EMBO/sd-character-level-ner huggingface dataset:
ΔPrPs are degraded by both lysosomal and proteasomal degradation systems.Figure 5A. Degradation of ΔPrPs by lysosomal and proteasomal systems. (Left) N2a cells transiently transfected with Δ159 or Δ159-175 were treated for 7 hours with inhibitors either for lysosomal degradation (bafilomycin A1, Baf, 120 nM), autophagy (3-methyladenine, 3MA, 10 mM), or the proteasome (MG132, 10 µM) and cell lysates analyzed in immunoblot for expression of Δ159 and Δ159-175 (mAb 3F4). Square bracket and bracket denote diglycoforms and nonglycoforms of ΔPrPs, respectively. An increment in diglycoforms is observed in Baf- and 3MA-treated cells, whereas in MG132-treated cells also the non-glycoforms are increased. (Right) A graph showing quantification of all PrP bands by densitometric analysis. Data from 3 independent (one in duplicate) experiments for Δ159 and 2 independent (one in duplicate) for Δ159-175 were statistically analyzed for mean and standard deviation (error bars). B. ΔPrPs can be localized in LAMP1-immunopositive vesicles. Immunofluorescence analysis shows distribution of ΔPrP (mAb 3F4) and LAMP1 in cells with or without 6M GdnHCl antigen-retrieval treatment. N2a cells transfected with ΔPrP159 were fixed, permeabilized and incubated without (upper panel) or with 6M GdnHCl (lower panel). With GndHCl treatment ΔPrP is diffusely distributed (arrowheads) and co-localizes poorly with LAMP1. After GdnHCl treatment (lower panel), bright 3F4-immunopositive puncta (arrows) are observed which partly co-localize with LAMP1-positive structures (merge). Scale bars, 10 µm.
LNS2SS2- and Nxph1-LNS2SS2A+ complexes.Figure 1Schematic showing the domain structure of Nrxn1α and Nxph1 with approximate positions of alternative splice-sites indicated by arrows. Colors of LNS2 and Nxph1 correspond to those in the structures in B. Crystal structures of Nxph1 in complex with Nrxn1 LNS2SS2- and LNS2SS2A+. β-strands are numbered, and N- and C-termini are indicated. SS2A insert is shown in magenta. Disordered regions are shown as dashed lines, and disulfide bonds are shown as red sticks.
Figure 1A. Flow cytometry of cells isolated from heart tissue when scaled against TMRM staining and also FSC and SSC parameters of flow cytometry. B. Cardiac troponin I (cTnI) and α-Sarcomeric actinin (α-SA) staining of the TMRM++ population (CMs) and Vimentin staining of the TMRM+ population (NCs). The scale bar represents 100 µm. C. Schematic showing early reprogramming processes of CMs and NCs isolated from triple transgenic mice. D. EGFP expression in iPSC-like colonies derived from CMs or NCs isolated from triple transgenic mice. E. EGFP expression combined with immunofluorescence staining of SSEA-1 and DAPI of iPSC-like colonies derived from CMs isolated from triple transgenic mice. F. Morphological changes of isolated CMs and NCs at different time points before AP positive colony formation at day 6. The scale bar represents 100 µm.
a) Interaction between LC3 and wild-type Dvl2 or its point mutants.figf4(a) Interaction between LC3 and wild-type Dvl2 or its point mutants. (b) Ubiquitylation of wild-type Dvl2 and its variants. Transfected HEK-293T cells were treated with BFA1 (0.2 μM) for 6 h. Ubiquitylated proteins were precipitated with Ni-nitrilotriacetate (Ni-NTA) beads, followed by anti-HA immunoblotting. Ub, ubiquitin. (c) Degradation kinetics of wild-type Dvl2 and its variants under starvation conditions. Lower panel: quantification of the immunoblotting results. (d) Starvation promotes co-localization of Dvl, ubiquitin and p62. Scale bar, 10 μm. (e) Starvation induces the formation of high-molecular-mass Dvl2 aggregates. (f) p62 knockdown alters Dvl localization. Scale bar, 10 μm. Uncropped images of blots are shown in Supplementary Information, Fig. S8.
Reduced PINK1 levels influence cell metabolism and population growth.Figure 44A) Oxygen consumption rate (OCR) was measured in nt and PINK1 knockdown (kd) SH-SY5Y cells without (nt) or with PINK1 knockdown (kd) cultivated in RPMI medium with 5% FCS. Although oxygen consumption appeared to be slightly impaired in cells with PINK1 kd, no significant changes were observed; n = 1 (quadruplicates). 4B) Energy charge (EC) of nt and PINK1 knockdown (kd) SH-SY5Y cells grown in RPMI+5% FCS was determined as detailed in Material Methods. PINK1 knockdown resulted in a lower EC compared to nt cells; n = 5, p0.05. 4C) nt and PINK1 knockdown (kd) SH-SY5Y cells were grown for 2 months in RPMI+5% FCS and their population doublings were determined. Starting from day 12 cells with PINK1 kd exhibited impaired cell population growth; n = 1 (triplicates); * p<0.05 (day 12) - p<1×10−6 (day 61). 4D) SH-SY5Y cells were grown in RPMI+5% FCS and the amount of nt and PINK1 knockdown (kd) cells in S-phase was determined by BrdU incorporation. No differences in cell proliferation were visible between cells without (nt) and with PINK1 kd; n = 3.
ATG16L1 Is Found in Recycling Endosomes.Figure 2(A) HeLa cells transfected with GFP-ATG16L1 for 24 hr were incubated for 15 min with 2.5 μg/ml with HRP-cholera toxin subunit B at 4°C and then for 10 min at 37°C. Cells were then fixed and treated for immunogold labeling on cryosections. ATG16L1 was detected with anti-ATG16L1 antibody (15 nm), and cholera toxin was detected using anti-HRP antibody. Scale bar, 150 nm. (B) HeLa cells were transiently transfected with GFP-ATG16L1 for 20 hr. Cells in basal conditions were fixed and labeled with anti-RAB11 antibody. (C) HeLa cells transfected with mStrawberry-ATG16L1 (20 hr) and starved (4 hr) were processed for immunogold labeling on cryosections. Double labeling was performed with rabbit anti-ATG16L1 (10 nm gold) and rabbit anti-RAB11 (15 nm gold). Scale bar, 150 nm. Schematic diagram shows membrane outlines from EM. Green lines indicate structures positive for both ATG16L1 and RAB11, whereas red lines are only positive for RAB11. Histogram shows the percentage by EM of ATG16L1 tubulovesicular structures that carry RAB11. 281 tubulovesiclular structures from different experiments were analyzed. Error bars, SD. (D) HeLa cells were transfected with EGFP-ATG16L1 and mCherry-RAB11, and 5 min movies were recorded. Representative images from movies are shown. (E) HeLa cells were loaded with mouse anti-transferrin receptor antibody (5E9C11) for 30 min and fixed for immunogold labeling for cryosections. The sample was labeled with rabbit anti-mouse antibody to recognize transferrin receptor (5 nm), anti-ATG16L1 (10 nm), and anti-mATG9 (15 nm). Scale bar, 150 nm. (F) HeLa cells were transfected with pmStrawberry-ATG16L1 and ATG9L1-pEGFP, and 5 min movies were recorded. Representative images from the movies are shown. (G) HeLa cells were transfected with pmStrawberry-ATG16L1 and pEGFP-LC3, starved for 2 hr, fixed, and labeled for mATG9.
Figure 6A) Hierarchical clustering of motif co-association enrichments in pDHSs. Zscores represent enrichment of observed versus background co-associations computed in 1,000 randomly selected, chromatin-accessible regions. B) Log2 mRNA expression levels of Runx1, Ets1 and B2m in untreated and PMA/I treated TN, TM, and TB from the microarray analysis.
Figure 8A. Representative western blot demonstrating ABCB8 overexpression in H9c2 cells. EV = empty vector. N=3 independent samples for each group. B. Complex I ROS production in mitochondria with ABCB8 downregulation with or without rotenone. ANOVA followed by post-hoc Tukey test was performed. N= 6 independent samples for each group. C. Complex III ROS production in mitochondria with ABCB8 downregulation with or without antimycin A. ANOVA followed by post-hoc Tukey test was performed. N= 6 independent samples for each group. D. Complex I ROS production in mitochondria with ABCB8 overexpression with or without rotenone. EV= empty vector. ANOVA followed by post-hoc Tukey test was performed. N=9 independent samples for EV-baseline and N=8 independent samples for the other groups. E. Complex IIIROS production in mitochondria with ABCB8 overexpression with or without antimycin A. EV= empty vector. ANOVA followed by post-hoc Tukey test was performed. N=9 independent samples for EV-baseline and N=8 independent samples for the other groups. F. Mitochondrial ROS in H9c2 cells with various treatments with or without oxidative stress. N=6 independent samples for each group. ANOVA followed by post-hoc Tukey test was performed. * P=2E-8 PBS-PBS vs. PBS-H2O2. * P=2E-8 DFO- H2O2 vs. BPD-H2O2. G. Mitochondrial membrane potential as measured by TMRE intensity in H9c2 cells treated with indicated iron chelator with or without oxidative stress. N= 6 independent samples for each group. ANOVA followed by post-hoc Tukey test was performed. * P=1E-8 PBS-PBS vs. PBS-H2O2. * P=1.8E-8 DFO-PBS vs. DFO-H2O2. H. The levels of lipid peroxidation products in ABCB8 KO mice with or without chelator treatment. N=4 mice for WT-vehicle, N=6 mice for ABCB8 KO-vehicle, and N=5 mice for all other groups. ANOVA followed by post-hoc Tukey test was performed. * P=0.004 WT-vehicle vs. ABCB8 KO-vehicle. * P=1E-6 ABCB8 KO-vehicle vs. ABCB8 KO-BPD. I. Lipid peroxidation products in ABCB8 TG and NTG mice two days after I/R. N=6 mice for NTG-sham, and N=4 mice for the other groups. ANOVA followed by post-hoc Tukey test was performed. * P=0.024 NTG-sham vs. NTG-I/R. * P=0.016 NTG-I/R vs. TG-I/R. All data are expressed as mean ± SEM. N.S. = not signigicant.
Figure 4A-B. In-silico docking of veal2 and Prkcbb identified putative 4 interaction motifs in Prkcbb. Motif1-(549, 551-553, 559, 560) in Purple, motif2-(584-587) in Orange, motif3-(678-680) in Red, motif4-(844-846) in Green. The 8bp deletion in veal2-Δ8 RNA led to change in folding and altered interaction with Prkcbb. C. Scatter plot showing the number of animals which displayed the hemorrhage phenotype across the progeny derived from an outcross of veal2gib005Δ8/+ zebrafish injected with IVTs of different veal2 variants. Control indicates veal2gib005Δ8/+ zebrafish without complementation. Data from three different experiments plotted as individual values; the middle bar represents mean percentage and the error bar represents ± standard deviation. D. The site of interaction of the motif3 of veal2 (CLM) with the Prkcbb protein indicates that it lies in a previously known DAG binding site. Both RNA (pink) and protein (purple) structures are shown as ribbon models. The distances between nucleotides of motif-3 of veal2 and the 2 amino acids of Prkcbb are given in Å units. E. Relative kinase activity of human PRKCB2 with various concentrations of DAG without or with different concentrations of veal2 WT RNA (0, 0.025nM, 0.25nM, 2.5nM, 25nM). PRKCB2 activity without DAG was used for normalization. Data from three different experiments plotted as mean fold change values ± standard deviation. F. Abundance of Prkcbb protein in total cell (T), cytoplasmic (C) and membrane (M) fractions of cells from 2 dpf gib004Tg(fli1a:EGFP;gata1a:DsRed) and veal2gib005Δ8/+ zebrafish embryos. Arrowhead indicates Prkcbb enrichment in the membrane fraction of veal2gib005Δ8/+ zebrafish embryos. G. Relative quantification of Prkcbb localization in cytoplasmic and membrane fractions of control and veal2gib005Δ8/+ embryos. Data from three different experiments plotted as mean percentage values ± standard deviation.
Figure 8Confocal single plane images of 1151>Ama-RNAi animals and 1151>mCherry-RNAi. A. Forming DFM at 40 h APF stained with anti-Zfh1 (green), anti-Futsch (22c10, red) and anti-Kettin (white). White arrows point to the wing hinge, wings pointing left, anterior up. B. Forming IFM (DLM) at 16 h APF, stained with anti-Futsch (22c10, red), anti-Zfh1 (green) and DAPI (cyan). Yellow-dashed box indicates magnified area (bottom panel). Anterior up. C. DFM at 96 h APF stained with Phalloidin (red), anti-Kettin (green) and DAPI (blue). Anterior up, dorsal right. DFM are numbered in white as in (Lawrence, 1982). D. IFM at 96 h APF, transverse section (top) stained with anti-PS-integrin (green), Phalloidin (red) and DAPI (blue), dorsal up; sagittal section (bottom) stained with anti-Kettin (green), Phalloidin (red) and DAPI (blue), anterior up, dorsal right.
phosphorylation cause lymphomas in humanized mice more frequently than EBV wt.Figure 6(A) Experimental set-up. (Images of animals, syringes and blood collection tubes are derived from Servier Medical Art.) (B) Survival curve of humanized mice infected with 105 GRU EBV wt (n = 13), EBV S379A (n = 11) or EBV S457A/T465V EBV (n = 9). (C) Percentage of mice having macroscopically visible tumors at the day of sacrifice. Numbers within bars indicate the total number of mice with or without tumors in the respective groups. (D) Viral loads in spleen of infected humanized mice at the day of sacrifice. Error bars indicate mean ± SEM. Number of biological replicates per group: EBNA2 WT EBV: n = 13; EBNA2 S379A EBV: n = 11; EBNA2 S457A T465V EBV: n = 9. (E) Development of viral loads in blood of infected mice over a period of five weeks. Error bars indicate mean ± SEM. Number of biological replicates per group: EBNA2 WT EBV week 2 - 5: n = 8; EBNA2 S379A EBV week 2 - 5: n = 7; EBNA2 S457A T465V EBV week 2: n = 9, week 3: n = 8, week 4 and week 5: n = 7 (F) Flow-cytometric analyses of CD8+/CD4+ T cell ratios in the blood of infected mice over a period of five weeks. : EBNA2 WT EBV week 0, 2, 4, 5: n = 13, week 3: n = 8 ; EBNA2 S379A week 0, 2, 4, 5: n = 11, week 3: n = 7 ; EBNA2 S457A T465V week 0, 2: n = 12, week 3: n = 7, week 4: n = 10, week 5: n = 9.
Figure 5A. Sequence alignment of FGF21, FGF21SS and FGF19 in the mutated region. The mutation sites are indicated by boxes. Cysteines for disulfide bond formation are highlighted on a yellow background. The secondary structure of FGF21 is shown on the top. The N-terminal residue IDs of FGF21SS are indicated under the sequence. # is used to identify the mutated residues (codes are colored blue) and the same residues having different sequence number in FGF21SS from wild type. B. 2D 1H-15N HSQC spectra comparison of FGF21core (black) and FGF21SS/core (red) revealing the elimination of the loop conformation in FGF21SS. Assignments for FGF21core and FGF21SS/core are labeled respectively. C. Temperature Denaturation CD experiments indicating the significant increase of thermostability. CD value was recorded and normalized at 222 nm. Tm value of the proteins are fitted using Boltzmann function. D. Superimposition of FGF21SS/core (skyblue) and FGF21core (green) structure ensembles (10 energy refined structures each) in ribbon representation showing the structure conservation upon SS bond mutation. E. Cartoon representation of FGF21SS/core. Cys31#-Cys43# (between β2 and β3) and Cys75-Cys93 (between β6 and β9) SS bonds are highlighted and shown as stick representations.
Figure 2(A) ASY1:GFP and ASY1T142D:GFP localization in late G2 and leptotene of male meiocytes of the wildtype and CDKA;1T161D mutants. ASY3:RFP, highlighting chromsomes, was used as a marker for the staging of meiosis. Scale bar: 5 μm. (B) Kinase assays of CDKA;1-SDS, -TAM, and -CYCA3;1 complexes using ASY1 puried from baculovirus-infected insect cells as a substrate. The upper panel shows the autoradiograph. The control reaction without CDKA;1-cyclin complex indicates a background activity co-purified from insect cells. The lower panel indicates protein loading by coomassie briliant blue (CBB) staining. Arrowheads indicate ASY1 proteins and asterisks depict the relevant cyclin used which also gets phosphorylated in the assay. Numbers indicate the relative intensities of ASY1 bands. (C) The upper panel shows a phos-tag gel analysis of ASY11-300 and ASY11-300/T142V;T184V with and without CDKA;1-SDS kinase complexes using an anti-MBP antibody. The lower panel denotes loading of CDKA;1 using an anti-Strep antibody. Arrowheads represent the proteins as indicated.
Figure 2A. Nucleosomes (N) decorating a relaxed plate (P1); insets in the upper right show additional examples. Nucleosomes are not visible as individual units in typical compact plates (P2). Short compact interdigitated solenoids are shown in the main image (S) and in the bottom-left insets. Data information: Scale bars: 50 nm B-E Slice (B) and segmentations in three different orientations (C) of a large relaxed plate decorated with many nucleosomes on its right side. Slice (D) and segmentation (E) of a relaxed plate forming a tube decorated with nucleosomes. Data information: Scale bars: 50 nm insets, B-E) F. Structure of the decorative particles (like those shown in N, panel A) after subtomogram averaging. The final density map was filtered to 25 Å and fitted with the molecular structure of the nucleosome core particle (Protein Data Bank code 2CV5). Data information: Scale bars: 5 nm (F).
Figure 7A. Crossover frequency (black) and SNP density (purple) per 200 kb measured from the Col×Ct F2 population. The position of the centromere (CEN, vertical dashed line) and 420 FTL transgenes (red and green vertical lines) used to measure crossover frequency are indicated. B. Plots of chromosome 3 showing positions of Col/Col (black) or Col/Ct (purple) homozygous or heterozygous genotypes at the indicated marker positions (circles). The positions of the 420 FTL transgenes are indicated (red and green vertical lines). C. 420 crossover frequency (cM) in the HOM-HOM, HET-HET, HET-HOM, HOM-HET genotypes shown in B, in either wild type or msh2. Values for replicate individuals are plotted, in addition to the mean (red). D. As for C, but comparing genotypes in wild type or HEI10-OE backgrounds. Values for replicate individuals are plotted, in addition to the mean (red).
Figure 3(A) Scheme of ribosome footprint analysis using PACE reporter mRNAs. An example of a 28-nt footprint is shown. The relative ribosome density of each codon at the A site was calculated using the formula shown on the right. (B-E) Scatter plots showing a correlation between relative ribosome density (28-30-nt footprints; x-axis) and the codon effects measured by PACE (y-axis). Each dot represents a single codon. Graphs for all codons (B), codons for polar amino acids (C), nonpolar amino acids (D), and charged amino acids (E) are shown. Data information: the regression line is shown in blue and the 95% confidence interval is shown in gray. P-values were calculated using Student's t-test. (F-I) Scatter plots showing a correlation between relative ribosome density (28-30-nt footprints; x-axis) and the tRNA levels (y-axis, Each dot represents a single codon. Graphs for all codons (F), codons for polar amino acids (G), nonpolar amino acids (H), and charged amino acids (I) are shown. Data information: the regression line is shown in blue and the 95% confidence interval is shown in gray. P-values were calculated using Student's t-test.
Figure 3Mitochondrial DNA (A) in C2C12 myocytes transfected with siRNA or plasmid for mGPDH 24 h after differentiation nuclear-encoded OXPHOS genes (B in C2C12 myocytes transfected with siRNA or plasmid for mGPDH 24 h after differentiation respirometry analysis (C in C2C12 myocytes transfected with siRNA or plasmid for mGPDH 24 h after differentiation immunoblots of mGPDH, phospho-Thr172 AMPK (p-AMPK), total AMPK (AMPK), phospho-Ser79-ACC (p-ACC), total ACC and PGC1α and corresponding quantifications represent mGPDH, p-AMPK, p-ACC and PGC1α protein level in C2C12 myocytes transfected with siRN for mGPDH 24 h after differentiation immunoblots of mGPDH, phospho-Thr172 AMPK (p-AMPK), total AMPK (AMPK), phospho-Ser79-ACC (p-ACC), total ACC and PGC1α and corresponding quantifications represent mGPDH, p-AMPK, p-ACC and PGC1α protein levels (D-F) in C2C12 myocytes transfecte wit plasmid for mGPDH 24 h after differentiation Immunoblots of p-AMPK, p-ACC and PGC1α and corresponding quantifications represent p-AMPK, p-ACC and PGC1α protein levels (G in C2C12 myocytes transfected by mGPDH plasmid with the AMPK inhibitor compound C (CC) 24 h after differentiation mitochondrial DNA (H in C2C12 myocytes transfected by mGPDH plasmid with the AMPK inhibitor compound C (CC) 24 h after differentiation nuclear-encoded OXPHOS genes combined by NDUFS8, SDHb, Uqcrc1, COX5b and ATP5a1 (I) in C2C12 myocytes transfected by mGPDH plasmid with the AMPK inhibitor compound C (CC) 24 h after differentiation NAD+/NADH ratio (J in C2C12 myocytes transfected with siRNA or plasmid for mGPDH 24 h after differentiation immunoprecipitation analysis for PGC-1α acetyl-lysine (Ac-Lys) level (K) in C2C12 myocytes transfected with siRNA or plasmid for mGPDH 24 h after differentiation Immunoblot of c-myc and myogenin (L in C2C12 myocytes transfected with mGPDH plasmid with the AMPK inhibitor CC at 24 h (L,M after differentiation corresponding quantifications represent c-myc and myogenin protein levels (M) in C2C12 myocytes transfected with mGPDH plasmid with the AMPK inhibitor CC at 24 h (L,M representative images of MyHC immunofluorescence (N), fusion index (O) and the distribution of nuclei per myotube (P) in C2C12 myocytes transfected with mGPDH plasmid with the AMPK inhibitor CC a 72 h (N-P) after differentiation Immunoblots of p-AMPK, p-ACC, PGC1α and myogenin in C2C12 myocytes transfected with mGPDH plasmid with the CaMKKβ inhibitor STO-609 at 24 h after differentiation. Quantifications represent p-AMPK, p-ACC, PGC1α and myogenin protein levels Immunoblots of p-AMPK and p-ACC in C2C12 myocytes transfected with mGPDH plasmid with the Ca2+ chelator BAPTA-AM at 24 h after differentiation. Quantifications represent p-AMPK and p-ACC protein levels.
Ubqln4 interacts with LC3.Figure 2Ubqln4 interacts with LC3. Confocal images of (A) Hela cells transfected with GFP‐LC3 and RFP‐Ubqln4 and starved for 1 h or (B) Hela cells starved for 1 h and stained for endogenous LC3 and Ubqln4. The squares show colocalization. The insets show the enlargement of the indicated areas. Linescan of the white line in the merged image indicates the intensities of the fluorescent molecules in the cross‐section. (C) 293 cells were transfected with GFP‐S, GFP‐S‐Ubqln1 or GFP‐S‐Ubqln4 and treated with 50 μM chloroquine for 14 h. GFP was immunoprecipitated and analysed by western blot using anti‐GFP and anti‐LC3 antibodies. (D) Recombinant His‐Ubqln1, His‐Ubqln4, His‐S5a or His‐p62 was mixed with LC3 covalently coupled to agarose beads. The beads were washed, eluted and detected by western blot using anti‐His antibody. The extent of Ubqln1, Ubqln4, s5a or p62 co‐immunoprecipitation with LC3 was measured. Bars for Ubqln1, Ubqln4 and s5a represents the average and s.e.m. of four independent experiments. *P‐value=0.0094. (E) 293 cells were transfected with GFP‐S‐Ubqln4 WT, GFP‐S‐Ubqln4 ΔSTI1 12, GFP‐S‐Ubqln4 Δ between STI1 or GFP‐S‐Ubqln4 ΔSTI1 34 and treated with 50 μM chloroquine for 14 h. GFP was immunoprecipitated and analysed by western blot using anti‐GFP and anti‐LC3 antibodies. GFP, green fluorescent protein; IP, immunoprecipitation; LC3, microtubule‐associated protein light chain 3; Ubqln, Ubiquilin; WCL, whole‐cell lysate; WT, wild‐type.
Figure 2A Schematic representation of the proteomic experiments used to isolate novel proteins enriched at the Shigella-septin cage. (a) A SEPT6-STREP-FLAG construct was (b) transfected in HeLa cells for 24h. (c) Cells were infected with S. flexneri AfaI for 4 h, harvested, and (d) SEPT6-STREP-FLAG cage-associated proteins isolated through co-immunoprecipitation (Co-IP). B HeLa cells stably expressing SEPT6-GFP were infected with S. flexneri AfaI for 4 h, harvested, and then tested with Co-IP. Empty vector and/or uninfected HeLa cells were used in parallel as control. GFP-trap magnetic agarose beads were used to isolate SEPT6-GFP from cells, and extracts were immunoblotted for SEPT6, GFP, SEPT7, or p62. The lysates were immunoblotted for GAPDH as control for cellular protein levels. C The protein pulldown experiments identified 56 proteins putatively associated to the Shigella-septin cage, and then categorised into 6 groups based on analysis from the Gene ontology database. See also Dataset EV1.