Journal: The Journal of Neuroscience

Article Title: Neuregulin1 Nuclear Signaling Influences Adult Neurogenesis and Regulates a Schizophrenia Susceptibility Gene Network within the Mouse Dentate Gyrus

doi: 10.1523/JNEUROSCI.0063-24.2024

Figure Lengend Snippet: Key resources table

Article Snippet: Recombinant mouse ErbB4/Her4 Fc chimera protein, CF , R&D Systems , Catalog #4387-ER-050.

Techniques: Affinity Purification, Virus, Recombinant, Protease Inhibitor, Sequencing, RNAscope, Variant Assay, Plasmid Preparation, Software

Journal: The Journal of Neuroscience

Article Title: Neuregulin1 Nuclear Signaling Influences Adult Neurogenesis and Regulates a Schizophrenia Susceptibility Gene Network within the Mouse Dentate Gyrus

doi: 10.1523/JNEUROSCI.0063-24.2024

Figure Lengend Snippet: The V 321 L substitution decreases nuclear back signaling. A , Immunoblot of triplicate nuclear fractions isolated from pooled cortical and hippocampal lysates. Nrg1 ICD was detected using Santa Cruz Biotechnology antibody sc-348. Histone H3 served as a nuclear loading control and Na+/K+ ATPase as a marker for the membrane fraction ( N = 3 mice/genotype). B , Immunoblot of two replicates of membrane fractions isolated from pooled cortical and hippocampal lysates. NRG1 ICD was detected using Santa Cruz Biotechnology antibody sc-348. Na+/K+ ATPase served as a marker for the membrane fraction; note the lack of the nuclear marker Histone H3 or cytoplasmic marker CyclophilinA indicating clean membrane preps. FL Nrg1 is indicated with a yellow arrowhead as “Nrg1-FL,” and the membrane-bound C-terminal fragment not cleaved by γ-secretase is indicated as “Nrg1 TM-CTF.” A positive control consisting of total lysate from N2A cells transfected with a Type III Nrg1 plasmid is shown in the lane labeled “C.” ( N = 2 mice/genotype.) C , Left, Hippocampal neurons from WT (dark blue) and V 321 L (light blue) neonatal pups (P4) were cultured for 17 d in vitro and were stimulated with either vehicle (Veh), 20 nM sERBB4 (sB4), or 20 nM sErbB4 after a 24 h pretreatment with 20 µM of the γ-secretase inhibitor DAPT (DAPT). Neurons were fixed and stained using an antibody directed against the Nrg1 ICD and counterstained with DAPI. Scale bar, 10 µm. Right, Quantification of nuclear clusters of Nrg1-ICD. Neurons from WT mice show increased nuclear ICD clusters in response to sB4 stimulation, which is counteracted by pretreatment with DAPT (DAPT). Neurons from V 321 L mice do not respond to sB4 stimulation ( N = 6–13 neurons, 3 platings/mouse, 3 mice/genotype; one-way ANOVA p values corrected for multiple comparisons using Tukey’s post hoc test; WT Veh vs WT sB4, p < 0.0001 (****); WT sB4 vs WT sB4 + DAPT, **** p < 0.0001; WT sB4 vs V 321 L Veh, **** p < 0.0001; WT sB4 vs V 321 L B4, **** p < 0.0001). All other comparisons are statistically not statistically significant. D , Cortical neurons from embryonic WT (dark blue) and V 321 L mice (light blue; E18.5) were cultured for DIV3 and were stimulated with soluble ErbB4 (sB4), PI3K inhibitor WM, γ-secretase inhibitor L-685,458 (L6), WM + B4, or L6 + B4. Neurons that underwent no drug treatments/sB4 stimulation are indicated as the control group (C). Neurons were fixed and axonal length was quantified. (Two-way ANOVA with Tukey’s post hoc correction, WT C vs WT B4, **** p = 0.0002; WT L6 vs WT L6 + B4, ** p = 0.0047; V 321 L C vs V 321 L B4, *** p = 0.001; V 321 L WM vs V 321 L WM + B4, p = 0.1; V 321 L L6 vs V 321 L L6 + B4, * p = 0.03.) N = 20–37 neurons per genotype per condition. ns, not significant. E , Treatment and conditions same as in D . Quantification is for dendritic length. (two-way ANOVA w/ Tukey’s post hoc correction: WT C vs WT B4, ** p = 0.002; WT WM vs WT WM + B4, **** p < 0.0001). N = 20–37 neurons per genotype per condition. ns, not significant.

Article Snippet: Recombinant mouse ErbB4/Her4 Fc chimera protein, CF , R&D Systems , Catalog #4387-ER-050.

Techniques: Western Blot, Isolation, Control, Marker, Membrane, Positive Control, Transfection, Plasmid Preparation, Labeling, Cell Culture, In Vitro, Staining

Journal: Cancer control : journal of the Moffitt Cancer Center

Article Title: ERBB4 Expression in Ovarian Serous Carcinoma Resistant to Platinum-Based Therapy

doi: 10.1177/107327481702400115

Figure Lengend Snippet: Mean level of ERBB4 expression in normal controls, tumor tissue, and ovarian cancer cell lines. Analysis of variance was statistically significant (P < .001). The mean scores of ERBB4 are significantly different for tumors vs controls vs cell lines, with cell lines greater than tumors and greater than controls.

Article Snippet: A mouse monoclonal antibody that reacts to ERBB4, NB100–2662 (Novus, Littleton, CO), was used at a 1:25 concentration in Antibody Diluent (Dako, Carpenteria, CA) and incubated for 60 minutes.

Techniques: Expressing

Journal: Cancer control : journal of the Moffitt Cancer Center

Article Title: ERBB4 Expression in Ovarian Serous Carcinoma Resistant to Platinum-Based Therapy

doi: 10.1177/107327481702400115

Figure Lengend Snippet: ERBB4 expression by immunohistochemistry. Shown are (A) rare membranous, (B) nuclear staining, (C–G) increasing H scores from less than 50 to 300 for particular fields, and (H) ERBB4 staining in cells lines.

Article Snippet: A mouse monoclonal antibody that reacts to ERBB4, NB100–2662 (Novus, Littleton, CO), was used at a 1:25 concentration in Antibody Diluent (Dako, Carpenteria, CA) and incubated for 60 minutes.

Techniques: Expressing, Immunohistochemistry, Staining

Journal: Cancer control : journal of the Moffitt Cancer Center

Article Title: ERBB4 Expression in Ovarian Serous Carcinoma Resistant to Platinum-Based Therapy

doi: 10.1177/107327481702400115

Figure Lengend Snippet: Using an unpaired t test with Welch correction, ERBB4 expression was higher patients with an IR than CR to platinum-based therapy (P = .033).

Article Snippet: A mouse monoclonal antibody that reacts to ERBB4, NB100–2662 (Novus, Littleton, CO), was used at a 1:25 concentration in Antibody Diluent (Dako, Carpenteria, CA) and incubated for 60 minutes.

Techniques: Expressing

Journal: Cancer control : journal of the Moffitt Cancer Center

Article Title: ERBB4 Expression in Ovarian Serous Carcinoma Resistant to Platinum-Based Therapy

doi: 10.1177/107327481702400115

Figure Lengend Snippet: Multivariate Survival Analysis Using a Cox Proportional Hazard Model

Article Snippet: A mouse monoclonal antibody that reacts to ERBB4, NB100–2662 (Novus, Littleton, CO), was used at a 1:25 concentration in Antibody Diluent (Dako, Carpenteria, CA) and incubated for 60 minutes.

Techniques:

Journal: Cancer control : journal of the Moffitt Cancer Center

Article Title: ERBB4 Expression in Ovarian Serous Carcinoma Resistant to Platinum-Based Therapy

doi: 10.1177/107327481702400115

Figure Lengend Snippet: Log-rank test results illustrating ERBB4 expression and OS. Probability values were as follows: all 3 levels, P = .020; low vs intermediate, P = .080; low vs high, P = .004; and intermediate vs high, P = .201.

Article Snippet: A mouse monoclonal antibody that reacts to ERBB4, NB100–2662 (Novus, Littleton, CO), was used at a 1:25 concentration in Antibody Diluent (Dako, Carpenteria, CA) and incubated for 60 minutes.

Techniques: Expressing

Journal: Diabetes

Article Title: Mutations of NRG4 Contribute to the Pathogenesis of Nonalcoholic Fatty Liver Disease and Related Metabolic Disorders.

doi: 10.2337/db21-0064

Figure Lengend Snippet: Figure 4—Enhancement of E47Q Nrg4 on the attenuation of de novo lipogenesis in hepatocytes and the loss of function of R44H Nrg4. A: TAG levels in hepatocytes transduced with ErbB4 adenovirus overnight and treated for 24 h with palmitate (PA) (0.6 mmol/L) and WT, R44H, and E47Q Nrg4 (20 mg/mL) simultaneously. B: Quantitative PCR analysis of gene expression in ErbB4 adenovirus transduced pri- mary hepatocytes treated with the combination of 2:1 oleate (OA) (0.6 mmol/L) and PA (0.3 mmol/L) in the presence of Nrg4 for 24 h. Con- trol (Con): hepatocytes transduced with ErbB4 adenovirus. C: Immunoblots of hepatocytes transduced with ErbB4 adenovirus and treated with WT, R44H, and E47Q Nrg4 for 20 min. D: Immunoblots of total hepatocyte lysates and nuclear extracts. *P < 0.05, **P < 0.01, ***P < 0.001 compared with WT group by ANOVA and SNK multiple pairwise comparison test; #P < 0.05, ##P < 0.01, ###P < 0.001 compared with GFP group by ANOVA and SNK multiple pairwise comparison test.

Article Snippet: ErbB4 protein from R&D Systems (4387-ER) was immobilized on a chip.

Techniques: Transduction, Real-time Polymerase Chain Reaction, Gene Expression, Western Blot, Comparison

Journal: Diabetes

Article Title: Mutations of NRG4 Contribute to the Pathogenesis of Nonalcoholic Fatty Liver Disease and Related Metabolic Disorders.

doi: 10.2337/db21-0064

Figure Lengend Snippet: Figure 5—Examination of the interaction between any of the three versions of Nrg4 (WT, R44H, E47Q) and ErbB4. A: SPR assay with Bia- core diagram and saturation curve of WT Nrg4 binding to the ErbB4 with an affinity of 2.20 mmol/L. The purified Nrg4 proteins were tested for binding with gradient concentrations of 0.195, 0.39, 0.78, 1.5625, 3.125, and 6.25 mmol/L in single-cycle mode. The binding profiles are shown with time (s) on the x-axis and response units (RU) on the y-axis. As the flowing protein Nrg4 concentration increases (x-axis), the chip RU (y-axis) continues to increase, indicating the interaction between two kinds of protein (right). B: Biacore diagram showing no binding between R44H Nrg4 and ErbB4. When gradient concentrations of the R44H Nrg4 solution flowed through the chip, there was no increase in RU, indicating that there was no interaction between the two types of protein. C: Biacore diagram and saturation curve of E47Q mutant of Nrg4 binding to ErbB4. The E47Q mutant can keep binding to the ErbB4 with an affinity of 1.66 mmol/L, which is a slightly higher affinity than that of WT Nrg4. D: Fluorescence micrographs of primary hepatocytes transduced with GFP-ErbB4 adenovirus follow- ing three kinds of Nrg4 stimulation. E–I: Binding capacity as measured by flow cytometry between GFP-ErbB4 after 36 h of transfection and incubation with Nrg4 (500 mg/mL for 1 h). Dot plot obtained from flow cytometry analysis indicates the four different cell populations. The binding capacity was measured by the proportion of ErbB41Nrg41 hepatocytes (top right) in ErbB41 hepatocytes (top right and bot- tom right). Ctl, control.

Article Snippet: ErbB4 protein from R&D Systems (4387-ER) was immobilized on a chip.

Techniques: SPR Assay, Binding Assay, Concentration Assay, Mutagenesis, Fluorescence, Transduction, Cytometry, Transfection, Incubation, Control

Journal: Cell stem cell

Article Title: Human pallial MGE-type GABAergic interneuron cell therapy for chronic focal epilepsy

doi: 10.1016/j.stem.2023.08.013

Figure Lengend Snippet: Key resources table

Article Snippet: Single cell suspension was incubated with a biotinylated primary antibody against human ERBB4 (BAF1131 R&D Systems), followed by incubation with Anti-Biotin MicroBeads (130-105-637, Miltenyi Biotec), and magnetic sorting was performed on the CliniMACS according to manufacturer instructions (Miltenyi Biotec).

Techniques: Control, Plasmid Preparation, Recombinant, Software

Journal: Cell stem cell

Article Title: Human pallial MGE-type GABAergic interneuron cell therapy for chronic focal epilepsy

doi: 10.1016/j.stem.2023.08.013

Figure Lengend Snippet: (A,B) Simplified schematics of neuronal subtypes derived from the MGE progenitor domain (FOXG1+ DLX1/2+ NKX2-1+), including GABAergic pallial interneurons (pINs) that migrate to the cortex and hippocampus (HC), as well as multiple lineages that remain in the subpallium. The latter include striatal GABAergic and cholinergic INs, cholinergic projection neurons (PN) of the basal telencephalon, and GABAergic PNs of the globus pallidus. Listed genes expressed in the immature neurons play critical roles in the specification of the indicated neuronal subtypes. (C,D) Representative ICC images of unsorted and sorted lot 1 after cryopreservation and thaw show expression of MGE pIN markers (LHX6, MAFB, MAF, ERBB4), and cholinergic neurons (ISL1). (E) Protein quantification across independent lots (n=10 to 18). (F,G) Quantification of GABA (F) and Acetylcholine (G) neurotransmitters in the culture supernatant from independent pIN lots (n=8), undifferentiated hESCs (n=1) and spinal motor neurons (n=1). (H) Cell viability post-thaw from independent pIN lots (n=16 unsorted/sorted pairs). In E-H, each dot is an average of technical replicates (2-3) from independently manufactured lots. All data are expressed as a mean ± SEM.

Article Snippet: Single cell suspension was incubated with a biotinylated primary antibody against human ERBB4 (BAF1131 R&D Systems), followed by incubation with Anti-Biotin MicroBeads (130-105-637, Miltenyi Biotec), and magnetic sorting was performed on the CliniMACS according to manufacturer instructions (Miltenyi Biotec).

Techniques: Derivative Assay, Expressing

Journal: Cell stem cell

Article Title: Human pallial MGE-type GABAergic interneuron cell therapy for chronic focal epilepsy

doi: 10.1016/j.stem.2023.08.013

Figure Lengend Snippet: Key resources table

Article Snippet: Single cell suspension was incubated with a biotinylated primary antibody against human ERBB4 (BAF1131 R&D Systems), followed by incubation with Anti-Biotin MicroBeads (130-105-637, Miltenyi Biotec), and magnetic sorting was performed on the CliniMACS according to manufacturer instructions (Miltenyi Biotec).

Techniques: Control, Plasmid Preparation, Recombinant, Software