rabbit polyclonal anti sec61β  (Proteintech)

Journal: EMBO Reports

Article Title: Sec61β maintains cytoplasmic proteostasis via ARIH1-mediated translational repression upon ER stress

doi: 10.1038/s44319-026-00690-y

Figure Lengend Snippet: ( A ) Gene Ontology (GO) analysis of Derlins-interacting proteins by thapsigargin (Tg) treatment. The bar graph shows the top 10 GO molecular function terms with a false discovery rate of <0.05 calculated from the DAVID online tool. P values were calculated using the modified Fisher’s exact test implemented in DAVID; 37 proteins identified as RNA binding in terms of molecular function are listed in Dataset . ( B ) Interactions of Derlins with Sec61β. HEK293 cells transfected with indicated plasmids and treated with or without 50 nM Tg and 200 nM MG132 for 16 h were immunoprecipitated (IPed) with an anti-Flag antibody and immunoblotted with indicated antibodies. ( C ) Endogenous interaction of Derlin-1 with Sec61β. Immunoprecipitation (IP) with anti-Sec61β antibody or control (Ctrl) IgG using Protein G Sepharose and immunoblotting (IB) with indicated antibodies in HepG2 cells treated with or without 200 nM Tg and/or 500 nM MG132 for 16 h. ( D – F ) IB of ERpQC substrates in HEK293 cells transfected with indicated siRNAs and plasmids and treated with or without 50 nM Tg and 200 nM MG132 for 16 h. All samples were immunoblotted with indicated antibodies. Black arrowhead, signal peptide-uncleaved NHK QQQ ( S NHK QQQ ); white arrowhead, signal peptide-cleaved NHK QQQ ( C NHK QQQ ). ( G ) IB of ERpQC substrate in wild-type (WT) or Derlin-1, -2 , and -3 triple knockout (TKO) HEK293 cells transfected with indicated siRNAs and plasmid for NHK QQQ and treated with or without 50 nM Tg and 200 nM MG132 for 16 h. All samples were immunoblotted with indicated antibodies. Expression levels of S NHK QQQ were calculated and shown as the percentage of S NHK QQQ out of the total amount of NHK QQQ ( S NHK QQQ and C NHK QQQ ). Black arrowhead, S NHK QQQ ; white arrowhead, C NHK QQQ . .

Article Snippet: Rabbit polyclonal anti-Sec61β (IB: 1:500) , Proteintech , Cat. #15087-1-AP; RRID: AB_2186411.

Techniques: Modification, RNA Binding Assay, Transfection, Immunoprecipitation, Control, Western Blot, Triple Knockout, Plasmid Preparation, Expressing

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet: (A) HeLa Fcγ cells were infected with L.p. for the indicated times, stained with anti-Rtn4 (sER marker), Sec61β (rER marker) antibodies, and Hoechst ( L.p .) for immunofluorescence analysis. Results represent three independent experiments (100 vacuoles were scored in each). Error bars represent mean ± SD. Bar, 5 μm. (B) HeLa Fcγ cells stably expressing GFP-Rtn4 and RFP-Sec61β were infected with Halo-tagged WT L.p . for 3 h for FLIP. The top panel represents GFP-Rtn4 before photobleaching, and the bottom represents GFP-Rtn4 after photobleaching for 80 s. Bar, 10 μm. (C) The graph represents the relative fluorescence intensity of GFP-Rtn4 (green lines) and RFP-Sec61β (red lines) in the ER and around LCV. Error bars represent mean ± SD. The purple and red boxes represent the area around LCV and bleach areas, respectively. (D) HeLa Fcγ cells stably expressing GFP-Rtn4 and RFP-Sec61β were infected with WT L.p . for 6 h. The top panel represents RFP-Sec61β before photobleaching, and the bottom represents RFP-Sec61β after photobleaching. The purple and red boxes represent the area around the LCV and bleach areas, respectively. Bar, 10 μm. (E) The graph represents the relative fluorescence intensity of GFP-Rtn4 (green lines) and RFP-Sec61β (red lines) in the ER and around LCV. (F) HeLa Fcγ cells stably expressing GFP-Rtn4 and RFP-Sec61β were infected for 6 h, and RFP-Sec61β was photobleached for FRAP. The green box represents the total area of bleach. The red and blue boxes represent the bleached area around the LCV and the ER, respectively. White arrows represent the area around LCV. Bar, 10 μm. (G) The graph represents the relative fluorescence intensity of GFP-Rtn4 (green lines) and RFP-Sec61β (red lines) in the ER and around LCV. Error bars represent mean ± SD.

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Infection, Staining, Marker, Immunofluorescence, Stable Transfection, Expressing, Fluorescence

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet: (A) HeLa Fcγ cells were subjected to subcellular fractionation, and immunoblot analysis was done against the indicated proteins. (B) Sucrose gradient fractionation was done to obtain the sER and rER fractions and immunoblotted against indicated antibodies. (C) HeLa Fcγ cells expressing mRFP-Rab4 or mRFP-Rab10 were infected with L.p. for the indicated times, stained with L.p . for immunofluorescence analysis, and the vacuoles positive for mRFP-Rab4 or -Rab10 were quantified. Bar, 5 μm. The purple box represents the magnified image. Results represent three independent experiments (100 vacuoles were scored in each). Error bars represent mean ± SD. (D) HEK293 Fcγ cells were transfected with either mock, Rab4, or Rab10 siRNA for 72 h, and cell extracts were immunoblotted against the indicated antibodies. (E and F) HeLa Fcγ cells were silenced for Rab4 or Rab10 for 72 h, infected with L.p. for 4 and 8 h, and the vacuoles positive for Rtn4 or Sec61β were counted. Results represent three independent experiments, and statistical significance was measured using one-way ANOVA (100 vacuoles were scored in each experiment). Bars represent mean ± SD. (G) HeLa Fcγ cells were silenced with Rab4 and Rab10 siRNA, and the ER morphology was observed by immunofluorescence analysis. Bars, 5 μm. Arrows indicate disruption of the sER tubular network. Arrowheads indicate diffusion of rER to the cell periphery. (H) Triplicate experiments (30 cells for each condition) from 3G were quantitated and are shown as a histogram. Bars represent mean ± SD.

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Fractionation, Western Blot, Expressing, Infection, Staining, Immunofluorescence, Transfection, Disruption, Diffusion-based Assay

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet: (A) HeLa Fcγ cells were infected with L.p . for the indicated time, fixed, and stained with anti-BAP31 antibody and Hoechst 33342. White boxes indicate the LCV. (B) HeLa Fcγ cells were transfected with either mock or BAP31 siRNA for 72 h and immunoblotted against the indicated antibodies. (C and D) Mock-treated or BAP31-silenced HeLa Fcγ cells were infected with L.p . for the indicated time, fixed, and stained with anti-Rtn4 or -Sec61β antibodies and DAPI, and the vacuoles were quantified. Results are representative of three independent experiments (100 vacuoles were scored in each experiment). Bars represent mean ± SD. (E) HeLa Fcγ cells were silenced with either mock or BAP31 for 72 h, followed by infection with L.p. for 8 h. Post infection, cells were fixed and stained with anti-Rtn4 antibody and Hoechst for immunofluorescence analysis. Bar, 5 μm. (F) HeLa Fcγ cells were silenced with either mock or BAP31 for 72 h, followed by infection with L.p. for 12 h at an MOI of 2. Post infection, cells were fixed and stained with anti- Legionella antiserum for detection of extracellular bacteria, then permeabilized and further stained with DAPI for detection of intracellular bacteria. Bar, 5 μm. The graph shows the percentage of vacuoles containing a single bacterium, 1–10 bacteria, 11–30 bacteria, and ≥31 bacteria in a single vacuole. Data are representative of three independent experiments (30 vacuoles were scored in each experiment). Results are shown as the mean ± SD. *** p < 0.001.

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Infection, Staining, Transfection, Immunofluorescence, Bacteria

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet: (A) Schematic representation of the chromosomal organization of genomic islands of L.p . (B) HeLa Fcγ cells were infected with the indicated genomic island deletion mutant L.p. strains for 4 h and processed for immunofluorescence against BAP31 antibody. The histogram represents the percentage of vacuoles positive for BAP31 post infection with different L.p. strains. (C) HeLa Fcγ cells were transfected with 3xFLAG-Lpg1152 plasmid and processed for immunofluorescence microscopy using FLAG (green) and BAP31 (red) antibodies. Nuclei were stained with DAPI. Bar, 10μm. (D and E) HEK293 Fcγ cells were transfected with the indicated plasmids. Cell extracts were subjected to immunoprecipitation assay post transfection with anti-FLAG antibody and immunoblotted against the indicated antibodies. (F) The docked model complex of Lpg1152 (deep salmon) and BAP31 (green). (G) The electrostatic potential surface of Lpg1152 is shown bound with BAP31 (green). (H) Interacting residues of Lpg1152 (orange, ball-stick) and BAP31 (lime, ball-stick) at the protein-protein interface. (I) HeLa Fcγ cells were infected with the indicated L.p. strains. Histograms represent the infection efficiency, i.e., the percentage of infected cells at MOI 10 1.5 h after infection ( n = 200 per triplicate experiment). (J) HeLa Fcγ cells were infected with the indicated strains of L.p. for the indicated time points. Following infection, LCV positive for BAP31 was scored using immunofluorescence analysis. (K) HeLa Fcγ cells were infected with the indicated strain, and the Rtn4- and Sec61β-positive vacuoles were scored using immunofluorescence. Graphs represent the percentage recruitment of Rtn4 and Sec61β at the indicated time points post infection. (L) HeLa Fcγ cells were infected with the indicated L.p. strains, and the number of L.p. per LCV was counted 8 h after infection ( n = 30 per triplicate experiment). (M) U937 cells were infected with the indicated L.p. strains for 1 h, and colony-forming units (CFU) were counted at the indicated time after infection. Data represent three independent experiments, and statistical significance was carried out with a Student’s t test. Bar represents mean ± SEM.

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Infection, Mutagenesis, Immunofluorescence, Transfection, Plasmid Preparation, Microscopy, Staining, Immunoprecipitation

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet: (A) HeLa Fcγ cells were silenced with mock, BAP31, or BAP29 siRNA for 72 h. Post silencing, cells were fixed and stained with anti-Rtn4 and Sec61β antibodies and were subjected to immunofluorescence imaging. Bar, 5 μm. White arrows indicate Sec61β localization to the cell periphery. (B) HeLa Fcγ cells were silenced with either mock or Stx-18 siRNA for 72 h. Post silencing, cells were fixed and stained with anti-Rtn4 and anti-Sec61β antibodies. Bar, 5 μm. (C and D) HeLa Fcγ cells were silenced for Stx18 alone or a combination of Stx18/BAP31 or Stx18/BAP29 siRNA. Following single or double silencing, cell extracts were prepared and immunoblotted against the indicated antibodies. (E) HeLa Fcγ cells were silenced for Stx18 alone, Stx18/BAP31, or Stx18/BAP29; following single or double silencing, cells were fixed and stained with an anti-Rtn4 antibody for immunofluorescence analysis. Bar, 5 μm.

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Staining, Immunofluorescence, Imaging

Journal: Cell reports

Article Title: Legionella uses host Rab GTPases and BAP31 to create a unique ER niche

doi: 10.1016/j.celrep.2024.115053

Figure Lengend Snippet:

Article Snippet: Anti-Sec61β rabbit polyclonal antibody , Proteintech , Cat# 14846-1-AP.

Techniques: Virus, Recombinant, Transfection, In Situ, Fluorescence, Stable Transfection, Expressing, Software

Journal: eLife

Article Title: Single-cell transcriptomics of vomeronasal neuroepithelium reveals a differential endoplasmic reticulum environment amongst neuronal subtypes

doi: 10.7554/eLife.98250

Figure Lengend Snippet: Pseudocolored immunofluorescence images of VNO coronal sections labeled with antibodies against KDEL (anti-SEKDEL), a common ER retention signal ( A ). The section is co-labeled with anti-Gnao1 to mark basal zone neurons ( B ) and anti-OMP to mark all mature neurons ( C ). Signal intensity of KDEL, Gnao1, and OMP channels are quantified from ROIs along the apical-basal axis as shown in example ( D ). Signal intensity measured from multiple sections (n>20 for each antibody) from three biological replicates was normalized and the trendline was fitted to show the Gnao1 neuron-enriched localization of anti-KDEL ( E ). Points on the plot show normalized intensity values color-coded for each antibody on which the trendline was fitted. Similar immuno-labeling and quantification of ER chaperone Hspa5 (BiP) ( F ), ER membrane translocon subunit Sec61β ( G ), and ER membrane protein Atlastin1 ( H ) indicate their enrichment in Gnao1 neurons compared to Gnai2 neurons. Distribution of additional ER chaperone and membrane proteins is shown in along with combined source data in . Scale bar: 50 µm. Figure 7—source data 1. Fluorescence intensity values of ROIs along an apical-basal axis from immunolabeled images of Vomeronasal organ (VNO) neuroepithelium.

Article Snippet: Antibody , anti-Sec61β (Rabbit polyclonal) , Invitrogen , Cat#PA3-015 RRID: AB_2239072 , Dilution –1:400.

Techniques: Immunofluorescence, Labeling, Immunolabeling, Membrane, Fluorescence

Journal: eLife

Article Title: Single-cell transcriptomics of vomeronasal neuroepithelium reveals a differential endoplasmic reticulum environment amongst neuronal subtypes

doi: 10.7554/eLife.98250

Figure Lengend Snippet:

Article Snippet: Antibody , anti-Sec61β (Rabbit polyclonal) , Invitrogen , Cat#PA3-015 RRID: AB_2239072 , Dilution –1:400.

Techniques: Purification, Labeling, Recombinant, Blocking Assay, Electron Microscopy, Marker, Software

Journal: Molecular Cell

Article Title: Structure of the Inhibited State of the Sec Translocon

doi: 10.1016/j.molcel.2020.06.013

Figure Lengend Snippet:

Article Snippet: Primary antibodies used were: mouse monoclonal anti-Sec61α; rabbit polyclonal anti-Sec61β (Kind gift from the Dobberstein lab) ( ); rabbit polyclonal anti-ribosomal protein S6 and mouse monoclonal anti-ribophorin 2 (Santa Cruz Biotechnology, sc-166421).

Techniques: Recombinant, Western Blot, Sequencing, Software, Dispersion, Membrane, Modification, Protease Inhibitor