = training session. Right panel = testing session. N = 9-10 mice/group; Mean ± SEM; F = 2.871. ( f ) contextual threat conditioning of Pvalb.Cre/GADD34 flfl . Left panel = training session. Right panel = testing session. N = 4-8 mice/group; Mean ± SEM.; F = 1.122.( g ) contextual threat conditioning of Sst.Cre/GADD34 flfl . Left panel = training session. Right panel = testing session. N = 7-11 mice/group; Mean ± SEM; F = 2.319. ( h ) Experimental plan for TRAP-seq of excitatory neurons from the dHPC of Camk2a.Cre ERT2 /GADD34 flfl mice. N = 3-5 mice/group. Mean ± SEM. ( i ) Volcano plot illustrating the DEGs found in WT (left panel) and KO (right panel). Light blue = downregulated. Dark blue = upregulated. Grey = not significant. ( j ) cumulative distribution fraction of shock vs home cage log2 fold changes in WT (orange line) and KO (dark blue line). ( k ) Dot plot evidencing DEGs in WT that are differentially modulated in KO mice subjected to contextual threat conditioning. Q1-4 = Quadrant numbers. Dashed lines represent abs(Log 2 FC) > 1. ( l ) Comparative cumulative distribution of genes belonging to Q1. ( m ) Map of the log 2 FC values in the WGCNA network previously obtained. The WT network is similar to what was previously found. Red = upregulated (log 2 FC > 0.35), blue = downregulated (log 2 FC < -0.35).

Article Snippet: Camk2a.Cre ERT2 /GADD34 flfl mice were injected with AAV9.CAG.FLEX.RPL10a::eGFP (Addgene).

Techniques: De-Phosphorylation Assay, Control

Journal: bioRxiv

Article Title: Neuron type-specific mRNA translation programs provide a gateway for memory consolidation

doi: 10.1101/2025.01.13.632784

Figure Lengend Snippet: ( a) Sketch illustrating the design of the machine learning model. Adapted from ref . ( b ) On the left: Receiver Operating Characteristics (ROC) curves on the left, categorized by condition tested. Diagonal line represents random classification performance (50% chance). On the right: Accuracy per cross validation performed ( k = 10). Horizontal dashed line represents random classification performance. Blue shades graphs = Camk2a ; Green shades graphs = Pvalb ; Red shades graphs = Sst . ( c ) Permutation importance test ( k = 10). ( d ) Average value of each mRNA feature per GO that was upregulated 15- or 60-min following training. Blue graphs = GOs upregulated in Camk2a + neurons; Green graphs = GOs upregulated in Pvalb + neurons; Red graphs = GOs upregulated in Sst + neurons. Light colors represent 15-minute time points, darker colors represent the 60-minute time point. Number of mice per group in the dataset used is listed in Methods section. NS = not significant; * = p <0.05; ** = p <0.01; *** = p <0.001.. Student’s t test, unpaired. The upper and lower whiskers represent the largest and smallest value within 1.5 times above or below the interquartile range, respectively.

Article Snippet: Camk2a.Cre ERT2 /GADD34 flfl mice were injected with AAV9.CAG.FLEX.RPL10a::eGFP (Addgene).

Techniques:

Journal: bioRxiv

Article Title: Neuron type-specific mRNA translation programs provide a gateway for memory consolidation

doi: 10.1101/2025.01.13.632784

Figure Lengend Snippet: (a) Average fold changes of mRNAs predicted to be targets of ARE-RBPs in Camk2a + , Pvalb + and Sst + neurons (names of the RBPs are highlighted inside each plot). Top left corner = R represents Pearson’s correlation coefficient; p is the significance of the correlation. On the right of each plot, density plots show fold change distribution of targets of hnRNPD (colored) versus non-targets (in gray). Number of mice per group in the dataset used is listed in Methods section. (b) Schematics of bi-directional manipulation approach to target hnRNPD expression in a cell type-specific way. Green neurons = GFP + neurons; red neurons = mCherry + neurons; yellow neurons = GFP + /mCherry + neurons. A/P = anterior/posterior. M/L = medial/lateral; D/V = dorsal/ventral coordinates. (c) On the left: representative images of the viral delivery to the dHPC. (d) Quantification of hnRNPD levels in AAV9.GFP-, OE- or shRNA-transduced neurons. Mean ± SEM. (e) contextual threat conditioning of mice injected with AAV9.GFP-, OE- or shRNA viruses. On the left, training session; On the right, test session. OE = overexpression. Hab = habituation (N=6-7 mice per group); Mean ± SEM; F for panel d = 0.7307; F for panel e = 0.6853. (f-h) Same as in c-e, but for Pvalb + neurons (N=6-7 mice per group); Mean ± SEM; F for panel g = 0.7307; F for panel h = 1.097. (i-k) Same as in c-e, but for Sst + neurons (N=3-4 mice per group); Mean ± SEM; F for panel j = 0.0421; F for panel k = 1.412. P values are demonstrated to the fourth digit above bar graphs in each panel. One-Way ANOVA corrected with Tukey’s post-hoc test.

Article Snippet: Camk2a.Cre ERT2 /GADD34 flfl mice were injected with AAV9.CAG.FLEX.RPL10a::eGFP (Addgene).

Techniques: Expressing, shRNA, Injection, Over Expression

Journal: Molecular Medicine

Article Title: Wnt5a-mediated autophagy contributes to the epithelial-mesenchymal transition of human bronchial epithelial cells during asthma

doi: 10.1186/s10020-024-00862-3

Figure Lengend Snippet: Schematic illustration of the mechanisms of Wnt5a-induced EMT in HBECs. This hypothesis illustrates a molecular pathway in HBECs leading to airway remodeling, typically associated with classical autophagy pathway and EMT. In short, Wnt5a induced by HDM or IL-4 leads to the overactivation of autophagy in HBECs through the Ca 2+ /CaMKII signaling pathway, subsequently promoting bronchial epithelial EMT and triggering airway remodeling

Article Snippet: The antibodies used in the present research were as follows: rabbit anti-α-Tubulin antibody (1:2000, Servicebio, Wuhan, China); rabbit anti-α-SMA antibody (1:1000, Cell Signaling Technology, USA); rabbit anti-E-cadherin antibody (1:2000, Cell Signaling Technology); rabbit anti-Beclin1 antibody (1:1500, Cell Signaling Technology); rabbit anti-Collagen I antibody (1:1000, Cell Signaling Technology); rabbit anti-LC3 antibody (1:1500, Cell Signaling Technology); rabbit anti-Wnt5a antibody (1:1000, BOSTER, Wuhan, China); rabbit anti-CaMKII antibody (1:1000, BOSTER); rabbit anti-p-CaMKII antibody (1:1000, Cell Signaling Technology).

Techniques:

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet: ( A ) Pedigree of a consanguineous Jordanian family with two affected siblings with germline homozygous mutations in CAMK2A . The genotypes of all individuals were verified by Sanger sequencing. ( B ) Photographs of the two affected siblings with normal head circumferences. ( C ) EEG graph of patient II.I showing abnormal epileptiform transients (red boxes) ( D ) Homozygosity mapping delineates one candidate locus on chromosome 5. ( E ) CAMK2A exonic structure and CAMK2A protein domains. Patients II:1 and II:4 carry biallelic missense mutation p. H477Y located in CAMK2A association domain (AD). Nucleotide change c.1429 C > T refers to position on CAMK2A cDNA.

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Sequencing, Mutagenesis

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet: List of homozygous variants identified by Whole Exome Sequencing.

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Sequencing, Variant Assay

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet: ( A ) Sequence conservation of CAMK2A homologs. Histidine 477 (H477) is highlighted in red. ( B ) X-ray crystal structure of human CAMK2A AD tetradecamer (PDB: 5IG3). H477 (red) is located at the equatorial dimer interface. ( C ) Defective oligomerization of the p.H477Y mutant. 293 T cells were transiently transfected with FLAG tagged wild-type CAMK2A and CAMK2A H477Y . A third mutant, CAMK2A H477X which lacks part of the AD (a.a. 478–489) was used as positive control. ( D ) Defective self-association of the p.H477Y mutant. The indicated FLAG- and HA-tagged CAMK2A wild-type and mutant proteins were synthesized in vitro using rabbit reticulocyte lysate. FLAG-GFP was used a negative control. FLAG-tagged proteins were immunoprecipitated using anti-FLAG M2 agarose resin in the presence of 1% NP40. Co-immunoprecipitated proteins were analyzed by SDS-PAGE. *, IgG light chain. ^, IgG heavy chain. ( E ) p.H477Y mutation lowers expression of CAMK2A in cells. 293 T cells were transfected with reporter plasmids encoding GFP-tagged wild-type CAMK2A, CAMK2A H477Y and CAMK2A H477X mutants, followed by T2A peptide and mCherry. Representative confocal images show lower expression of mutant GFP- CAMK2A H477Y compared to wild-type. Scale bar represents 100 µm. ( F ). p.H477Y decreases CAMK2A stability via proteasomal degradation. 293 T cells were transfected as in ( E ) and treated with 10 µM MG132 or DMSO for 16 hr. 10 µg total cell lysate was used for SDS-PAGE and Western blot.

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Sequencing, Mutagenesis, Transfection, Positive Control, Synthesized, In Vitro, Negative Control, Immunoprecipitation, SDS Page, Expressing, Western Blot

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet: ( A ) Sequence conservation of CAMK2 paralogs. His477 (red) is invariant in all human CAMK2 paralogues with an association domain. ( B ) p.H477Y mutant is subject to proteasomal degradation. 293 T cells were transfected with plasmids encoding GFP-tagged wild-type CAMK2A, p.H477Y and p.H477X mutants. Cells were incubated with DMSO or 2.5 μM MG132 for 16 hr 1 day post transfection. GFP intensity in the cells is increased in the CAMK2A p.H477Y and p.H477X mutants after MG132 treatment. Scale bar represents 100 µm.

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Sequencing, Mutagenesis, Transfection, Incubation

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet: ( A ) Schematic drawing of C. elegans motor neuron, DA9 in the tail region. DA9 extends a dendrite (red) anteriorly and an axon (blue) that extends posteriorly crosses the midline of the animal and anteriorly in the dorsal nerve cord (DNC). It forms approximately 20 en passant synapses within a discrete stretch along the DNC (blue box). DA9 presynaptic vesicles were marked with RAB-3 (GFP::RAB-3). The asynaptic region (yellow box) is devoid of any synaptic vesicle accumulation. ( B ) The localization of mCherry::UNC-43 and mCherry::UNC-43(H466Y) in DA9 synapses. Note that UNC-43 accumulates at synaptic boutons while UNC-43(H466Y) is diffusely localized. Fluorescent intensity of mCherry::UNC-43 was measured at synaptic boutons and along the axonal shaft. Graph plots the ratio of fluorescence intensity at synaptic boutons compared to the axonal shaft of 30 synapses from three animals. Graph shows the mean and error bars show SEM, ***p-value 6.32e −19 , Student’s T-test. ( C ) Representative confocal images demonstrating presynaptic puncta size changes between WT and unc-43(e408) mutants. unc-43 mutants have smaller presynaptic puncta along the DNC. This defect is rescued by expression of either UNC-43 or CAMK2A in DA9 whilst the mutated UNC-43 H466Y and CAMK2A H477Y fail to rescue. ( D ) Quantification of average puncta intensity from WT and unc-43(e408) animals. Error bars represent SEM with number of synaptic puncta quantified n > 80, N.S. is not significant, ***p-value<0.001 (uninjected vs unc-43 p-value 5.0e 8 , uninjected vs unc-43 H466Y p-value 4.17, uninjected vs CAMK2A p-value 4.25e −12 , uninjected vs CAMK2A H477Y p-value 9.40), One-Way ANOVA with Bonferroni posthoc test. ( E ) Representative confocal images showing mislocalization of GFP::RAB-3 into the asynaptic region (yellow box) in unc-43 DA9 neuron. ( F ) Rescue of the unc-43(e408) phenotype by DA9 cell-specific expression of UNC-43 or CAMK2A. UNC-43 H466Y and CAMK2A H477Y fail to rescue the unc-43 phenotype. Graph shows the percentage of animals with the WT and unc-43 mutant phenotypes. ***p<0.001 ( unc-43 vs unc-43 H466Y p-value 2.13e −51 , CAMK2A vs CAMK2A H477Y p-value 3.77e −50 ), Fisher Exact test with n = 100 animals scored for each line. ( G ) Behavioral rescue by expressing wild-type UNC-43 or UNC-43 H466Y in unc-43(e408) mutants. The behavior was scored as either wild-type or unc-43. Two independent worm lines were analyzed for each condition. *** p-value 5.29e −41 , Fisher Exact test with n = 100 animals scored for each line.

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Fluorescence, Expressing, Mutagenesis

Journal: eLife

Article Title: A homozygous loss-of-function CAMK2A mutation causes growth delay, frequent seizures and severe intellectual disability

doi: 10.7554/eLife.32451

Figure Lengend Snippet:

Article Snippet: CAMK2A proteins were synthesized in vitro using TNT T7 Quick Coupled Transcription/Translation (Promega) with 1 μg of plasmids in 20 μl reaction volumes for 90 mins at 30°C.

Techniques: Recombinant, Plasmid Preparation, Clone Assay, Modification, Derivative Assay, Cell Culture, Knock-Out

Journal: Molecular Medicine Reports

Article Title: Effects of intrathecal bupivacaine on the NR2B/CaMKIIα/CREB signaling pathway in the rat lumbar spinal cord

doi: 10.3892/mmr.2018.8448

Figure Lengend Snippet: Intrathecal bupivacaine reduces the expression of p-CaMKIIα in rat spinal cord tissue. (A) Protein expression of CaMKIIα/p-CaMKIIα as measured by western blot analysis; (B) mRNA expression of CaMKIIα as measured by reverse transcription-quantitative polymerase chain reaction; (C) Protein expression of p-CaMKIIα as measured by immunohistochemistry (magnification, ×400). Data are expressed as the mean ± standard deviation of six rats in each group. **P<0.01 vs. the NS group. CaMKII, calcium/calmodulin-dependent protein kinase II; p, phosphorylated; Bup, bupivacaine; NS, normal saline; OD, optical density.

Article Snippet: Following incubation in 5% non-fat milk for 2 h at room temperature, membranes were incubated with primary antibodies against GAPDH (10494–1-AP; 1:2,000; ProteinTech Group, Inc., Chicago, IL, USA), NR2B (21920–1-AP; 1:1,000; ProteinTech Group, Inc.), CaMKIIα (13730–1-AP; 1:1,000; ProteinTech Group, Inc.), CREB (9197; 1:900; Cell Signaling Technology, Inc., Danvers, MA, USA), phosphorylated (p)-CaMKIIα (11278; 1:700; Signal-Aldrich; Merck KGaA, Darmstadt, Germany), and p-CREB (9198; 1:600; Cell Signaling Technology, Inc., USA) at 4°C overnight.

Techniques: Expressing, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction, Immunohistochemistry, Standard Deviation, Saline