Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: ADAM9-regulated genes with vascular remodeling functions. ( A ) Ingenuity Pathway Analysis, by comparing gene expression in control versus ADAM9 knockdown cells, showed that the gene network functioned in cell death and survival, cardiovascular disease, and hematological disease. ( B ) Ratio of differential gene expression in microarray analysis (control shGFP Bm7 cells/ ADAM9 knockdown Bm7 cells). ( C – E ) Western blot of ADAM9, ANGPT2, and VEGFA in control and ADAM9 knockdown cells from Bm7 lung cancer cells ( C ), Brmx2 lung cancer cells ( D ), and A549 lung cancer cells ( E ). ( F ) Western blot of ADAM9, VEGFA, and ANGPT2 in Bm7 cells treated with BB94 (a broad spectrum of metalloproteinase inhibitor). ( G ) Western blot of ADAM9, VEGFA, and ANGPT2 in control and ADAM9 knockout Brmx2 cells transiently transfected with plasmids of empty control ( E ), ADAM9 wild-type (WT), or ADAM9 mutant (MT) (E348A). EF1α or GAPDH served as the loading control in ( C – G ). Some cropped blots in ( C – G ) were displayed and the full-length blots were shown in Supplementary Figure .

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

Techniques: Gene Expression, Control, Knockdown, Microarray, Western Blot, Knock-Out, Transfection, Mutagenesis

Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: ADAM9 promotes angiogenesis through VEGFA. ( A ) Concentrations of VEGFA were measured by ELISA in conditioned media of control and ADAM9 knockdown Bm7 cells. ( B , C ) Tube formation assays of HUVECs were performed by treatment with growth media alone (as control) or the conditioned media of indicated cells. Conditioned media of shADAM9-E Bm7 cells (B) and shADAM9-H Bm7 cells ( C ) were used for tube formation assays. ( D ) Tube formation assays were performed by treating HUVECs with the conditioned media of Bm7-shGFP cells in the presence or absence of 5 μg/ml Avastin pre-incubation (white bars), or with the conditioned media of Bm7-shADAM9 cells in the presence or absence of 100 ng/ml VEGF (black bars). Top, photographs of representative experiments for control, GFP knockdown, and ADAM9 knockdown; bottom, quantitative data for tube formation. Bars represent the mean ± SD of three replicate samples from one representative experiment. * P < 0.05, ** P < 0.01.

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

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

Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: ADAM9 disrupts the integrity of brain endothelial cells through ANGPT2 and PLAT. ( A ) VE-cadherin staining of HBMECs treated with concentrated media from the indicated cancer cells. After incubation, cells were stained with anti-VE-cadherin antibody (red) and DAPI (blue). For quantitation, the membrane length stained by anti-VE-cadherin antibody in HBMECs was calculated and normalized to HBMEC nuclear numbers. Anti-ANGPT2 antibody was pre-incubated with concentrated conditioned media before treatment of HBMECs. ( B ) Permeability of HBMECs treated with concentrated conditioned media from the indicated cancer cells. Anti-ANGPT2 antibody was pre-incubated with concentrated conditioned medium before treatment of HBMECs. * P < 0.05. ( C ) VE-cadherin staining of HBMECs treated with the indicated conditioned media or with recombinant tPA protein reveals the integrity of HBMECs.

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

Techniques: Staining, Incubation, Quantitation Assay, Membrane, Permeability, Recombinant

Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: ADAM9 knockout significantly inhibits tumor growth and metastasis in vivo . ( A ) Western blot of ADAM9 knockout in mouse lung cancer TC1 cells. WT, wild-type; #1 and #2 represent two independent stable clones with ADAM9 knockout. ( B ) Wild-type ( n = 7) or ADAM9 knockout ( n = 7) TC1 cells were injected subcutaneously into C57BL/6 mice. The photograph ( top ) shows subcutaneous tumors removed from each group after sacrifice; the graph ( bottom ) shows the quantification of tumor volume at the indicated days after inoculation. ( C ) Western blot of ADAM9, ANGPT2, and VEGFA from subcutaneous (s.c.) tumors of each group. ( D ) Tumor tissues were processed for IHC staining of CD31. The images were magnified at 100x ( top , scale bar 200 μm). CD31 staining was quantified as the percentage of positively stained density ( bottom ). ( E ) WT ( n = 5) and ADAM9 KO ( n = 5) TC1 cells were injected intravenously via the tail vein of mice. The photograph shows metastatic tumors in the lung from each group ( top ) and the weight of whole lung tumors was measured from the two groups ( bottom ). ( F ) Metastasis in the lung from WT or ADAM9 KO TC1 cells injection intravenously appeared as dense tumor clusters (T) following hematoxylin and eosin staining (H&E). ( G ) Lung tumor tissues were processed for IHC staining of ADAM9. Images were scanned with Aperio software; digital resolution is 0.5 μm per pixel. Scale bar is 500 μm. Vessels were highlighted by red broken line (left, upper). The representative images of ADAM9 staining in tumor with or without vessels were showed (left, lower). The scale bar is 100 μm. The intensities of ADAM9, H score, were quantified by Aperio ImageScope software program (right). * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

Techniques: Knock-Out, In Vivo, Western Blot, Clone Assay, Injection, Immunohistochemistry, Staining, Software

Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: Simultaneous high expression of ADAM9 and VEGFA correlates with poor outcome in lung adenocarcinoma patients. ( A ) Kaplan-Meier survival analyses of lung adenocarcinoma patients in each indicated data set, with patients categorized by ADAM9 and VEGFA gene expression. DH, dual high; SH, single high; DL, dual low. High and low are above and below the median, respectively. ( B ) Cox proportional hazards regression analyses of overall survival in each dataset of lung adenocarcinoma patients, grouped by ADAM9 and VEGFA expression. ( C ) Ranking the P -value of ADAM9 plus VEGFA among all P -values of profile genes by combining a fixed predictor with the other randomly selected genes from the same dataset. Fixed predictor: ADAM9 in the GSE8894 dataset and VEGFA in the Shedden dataset. The P -value of ADAM9 high /VEGFA high versus ADAM9 low /VEGFA low is marked by the red line.

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

Techniques: Expressing, Gene Expression

Journal: Scientific Reports

Article Title: ADAM9 promotes lung cancer progression through vascular remodeling by VEGFA, ANGPT2, and PLAT

doi: 10.1038/s41598-017-15159-1

Figure Lengend Snippet: Simultaneous high expression of ADAM9 and ANGPT2 correlates with poor outcome in lung adenocarcinoma patients. ( A ) Kaplan-Meier survival analyses of lung adenocarcinoma patients in each indicated data set, with patients categorized by ADAM9 and ANGPT2 gene expression. Labels are defined as in Fig. . ( B ) Cox proportional hazards regression analyses of overall survival in each dataset of lung adenocarcinoma patients, grouped by ADAM9 and ANGPT2 expression. ( C ) Ranking the P -value of ADAM9 plus ANGPT2 among all P -values of profile genes by combining a fixed predictor with the other randomly selected genes from the same dataset. Fixed predictor: ADAM9 in the GSE8894 and GSE11969 datasets and ANGPT2 in the Shedden dataset. The P -value of ADAM9 high /ANGPT2 high versus ADAM9 low /ANGPT2 low is marked by the red line.

Article Snippet: The antibodies used in the study were against ADAM9 (MAB939, R&D Systems, Minneapolis, MN, USA), mouse ADAM9 (AF949, R&D Systems), ANGPT2 (AF623, R&D Systems), VEGFA (ABS82, Millipore, Billerica, MA, USA), CD31 (ab28364, Abcam), and elongation factor 1 α (EF1α, #05–235, Millipore).

Techniques: Expressing, Gene Expression

Journal: Cancer Science

Article Title: ADAM9 is over‐expressed in human ovarian clear cell carcinomas and suppresses cisplatin‐induced cell death

doi: 10.1111/cas.13469

Figure Lengend Snippet: Immunohistochemical localization of ADAM9 in ovarian serous, endometrioid, mucinous, and clear cell carcinomas and control non‐neoplastic ovarian tissue. Insets show high‐power view of the rectangle areas of ADAM9 immunostaining. NI, negative control with non‐immune IgG. Bar, 50 μm. Note the intense and circumscribed membranous staining with granular intracytoplasmic staining of ADAM9 in the clear cell carcinoma. ADAM, a disintegrin and metalloproteinase

Article Snippet: Paraffin sections of the ovarian tissues were subjected to immunostaining with goat anti‐human ADAM9 polyclonal antibody (AF939; R&D Systems, Minneapolis, MN, USA) or non‐immune goat IgG (AB‐108‐C; R&D Systems) after blocking endogenous peroxidase and antigen retrieval by autoclaving, followed by reactions with biotinylated secondary antibody and 3‐3′‐diaminobenzidine., Sections were counterstained with hematoxylin.

Techniques: Immunohistochemical staining, Control, Immunostaining, Negative Control, Staining

Journal: Cancer Science

Article Title: ADAM9 is over‐expressed in human ovarian clear cell carcinomas and suppresses cisplatin‐induced cell death

doi: 10.1111/cas.13469

Figure Lengend Snippet: Intensity, membrane, and total scores of ADAM9 immunostaining in ovarian serous, endometrioid, mucinous, and clear cell carcinomas. A,C,E, Distribution of the intensity (0, 1+ and 2+), membrane (0 to 3+) and total scores (0 to 5+) of ADAM9 immunostaining. B,D,F, Distribution of the intensity, membrane and total scores of ADAM9 immunostaining in non‐clear cell and clear cell carcinoma groups. * P < .05; *** P < .001. ADAM, a disintegrin and metalloproteinase

Article Snippet: Paraffin sections of the ovarian tissues were subjected to immunostaining with goat anti‐human ADAM9 polyclonal antibody (AF939; R&D Systems, Minneapolis, MN, USA) or non‐immune goat IgG (AB‐108‐C; R&D Systems) after blocking endogenous peroxidase and antigen retrieval by autoclaving, followed by reactions with biotinylated secondary antibody and 3‐3′‐diaminobenzidine., Sections were counterstained with hematoxylin.

Techniques: Membrane, Immunostaining

Journal: Cancer Science

Article Title: ADAM9 is over‐expressed in human ovarian clear cell carcinomas and suppresses cisplatin‐induced cell death

doi: 10.1111/cas.13469

Figure Lengend Snippet: Knock‐down of ADAM9m in human ovarian clear cell carcinoma cell lines (RMG‐I and TOV21G) and its effect on cell migration and EGFR activation. A, mRNA expression of ADAM9m, ADAM9s, ADAM10 and ADAM17 in cells transfected with shRNAs (sh1 and sh2) for ADAM9 or non‐targeting shRNA (Mock) vectors by RT‐PCR. β‐actin, a loading control. B, Protein expression of ADAM9m in cells transfected with sh1, sh2 or Mock vectors by immunoblotting. C, Migration assay of cells transfected with sh1, sh2 or Mock vectors. Migration activity was measured at 24 h after scratch‐wounding. D, Activation of EGFR of cells transfected with sh1, sh2 or Mock vectors. Expression of phosphorylated EGFR (pEGFR) and EGFR was examined by immunoblotting. All assays were carried out in triplicate, and repeated 3 times. Bars, mean value ± SD. ** P < .01; *** P < .001; NS, not significant. ADAM, a disintegrin and metalloproteinase

Article Snippet: Paraffin sections of the ovarian tissues were subjected to immunostaining with goat anti‐human ADAM9 polyclonal antibody (AF939; R&D Systems, Minneapolis, MN, USA) or non‐immune goat IgG (AB‐108‐C; R&D Systems) after blocking endogenous peroxidase and antigen retrieval by autoclaving, followed by reactions with biotinylated secondary antibody and 3‐3′‐diaminobenzidine., Sections were counterstained with hematoxylin.

Techniques: Knockdown, Migration, Activation Assay, Expressing, Transfection, shRNA, Reverse Transcription Polymerase Chain Reaction, Control, Western Blot, Activity Assay

Journal: Cancer Science

Article Title: ADAM9 is over‐expressed in human ovarian clear cell carcinomas and suppresses cisplatin‐induced cell death

doi: 10.1111/cas.13469

Figure Lengend Snippet: Effects of ADAM9m knock‐down, inhibition of ADAM9m activity and ADAM9m over‐expression on ovarian clear cell carcinoma cells after treatment with cisplatin. A, ADAM9m knock‐down effect on cell viability after treatment with cisplatin. Viability of RMG‐I and TOV21G cells treated without (CDDP−) or with cisplatin (CDDP+) was evaluated by MTT assay. B, Effect of ADAM9m knock‐down on cisplatin‐induced apoptotic cell death. Apoptotic cells showing positive reaction of propidium iodide (PI) and Annexin‐V in Mock and ADAM9m shRNA transfectants (sh1 and sh2) were monitored by flow cytometric assay. C, Effect of neutralizing anti‐ADAM9m antibody on cisplatin‐treated cells. Viability of the cells treated with LD50 concentration of cisplatin in the presence of non‐immune IgG (NI) or anti‐ADAM9m antibody (αADAM9) was evaluated by MTT assay. Cont, the cells treated with LD50 cisplatin alone. D, ADAM9m over‐expression effect on cisplatin‐treated cells. mRNA expression and cell viability of TOV21G cells transfected with ADAM9m expression vectors (A9m‐t) or mock vectors (Mock) were examined by RT‐PCR and MTT assay, respectively. All assays were carried out in triplicate, and repeated 3 times. Bars, mean value ± SD. NS, not significant; * P < .05; ** P < .01; *** P < .001. ADAM, a disintegrin and metalloproteinase

Article Snippet: Paraffin sections of the ovarian tissues were subjected to immunostaining with goat anti‐human ADAM9 polyclonal antibody (AF939; R&D Systems, Minneapolis, MN, USA) or non‐immune goat IgG (AB‐108‐C; R&D Systems) after blocking endogenous peroxidase and antigen retrieval by autoclaving, followed by reactions with biotinylated secondary antibody and 3‐3′‐diaminobenzidine., Sections were counterstained with hematoxylin.

Techniques: Knockdown, Inhibition, Activity Assay, Over Expression, MTT Assay, shRNA, Flow Cytometry, Concentration Assay, Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction

Journal:

Article Title: Transmembrane collagen XVII, an epithelial adhesion protein, is shed from the cell surface by ADAMs

doi: 10.1093/emboj/cdf532

Figure Lengend Snippet: Fig. 4. Expression of ADAMs in human keratinocytes. (A) Immuno fluorescence staining of collagen XVII and ADAMs in the skin. Collagen XVII antibody NC16A produced a linear signal at the basement membrane zone (a). TACE (b) and ADAM-9 (c) immunoreactivity was stronger in the lower epidermis, in the neighborhood of collagen XVII, whereas ADAM-10 immunoreactivity (d) was distributed throughout the entire epidermis. Scale bar: 250 µm. (B) Immunoblot of keratinocyte extracts with antibodies to the ectodomains of TACE and ADAM-9, and the endodomain of ADAM-10. TACE and ADAM-9 antibodies recognized two distinct protein bands of ∼120 and 85 kDa, and 110 and 79 kDa, respectively, representing the proform and the active form of the enzymes. ADAM-10 antibody recognized one band of ∼60 kDa, corresponding to the active form. (C) Activation of TACE in keratinocytes was inhibited by a furin inhibitor, but not by hydroxamates. Keratinocytes were cultured for 24 h in serum-free medium with or without 50 µM furin inhibitor decanoyl-RVKR-chloromethyl ketone or 25 µM hydroxamate BB 3103. Keratinocyte lysates were immunoblotted with TACE antibodies. The furin inhibitor, but not BB 3103, prevented the conversion of pro-TACE to active TACE. Molecular weight markers are shown on the left.

Article Snippet: As first antibodies, collagen XVII antibody NC16A, TACE antibodies against the extracellular pro- and metalloproteinase domains (Santa Cruz Biotechnology) and the intracellular C-terminus (Chemicon International, Hofheim, Germany), and antibodies to ADAM-10 endodomain (Chemicon) and ADAM-9 ectodomain (R&D Systems, Wiesbaden, Germany) were used.

Techniques: Expressing, Fluorescence, Staining, Produced, Membrane, Western Blot, Activation Assay, Cell Culture, Molecular Weight

Journal:

Article Title: Transmembrane collagen XVII, an epithelial adhesion protein, is shed from the cell surface by ADAMs

doi: 10.1093/emboj/cdf532

Figure Lengend Snippet: Fig. 5. Transient transfections of HaCaT cells with cDNAs for TACE, ADAM-9 and ADAM-10. (A) Immunofluorescence staining of transfected cells. Cells transfected with empty vector (control) remained negative, but ADAM-transfected cells showed strong positive signals. Scale bar: 100 µm. (B) HaCaT cells were transfected with different concentrations of cDNA for TACE (upper panel), ADAM-10 (middle panel) and ADAM-9 (lower panel) and shedding of collagen XVII was assessed with immunoblotting with a mixture of antibodies Endo-2, NC16A and Ecto-1. Densitometric analysis of the signals (expressed as a percentage of the control ± SD; n = 3) showed a dose-dependent increase of ectodomain shedding, with a concomitant decrease of full-length collagen XVII.

Article Snippet: As first antibodies, collagen XVII antibody NC16A, TACE antibodies against the extracellular pro- and metalloproteinase domains (Santa Cruz Biotechnology) and the intracellular C-terminus (Chemicon International, Hofheim, Germany), and antibodies to ADAM-10 endodomain (Chemicon) and ADAM-9 ectodomain (R&D Systems, Wiesbaden, Germany) were used.

Techniques: Transfection, Immunofluorescence, Staining, Plasmid Preparation, Western Blot

Journal:

Article Title: Transmembrane collagen XVII, an epithelial adhesion protein, is shed from the cell surface by ADAMs

doi: 10.1093/emboj/cdf532

Figure Lengend Snippet: Fig. 8. Increased collagen XVII shedding correlates with decreased cell motility. In wound closure assays cells migrate from the edges of a ‘scrape wound’ in the monolayer (interrupted lines) to cover the denuded space (between the lines). Scrape wounds were made in serum-free cultures, and after washing the cells were allowed to re-epithelialize for 22 h at 37°C. Micrographs of non-fixed cell layers at 0, 11, 16 or 22 h after wounding are shown. Scale bars: 200 µm. (A) Transfection of HaCaT cells with ADAM cDNA negatively regulated cell motility. Transfections were with 15 µg of empty vector (control), 15 µg of TACE, 15 µg of ADAM-10, or 15 µg of ADAM-9 cDNA. (B) Addition of purified collagen XVII ectodomain to normal human keratinocytes inhibited cell motility. After wounding, 1 nM collagen XVII ectodomain was added to the culture, and the cells were permitted to migrate into the denuded space for 16 h. Ectodomain-treated cells did not cover the denuded area as efficiently as control cells.

Article Snippet: As first antibodies, collagen XVII antibody NC16A, TACE antibodies against the extracellular pro- and metalloproteinase domains (Santa Cruz Biotechnology) and the intracellular C-terminus (Chemicon International, Hofheim, Germany), and antibodies to ADAM-10 endodomain (Chemicon) and ADAM-9 ectodomain (R&D Systems, Wiesbaden, Germany) were used.

Techniques: Transfection, Plasmid Preparation, Purification

Journal: bioRxiv

Article Title: Membrane-type 1 matrix metalloproteinase (MMP-14) modulates tissue homeostasis by a non-proteolytic mechanism

doi: 10.1101/631739

Figure Lengend Snippet: A . Western blotting analysis of MT1-MMP, CD44 and DDR1 expression and degradation in abdominal fat, joint cartilage and bone (femur) from 3-month old Mmp14 wt/wt (W) and Mmp14 Y573D/Y573D (Y) male mice. Tissues from three littermates per genotype were pooled for protein extraction. Lamin B is shown as a loading control. A representative result of multiple experiments is shown. B. Western blotting analysis of CD44, DDR1 and ADAM9 in cell extracts of primary fibroblasts from Mmp14 wt/wt (wt/wt) and Mmp14 Y573D/Y573D (YD/YD) mice grown in the presence (+) or absence (-) of GM6001 (50 μM) for 24 h. α-tubulin is shown as a loading control. A representative result of multiple experiments is shown. Bottom panel: gelatin zymography analysis of proMMP-2 activation is shown as a control for GM6001 inhibition of MT1-MMP. The cells were grown for 16 h in serum-free medium supplemented with human recombinant proMMP-2 without (-) or with (+) GM6001 (50 μM), and the conditioned medium was analyzed by gelatin zymography. See also .

Article Snippet: MT1-MMP antibody : Anti-MMP14 antibody [EP1264Y] rabbit mAb (Abcam, cat. # ab51074; Cambridge, MA), dilution: 1:5,000; phospho-ERK1/2 antibody : Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (197G2) Rabbit mAb (Cell Signaling Technology, cat. # 4377; Danvers, MA), dilution: 1:1,000; phospho-Akt antibody : Phospho-Akt (Ser473) (D9E) XP ® Rabbit mAb (Cell Signaling Technology, cat. # 4060), dilution: 1:2,000; phospho-FAK antibodies : Phospho-FAK (Tyr861) Recombinant Rabbit Monoclonal Antibody (26H16L4) (Invitrogen, cat. # 700154), concentration: 2 μg/ml; or Phospho-FAK (Tyr397) Polyclonal Antibody (Life technology, cat. # 44-624G), dilution: 1,1000; total ERK1/2 antibody : p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb (Cell Signaling Technology, cat. # 4695), dilution: 1:1,000; total Akt antibody : Akt (pan) (C67E7) Rabbit mAb (Cell Signaling Technologies, cat. # 4691), dilution: 1:1,000; total FAK antibody : FAK Antibody C20 (Santa Cruz Biotechnology, cat. # sc-558), dilution: 1:200; CD44 antibody : mouse monoclonal antibody to mouse CD44 (DSHB; cat. # 5D2-27-s), dilution: 0.2 - 0.5 μg/ml; ADAM9 antibody : mouse anti-ADAM-9 monoclonal antibody (R&D Systems; cat. # MAB939); dilution: 1 μg /ml; DDR1 antibody : rabbit anti-DDR1 monoclonal antibody D1G6 (Cell Signaling Technology; cat. # 5583, dilution: 1:1,000; or Phospho-DDR1 (Tyr792) Antibody (Cell Signaling Technology; cat. #11994), dilution: 1:1,000; α-tubulin antibody : rabbit anti- tubulin monoclonal antibody 11H10 (Cell Signaling Technology; cat. #2125), dilution: 1:1,000; β-tubulin antibody : β-Tubulin Antibody #2146 (Cell Signaling Technology; cat. #21465), dilution: 1:1,000; lamin B antibody : goat anti-lamin B antibody B-20 (Santa Cruz; cat. # sc-6217); dilution: 1:1,000; rabbit IgG antibody : anti-rabbit IgG, HRP-linked antibody (Jackson ImmunoResearch; cat. # AB_10015289), dilution 1:10,000, or (Cell Signaling Technologies, cat. # 7074), dilution: 1:10,000; mouse IgG antibody: anti-mouse HRP-linked antibody (Jackson ImmunoResearch; cat. # AB_2307391), dilution 1:10,000.

Techniques: Western Blot, Expressing, Protein Extraction, Control, Zymography, Activation Assay, Inhibition, Recombinant