Journal:

Article Title: Inhibition of Rac GTPase triggers a c-Jun- and Bim-dependent mitochondrial apoptotic cascade in cerebellar granule neurons

doi: 10.1111/j.1471-4159.2005.03252.x

Figure Lengend Snippet: ToxB induces apoptosis and elicits a selective caspase-mediated degradation of Rac1 GTPase in CGNs. (a) CGNs were incubated for 24 h in complete medium (containing serum and 25 mm KCl) in the presence of either vehicle (Veh; 2 μg/mL BSA in PBS) or ToxB (40 ng/mL). Following incubation, cells were fixed in paraformaldehyde and nuclei were stained with Hoechst dye. CGNs exposed to ToxB showed marked nuclear condensation and fragmentation characteristic of apoptosis. Scale bar, 10 μm. (b) Representative immunoblots of lysates obtained from CGNs incubated as described in (a). The blots shown were obtained from the BD PowerBlot™ (see Experimental procedures for details) using specific monoclonal antibodies against Rac1 (upper blots), RhoA (middle blots), or Cdc42 (lower blots). The immunoreactive Rho family GTPases are indicated by the arrows. Other protein bands apparent on the membranes represent adjacent lanes of the PowerBlot™ that were probed for distinct proteins and are shown to give an indication of the overall equality of protein loading. (c) The fold change in protein content induced by 24 h of ToxB treatment for each of the Rho family members blotted for in (b) is shown. Data represent the means ± range of duplicate PowerBlot™ comparisons for each GTPase. The dotted lines indicate no change (-1 to +1 fold). (d) CGNs were exposed for 8 h to ToxB in the absence or presence of either a pan-caspase inhibitor (Boc-Asp(OMe)-FMK; 200 μm) or zVAD-FMK; 200 μm) or a proteasome inhibitor (MG132; 10 μm). Following incubation, cell lysates were immunoblotted for Rac1.

Article Snippet: The blots shown were obtained from the BD PowerBlot™ (see Experimental procedures for details) using specific monoclonal antibodies against Rac1 (upper blots), RhoA (middle blots), or Cdc42 (lower blots).

Techniques: Incubation, Staining, Western Blot

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: The previously identified TEAD4 148 isoform is also shown for comparison. Schematic indicates the TEA DNA-binding domain (TEAD), a putative nuclear localization signal (NLS), a proline rich domain (PRD) and serine-threonine-tyrosine (STY) domains.

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Comparison, Binding Assay

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: The TEAD4 216 isoform can repress expression from the full length human VEGF promoter F1–R3 (p<0.01, n = 9) in contrast to the TEAD4 434 and TEAD4 148 enhancers.

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Expressing

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: The TEAD4 216 isoform can repress expression from the human VEGF promoter (F2–R3) that lacks the HRE (p<0.01, n = 6). The TEAD4 148 and TEAD4 311 enchancer isoforms do not require the HRE to promote reporter gene expression (p<0.001). However the full length TEAD4 434 did not significantly enhance expression (p>0.02).

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Expressing, Gene Expression

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: The TEAD4 216 isoform can competitively repress expression from the human VEGF promoter in the presence of either the TEAD4 434 or TEAD4 148 enhancer isoforms (p<0.005, n = 6).

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Expressing

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: The TEAD4 216 isoform can still repress expression mediated from the human VEGF promoter (F1–R3) that includes the HRE sequence under conditions of hypoxia (p<0.001, n = 6).

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Expressing, Sequencing

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: Transient plasmid transfection or stable lentiviral mediated introduction of TEAD4 216 into human cells results in reduction of native VEGF protein. Solid bars represent VEGF 165 levels, quantified by ELISA, within conditioned media collected 48 hours after transfection of pcDNA plasmid vector containing the TEAD4 216 isoform into ( A ) 293T cells, ( B ) ARPE-19 retinal pigment epithelial (RPE) cells in culture (n = 4), (p<0.05). ( C ) Lentiviral (LV) expression of the TEAD4 216 isoform in human D407 RPE cells can inhibit endogenous VEGF production. Solid bars represent VEGF 165 levels, quantified by ELISA, within conditioned media collected 48 hours after transduced RPE cells were FAC sorted and plated (n = 3),(p<0.02).

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Plasmid Preparation, Transfection, Enzyme-linked Immunosorbent Assay, Expressing

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: A Cyquant cell proliferation assay determining the DNA content of cells indicate that 4 days after initial seeding of equal cell numbers the control untransduced cells proliferate at a faster rate than the LV-TEAD4 216 transduced cells (n = 8, p<0.05).

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: CyQUANT Assay, Proliferation Assay, Control

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: TEAD4 isoforms were expressed as fusion proteins within 293T cells with various fluorescent proteins (FP) (TEAD4 434 -Green-FP, and TEAD4 148 -Yellow-FP (pseudo-colored green) and TEAD4 216 -Green-FP) to visualize cellular localization.

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques:

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: RT-PCR for TEAD4 from choroidal, retinal and iris tissue isolated from a non-human primate eye ( Rhesus macaque ) 24 hrs after occlusion of the central retinal artery (CRAO), indicates that the full length TEAD4 434 transcript is increased and the TEAD4 148 enhancer isoform is produced in the lasered eye. L = Lasered CRAO eye; C = control eye.

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Reverse Transcription Polymerase Chain Reaction, Isolation, Produced, Control

Journal: PLoS ONE

Article Title: The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4 216 , Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

doi: 10.1371/journal.pone.0031260

Figure Lengend Snippet: Photomicrograph showing a subretinal neovascular membrane in a human ocular tissue section (Fast Red, TEAD4; hematoxylin counterstain; original magnification x 100). Insert (location indicated by rectangle; original magnification x 1000) shows positive staining for TEAD4 by vascular endothelium of a choroidal new vessel that has bridged the elastic lamina of Bruch’s membrane.

Article Snippet: TEAD4 was visualized in tissue sections, according to a previously published indirect immunostaining method , using rabbit polyclonal anti-TEAD4 antibody (made to order by Genemed Synthesis, San Antonio, TX, see for details of custom made antibody) or, as negative control, rabbit IgG (Vector Laboratories, Burlingame, CA), at a concentration of 11 ug/mL.

Techniques: Membrane, Staining

Journal: Current biology : CB

Article Title: Protein interaction analysis provides a map of the spatial and temporal organization of the ciliary gating zone

doi: 10.1016/j.cub.2017.06.044

Figure Lengend Snippet: (A) Schematic of BiFC assay. The N- and C-terminal halves of a YFP (half-stars labeled N or C), when fused to proteins of interest, can reconstitute a fluorescent protein only when brought in close proximity via the interactions of their fusion partners. TZ, transition zone proteins. MT, axonemal microtubules. (B) Localization of Nup62-YC (left) and KIF17-YN (right) expressed in the absence of a BiFC partner. Arl13b-Cer or acetylated α-tubulin (AcTubulin) was used as a cilium marker. Images are of cropped regions containing the cilium, contrast-enhanced for viewing, with a schematic of the observed fluorescence localization shown below each fluorescence image. (C–E) Representative images and schematic depictions show the locations of BiFC interactions detected for Nup62-YC with (C) kinesin-2 motor KIF17-YN, (D) KIF17 with mutated CLS (KIF17ΔCLS-YN), or (E) non-ciliary protein (YN-SAH-FKBP). Nup62-YC was detected with an antibody to the HA tag (Nup62-YC-HA) whereas the KIF17 and SAH constructs were detected with an antibody to the Myc tag. See Table S1 for full description of constructs. See Figure S1 for uncropped images. (F) Quantification of the locations of BiFC interactions. The number of cells observed for each BiFC location category is indicated on the bar graph. (G–I) Time course of interaction between Nup62 and KIF17. Nup62-FRB-Cer and mCit-KIF17-FKBP were co-expressed and the cells were fixed after treatment with (G) ethanol (- Rapamycin control), (H) 10 min Rapamycin, or (I) 30 min Rapamycin. Graphs show the mean fluorescence of Nup62-FRB-Cer at the base or tip of the cilium (normalized to the fluorescence at the base) or the mean fluorescence of mCit-KIF17-FKBP at the base or tip of the cilium (normalized to the fluorescence at the tip). *p<0.01 compared to control (-rapamycin) by Student’s t-test. Error bars, S.D. n = 10–14 cells each.

Article Snippet: METHOD DETAILS Antibodies and plasmids Commercial antibodies for acetylated α-tubulin (1:10,000, Sigma, 6-11B-1) and the epitope tags Myc (1:500, Sigma, C3956) and HA (1:1,000, Covance, 16B12) were used.

Techniques: Bimolecular Fluorescence Complementation Assay, Labeling, Marker, Fluorescence, Construct