Journal: Cancer Research

Article Title: Tumor-Associated MICA Is Shed by ADAM Proteases

doi: 10.1158/0008-5472.can-07-6768

Figure Lengend Snippet: Figure 5. ADAM10 silencing impairs constitutive MICA shedding. A to C, ADAM10 siRNA expression in 293T-MICA cells down-regulates ADAM10 transcripts and total and cell surface ADAM10 protein expression. A, relative copy numbers of ADAM10 and MICA transcripts in 293T-MICA cells 48 h after transfection with pSuper-puro or pSuper-ADAM10 and in untreated 293T-MICA were determined by real-time PCR after normalization with 18S rRNA. B, total cellular ADAM10 protein was determined in lysates of 293T-MICA cells 72 h after transfection and of untreated 293T-MICA cells by immunoblotting. C, surface expression of MICA or ADAM10 (gray histograms) was determined 72 h after transfection. Overlays are respective stainings of untreated 293T-MICA cells (thick lines) and isotype control stainings of transfected 293T-MICA cells (thin lines). D, sMICA released from 293T-MICA transfected with pSuper or pSuper-ADAM10 was determined by sMICA ELISA in medium used for culture from 72 to 75 h after transfection and compared with 3-h culture supernatants of untreated 293T-MICA. **, P < 0.001.

Article Snippet: Antibodies specific for human ADAM10 (mAb 163003) and human ADAM17 (mAb 111633) were from R&D Systems.

Techniques: Expressing, Transfection, Real-time Polymerase Chain Reaction, Western Blot, Control, Enzyme-linked Immunosorbent Assay

Journal: Cancer Research

Article Title: Tumor-Associated MICA Is Shed by ADAM Proteases

doi: 10.1158/0008-5472.can-07-6768

Figure Lengend Snippet: Figure 6. ADAM17 mediates PMA-stimulated MICA shedding. A, transient knockdown of ADAM10 and/or ADAM17 in 293T-MICA and HeLa-MICA cells by transfection of siRNA results in decreased levels of cell surface ADAM10 and/or ADAM17. Stainings of MICA, ADAM10, and ADAM17 (gray histograms) 72 h after transfection with control siRNA (nontargeting), ADAM10 siRNA, ADAM17 siRNA, or a mixture of ADAM10/ADAM17 siRNA are overlayed with stainings of untreated cells (thick lines) and isotype controls (dotted lines). Histograms show cells that have not been treated with PMA. B and C, levels of sMICA were determined by sMICA ELISA in supernatants 72 to 75 h after transfection (B) and after subsequent PMA treatment (100 ng/mL) at 75 h after transfection (supernatants of 75–77.5 h after transfection; C). *, P = 0.003; **, P < 0.001. ns, not significant. Significance was calculated for difference to control-treated (nontargeting siRNA treated) cells.

Article Snippet: Antibodies specific for human ADAM10 (mAb 163003) and human ADAM17 (mAb 111633) were from R&D Systems.

Techniques: Knockdown, Transfection, Control, Enzyme-linked Immunosorbent Assay

Journal: Alzheimer's Research & Therapy

Article Title: Selective reduction of ADAM10 in brain and cerebrospinal fluid of Alzheimer’s disease patients

doi: 10.1186/s13195-026-02007-6

Figure Lengend Snippet: Protein levels of ADAM10 species and mRNA in AD brain samples. Non-dementia controls (NDC, n = 13) and AD extracts ( n = 16) from prefrontal cortex were resolved by SDS-PAGE/electrophoresis prior to western blot assay. Each individual ADAM10 immunoreactive band was quantified, and levels normalized using GAPDH. A Representative western blot of ADAM10 using the rabbit anti- C-terminal region monoclonal antibody (ab124695, Abcam) and GAPDH with monoclonal antibody (60004-1-Ig, Proteintech) ( B ) Densitometric quantification of ADAM10 immunoreactive bands assigned to immature (iADAM10) and C mature (mADAM10) species normalized with respect to GAPDH. D Ratio mADAM10/iADAM10 that represents the amount of mature species vs. immature. E Relative mRNA levels of ADAM10 in NDC vs. AD patients’ samples were analyzed by qRT-PCR. Transcript levels were calculated by the comparative 2 − ΔCt method with respect to GAPDH. The graphs represent mean ± SEM. Significant P < 0.05 values assayed by t-test are indicated

Article Snippet: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) amplification was performed using QuantStudioTM 3 Real-Time PCR System (Applied Biosystems) with TaqMan Gene Expression PCR Master Mix (Applied Biosystems) and specific TaqMan Gene Expression Assay for ADAM10 (Hs00153853_m1).

Techniques: SDS Page, Electrophoresis, Western Blot, Quantitative RT-PCR

Journal: Alzheimer's Research & Therapy

Article Title: Selective reduction of ADAM10 in brain and cerebrospinal fluid of Alzheimer’s disease patients

doi: 10.1186/s13195-026-02007-6

Figure Lengend Snippet: Aβ affects ADAM10 levels, but not ADAM17, in SH-SY5Y-differentiated neurons. SH-SY5Y cell cultures were differentiated to neurons with 10 µM retinoic acid treatment and then treated with 3 µM of Aβ42 for 48 h. A Representative western blot of control ( C ) and Aβ-treated (Aβ42) cell samples that was resolved with the anti C-terminal ADAM10 antibody (ab124695, Abcam). B Densitometric quantification of immunoreactive bands of iADAM10 and C mADAM10 with respect to GAPDH. Values represent the percentage with respect to control. D Values of the ratio between mature vs. immature species of ADAM10 (mADAM10/iADAM10). E Representative western blot of ADAM17 species resolved with anti-ectodomain region ADAM17 antibody (AF9301, R&D Systems) and the respective quantifications of F iADAM17 and G mADAM17 immunoreactive bands normalized to the ubiquitous protein GAPDH. H Result of the ratio of mADAM17/iADAM17. The graphs represent mean ± SEM of n = 12 samples of 3 independent experiments. Significant P < 0.05 values assayed by t-test are indicated

Article Snippet: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) amplification was performed using QuantStudioTM 3 Real-Time PCR System (Applied Biosystems) with TaqMan Gene Expression PCR Master Mix (Applied Biosystems) and specific TaqMan Gene Expression Assay for ADAM10 (Hs00153853_m1).

Techniques: Western Blot, Control

Journal: Alzheimer's Research & Therapy

Article Title: Selective reduction of ADAM10 in brain and cerebrospinal fluid of Alzheimer’s disease patients

doi: 10.1186/s13195-026-02007-6

Figure Lengend Snippet: ADAM10 and ADAM17 CSF levels are not altered with aging. Representative blots of CSF samples of non-AD control individuals with large age amplitude (the age of the subjects are shown on top) probed with ( A ) anti-ADAM10 polyclonal antibody to ectodomain region (OAGA02442, Aviva) and ( B ) ADAM17 antibody towards the ectodomain region (AF9301, R&D Systems). Correlations between the age and the levels of ADAM10 and ADAM17 species ( C ) iADAM10, ( D ) mADAM10, ( E ) sADAM10, ( F ) iADAM17, ( G ) mADAM17 and ( H ) sADAM17 were assayed. All graphs include their Spearman coefficient (r) and the P value. None of the ADAM10 species nor ADAM17 correlate with age. * Refers to unspecific bands

Article Snippet: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) amplification was performed using QuantStudioTM 3 Real-Time PCR System (Applied Biosystems) with TaqMan Gene Expression PCR Master Mix (Applied Biosystems) and specific TaqMan Gene Expression Assay for ADAM10 (Hs00153853_m1).

Techniques: Control

Journal: Alzheimer's Research & Therapy

Article Title: Selective reduction of ADAM10 in brain and cerebrospinal fluid of Alzheimer’s disease patients

doi: 10.1186/s13195-026-02007-6

Figure Lengend Snippet: Levels of ADAM10 and ADAM17 species in AD CSF samples. Analysis of CSF ADAM10 and ADAM17 in non-AD controls (NADC) and in AD patients. A Representative blot of CSF probed against anti-ADAM10 ectodomain antibody. Quantification of immunoreactive band values obtained from B iADAM10, C mADAM10 and D sADAM10. E Values of the ratio between mature vs immature species of ADAM10 (mADAM10/iADAM10). F Representative blot of CSF probed against ADAM17 and the quantifications of the immunoreactivity of the bands for G iADAM17, H mADAM17 and I sADAM17. J Result of the ratio mADAM10/iADAM10. The graphs represent mean ± SEM. Significant P < 0.05 values assayed by t-test are indicated. * Refers to unspecific bands

Article Snippet: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) amplification was performed using QuantStudioTM 3 Real-Time PCR System (Applied Biosystems) with TaqMan Gene Expression PCR Master Mix (Applied Biosystems) and specific TaqMan Gene Expression Assay for ADAM10 (Hs00153853_m1).

Techniques:

Journal: World Journal of Gastroenterology

Article Title: Fas counterattack in cholangiocarcinoma: A mechanism for immune evasion in human hilar cholangiocarcinomas

doi: 10.3748/wjg.v7.i6.860

Figure Lengend Snippet: FasL positive in human hilar cholangiocarcinom as (brown). SABC × 200

Article Snippet: Detection of FasL expression and CD45 positive cells was performed using a rabbit polyclonal anti-human FasL specific IgG and a mouse anti-human monoclonal antibody on paraffin sections of human cholangiocarcinoma respectively (Boster Biological Technology Company, Wuhan, China).

Techniques:

Journal: World Journal of Gastroenterology

Article Title: Fas counterattack in cholangiocarcinoma: A mechanism for immune evasion in human hilar cholangiocarcinomas

doi: 10.3748/wjg.v7.i6.860

Figure Lengend Snippet: Expression of FasL in cholangiocarcinoma cell line. QBC939 × 200

Article Snippet: Detection of FasL expression and CD45 positive cells was performed using a rabbit polyclonal anti-human FasL specific IgG and a mouse anti-human monoclonal antibody on paraffin sections of human cholangiocarcinoma respectively (Boster Biological Technology Company, Wuhan, China).

Techniques: Expressing

Journal: World Journal of Gastroenterology

Article Title: Fas counterattack in cholangiocarcinoma: A mechanism for immune evasion in human hilar cholangiocarcinomas

doi: 10.3748/wjg.v7.i6.860

Figure Lengend Snippet: Expression of FasL mRNA in human cholangiocarcinoma cells QBC939. M: DL 2000 Marker; 1: FasL; 2: FasL+β-actin

Article Snippet: Detection of FasL expression and CD45 positive cells was performed using a rabbit polyclonal anti-human FasL specific IgG and a mouse anti-human monoclonal antibody on paraffin sections of human cholangiocarcinoma respectively (Boster Biological Technology Company, Wuhan, China).

Techniques: Expressing, Marker

Journal: World Journal of Gastroenterology

Article Title: Fas counterattack in cholangiocarcinoma: A mechanism for immune evasion in human hilar cholangiocarcinomas

doi: 10.3748/wjg.v7.i6.860

Figure Lengend Snippet: Western blotting of FasL protein with mAb from QBC939 cell cultures clone 33 from QBC939 cell cultures

Article Snippet: Detection of FasL expression and CD45 positive cells was performed using a rabbit polyclonal anti-human FasL specific IgG and a mouse anti-human monoclonal antibody on paraffin sections of human cholangiocarcinoma respectively (Boster Biological Technology Company, Wuhan, China).

Techniques: Western Blot

Journal: Journal of Cancer

Article Title: Modification of proteolytic activity matrix analysis (PrAMA) to measure ADAM10 and ADAM17 sheddase activities in cell and tissue lysates

doi: 10.7150/jca.20779

Figure Lengend Snippet: PrAMA approach for solid tissue analysis. ( A ) Summary table and heat-map show 7 FRET-based polypeptide substrates with cleavage sequences (HomoPhe, homophenylalanine; Cha, 3-cyclohexylalanine), endogenous-protein protease substrates from which the peptides were designed, and the catalytic efficiencies with which various recombinant proteases cleave the substrates (for more information see ref. 35). ( B ) Workflow illustrates PrAMA for cell and tissue lysates. ( C ) Proof-of-principle is shown for PrAMA applied to known solutions, and one combination, of recombinant enzymes (abbreviations: rM2, recombinant MMP2; rA10, recombinant ADAM10; rA17, recombinant ADAM17). Normalized cleavage rates for each of the 4 solutions were measured across the 7 substrates (left), and data were interpreted using known enzyme-substrate preferences [as in ( A )] to infer which recombinant enzymes were present in the mixture. Actual mixture composition (top right) and PrAMA results (bottom right) are presented. ( D ) Surface plots depict three-dimensional “systematic PrAMA” inference as a function of the two parameters sensitivity (Syntherror) and specificity (Sigmathreshold). Processing data of seven substrates obtained with recombinant MMP2, ADAM10, and ADAM17 were analyzed by PrAMA across varying combinations of Syntherror and Sigmathreshold parameters to reveal how these two parameters influence PrAMA sensitivity and specificity. The three rows of surface-plots correspond to the analyzed individual three recombinant enzyme solutions (rMMP2, rADAM10 and rADAM17), and the three columns of surface-plots correspond to the three individual PrAMA-inferred enzyme activities from these solutions (MMP2a, ADAM10sa and ADAM17sa). The color scale ranges from red to blue, which reflects the surface heights as labeled on the vertical axis. The three-dimensional surface plots shown in the figure depict a representation of the two-dimensional “systematic PrAMA” shown in Figs. C- H. In the latter cases (Figs. C-H) and the rest of presented data, Syntherror is held constant (0.5) across a range of Sigmathreshold values. The experimental details follow those described for Fig. C.

Article Snippet: Human ADAM10 and ADAM17 siRNA (pools of three target specific siRNA), control (scrambled siRNA), siRNA transfection medium, and siRNA transfection reagent were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Recombinant, Labeling

Journal: Journal of Cancer

Article Title: Modification of proteolytic activity matrix analysis (PrAMA) to measure ADAM10 and ADAM17 sheddase activities in cell and tissue lysates

doi: 10.7150/jca.20779

Figure Lengend Snippet: Modified PrAMA detects ADAM10sa and ADAM17sa presence in wild-type and absence in ADAM10 -/- and ADAM17 -/- MEF lysates, respectively. ADAM10 +/- , ADAM10 -/- ( A, C, E ) , ADAM17 +/+ and ADAM17 -/- ( B, D, F ) MEFs were activated with PMA/Ionomycin and lysed. The 150 μL Tris solutions of 2 μg cell lysates, 10 μM PEPDAB substrates and 0.5% Roche/Halt protease inhibitors were incubated for 4 h at 37 o C, and the developed fluorescence was recorded hourly using TECAN fluorimeter. The presented experiments are representative of 5 performed. The enzymatic activity data are shown as pM means of duplicate measurements ± SD of the processed substrates ( A, B ). The decreased processing of PEPDAB005, 010, 011, 014 and 022 with knockout MEF lysates is significant ( ADAM10 +/- vs ADAM10 -/- MEFs and ADAM17 +/+ vs ADAM17 -/- MEFs: p<0.0001 and p<0.001, p=0.0056 and p<0.001, p=0.0016 and p=0.0025, p=0.0016 and p=0.009, and p=0.006 and p=0.004, respectively). Systematic PrAMA data using Syntherror/Sigmathreshold parameters 0.5/0.0 to 0.5/2.0 and 0.5/0.0 to 0.5/1.0 are presented as ADAM10sa AU ( C ) and ADAM17sa AU ( D ) , respectively. Standard errors of PrAMA data were 2.5% to 5.8%. True-positive ADAM10sa and ADAM17sa in ADAM10 +/- and ADAM17 +/+ MEFs, respectively, are presented as % of specific enzyme activities in wild-type MEFs, containing both the true-positive and false-positive activities, relative to ADAM10 -/- (E) and ADAM17 -/- (F) ME Fs containing only the false-positive activities, respectively.

Article Snippet: Human ADAM10 and ADAM17 siRNA (pools of three target specific siRNA), control (scrambled siRNA), siRNA transfection medium, and siRNA transfection reagent were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Modification, Incubation, Fluorescence, Activity Assay, Knock-Out

Journal: Journal of Cancer

Article Title: Modification of proteolytic activity matrix analysis (PrAMA) to measure ADAM10 and ADAM17 sheddase activities in cell and tissue lysates

doi: 10.7150/jca.20779

Figure Lengend Snippet: Modified PrAMA detects decreases of ADAM10sa and ADAM17sa in human cancer cells after silencing of the corresponding enzyme genes. H441 cells were transfected with human ADAM10 siRNA, ADAM17 siRNA or scrambled siRNA or were treated with transfection reagents alone for 48 h. In some experiments, control cells were also untreated. ( A , B ) Cells were stained with PE-conjugated IgG control mAb, anti-human ADAM10 ( A ) or anti-human ADAM17 mAb ( B ) and analyzed by flow cytometry. Empty histograms represent MFI of H441 cells stained with isotype control mAb. Dark gray histograms represent MFI of H441 cells treated with scrambled siRNA and stained with PE-conjugated anti-ADAM10 ( A ) or anti-ADAM17 ( B ) mAbs. Light grey histograms represent MFI of H441 cells treated with ADAM10 ( A ) or ADAM17 ( B ) siRNA and stained with PE-conjugated anti-ADAM10 or anti-ADAM17 mAbs, respectively. Data are from a representative experiment of 8 similar performed ( Suppl. Figs. A, 4B ). In the presented experiment, ADAM10 and ADAM17 siRNA induced 90% and 45% decreases of ADAM10 and ADAM17 protein expression on H441 cell surface, respectively. ( C ) After performing transfection, 2 μg of cell lysates were tested for processing PEPDABs in the presence of 0.5% Roche/Halt protease inhibitors. Data are from one of seven similar experiments performed. They are pM means of duplicate measurements ± SD of processed substrates. Processing of PEPDAB substrates was differently decreased in H441 cells transfected with ADAM10 or ADAM17 siRNA (PEPDAB005: p=0.0023 and p=0.0077; PEPDAB010: p=0.012 and p=0.017; PEPDAB011: p=0.048 and p=0.06; and PEPDAB014: p=0.07 and p=0.0.019, respectively). The substrate processing data obtained at 4 h of incubation were analyzed using the systematically increased Syntherror/Sigmathreshold parameters from 0.5/0.0 to 0.5/2.0. The resulted PrAMA ADAM10sa ( D) and ADAM17sa ( E ) AU are shown. PrAMA standard errors were 1.3% to 8.9%. Decreases of ADAM10sa and ADAM17sa in ADAM10 ( F ) and ADAM17 ( G ) siRNA transfected H441 cells, respectively, are presented as % of the specific enzyme activities in the siRNA transfected cells relative to transfection reagent-treated cells. Proportion-equation analysis of PrAMA-ADAM10sa and PrAMA-ADAM17sa of rADAM10 and rADAM17 vs the scrambled siRNA-transfected H441-cell lysates, respectively, showed that 10 μg of H441-cell lysate contained 17.0 ng of ADAM10sa and 1.65 ng of ADAM17sa.

Article Snippet: Human ADAM10 and ADAM17 siRNA (pools of three target specific siRNA), control (scrambled siRNA), siRNA transfection medium, and siRNA transfection reagent were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Modification, Transfection, Control, Staining, Flow Cytometry, Expressing, Incubation

Journal: Journal of Cancer

Article Title: Modification of proteolytic activity matrix analysis (PrAMA) to measure ADAM10 and ADAM17 sheddase activities in cell and tissue lysates

doi: 10.7150/jca.20779

Figure Lengend Snippet: Modified PrAMA efficiently measures ADAM10sa and ADAM17sa in multiple human tumor-tissue specimens. ( A ) NSCLC tumor lysates contain less ADAM10 than ADAM17 protein. ADAM10 and ADAM17 were quantified in lysates of 5 human NSCLC tumor tissues using ELISAs. Presented data are means pg/10 μg lysates of ADAM10 and ADAM17 proteins ± SD of 5 tumor tissues. ( B ) NSCLC tissue lysates process high amounts of PEPDAB05, 008 and 010, moderate amounts of PEPDAB014 and 022, and low amounts of PEPDAB011 and 013. Duplicates of 150 μL of Tris-based reaction buffer supplemented with NSCLC tumor-tissue lysates (10 μg/replicate), 0.5% Roche/Halt protease inhibitors and 10 μM of PEPDABs were incubated at 37 o C, and fluorescence quantified hourly for 4 h. Data are pM means of duplicate measurements ± SD of processed substrates with the 5 tissue lysates. ( C , E ) PrAMA ADAM10sa and ( D , F ) PrAMA ADAM17sa are robust but quantitatively different. The 4 h substrate processing data were analyzed using the systematically increased Syntherror/Sigmathreshold scripts from 0.5/0.0 to 0.5/2.0. Presented data are of the individual tissue lysates. PrAMA standard errors were 2.7% to 5.4%. Proportion-equation analysis of rADAM10 and rADAM17, and the tissue lysate PrAMA-ADAM10sa and PrAMA-ADAM17sa, respectively, showed that 10 μg of these tissue lysates contained 20.0 ng of ADAM10sa and 1.25 ng of ADAM17sa.

Article Snippet: Human ADAM10 and ADAM17 siRNA (pools of three target specific siRNA), control (scrambled siRNA), siRNA transfection medium, and siRNA transfection reagent were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Modification, Incubation, Fluorescence

Journal: PLoS ONE

Article Title: A Soluble Form of the Giant Cadherin Fat1 Is Released from Pancreatic Cancer Cells by ADAM10 Mediated Ectodomain Shedding

doi: 10.1371/journal.pone.0090461

Figure Lengend Snippet: The figure in A) shows a representative Western blot. PaCa44 and Panc1 cells were incubated in serum free medium for two days with the addition of either the broad spectrum metalloprotease inhibitor Batimastat (B, 10 µM), or the ADAM10-specific inhibitor GI245023X (GI, 5 µM) or solvent as a control (C, DMSO). Secretomes, 8 µg protein per sample, were analyzed by Western blot analysis using the Fat1 ECD2 polyclonal antibody. Blotting against transferrin was used as a loading control. (B) Fat1 specific signals from three independent experiments were quantified and normalized with the transferrin signals. The bar graphs show the mean values +/− S.E.M. from three experiments.

Article Snippet: The ADAM10 specific inhibitor GI254023X , and the broad spectrum metalloproteinase inhibitor Batimastat (Tocris Bioscience, Bristol, UK) were both dissolved in DMSO as stock solutions prior to dilution and use.

Techniques: Western Blot, Incubation, Solvent, Control

Journal: PLoS ONE

Article Title: A Soluble Form of the Giant Cadherin Fat1 Is Released from Pancreatic Cancer Cells by ADAM10 Mediated Ectodomain Shedding

doi: 10.1371/journal.pone.0090461

Figure Lengend Snippet: ADAM10 expression was suppressed by siRNA mediated knockdown experiments using ADAM10-specific and control siRNA pool's (30 µM each). A) Western blot analysis (75 µg of cell lysates) confirms efficient knockdown of B) Two independent experiments provided similar results as indicated by the normalized quantification.

Article Snippet: The ADAM10 specific inhibitor GI254023X , and the broad spectrum metalloproteinase inhibitor Batimastat (Tocris Bioscience, Bristol, UK) were both dissolved in DMSO as stock solutions prior to dilution and use.

Techniques: Expressing, Knockdown, Control, Western Blot

Journal: PLoS ONE

Article Title: A Soluble Form of the Giant Cadherin Fat1 Is Released from Pancreatic Cancer Cells by ADAM10 Mediated Ectodomain Shedding

doi: 10.1371/journal.pone.0090461

Figure Lengend Snippet: Western blot analysis of secretome fractions (8 µg) from PaCa44 and Panc1 cells after ADAM10 knockdown (compare ) showed significant reductions in Fat1 ectodomain shedding. Again, the two experiments provided similar results as indicated by the normalized quantification.

Article Snippet: The ADAM10 specific inhibitor GI254023X , and the broad spectrum metalloproteinase inhibitor Batimastat (Tocris Bioscience, Bristol, UK) were both dissolved in DMSO as stock solutions prior to dilution and use.

Techniques: Western Blot, Knockdown

Journal: PLoS ONE

Article Title: A Soluble Form of the Giant Cadherin Fat1 Is Released from Pancreatic Cancer Cells by ADAM10 Mediated Ectodomain Shedding

doi: 10.1371/journal.pone.0090461

Figure Lengend Snippet: Full-length Fat1 can be expressed on the cell surface; alternatively, the full length protein can be cut by a furin-type protease to yield the membrane-integrated heterodimer as described by Sadeqzadeh et al. and designated as “classical processing”. Another alternative is ectodomain shedding as described in this work, where the full-length protein is cut by ADAM10 into a p60 C-terminal remnant fragment (which may be further processed) and a gp515 soluble form released into the secretome.

Article Snippet: The ADAM10 specific inhibitor GI254023X , and the broad spectrum metalloproteinase inhibitor Batimastat (Tocris Bioscience, Bristol, UK) were both dissolved in DMSO as stock solutions prior to dilution and use.

Techniques: Membrane

Journal: PLoS ONE

Article Title: A Soluble Form of the Giant Cadherin Fat1 Is Released from Pancreatic Cancer Cells by ADAM10 Mediated Ectodomain Shedding

doi: 10.1371/journal.pone.0090461

Figure Lengend Snippet: Oncomine analysis of five mRNA datasets indicate overexpression of Fat1 and of its sheddase ADAM10 in pancreatic cancer. Copyright 2008–11 Compendia Bioscience, Inc. Oncomine (Compendia Bioscience, Ann Arbor, MI) was used for analysis and visualization.

Article Snippet: The ADAM10 specific inhibitor GI254023X , and the broad spectrum metalloproteinase inhibitor Batimastat (Tocris Bioscience, Bristol, UK) were both dissolved in DMSO as stock solutions prior to dilution and use.

Techniques: Over Expression