Journal: Biomaterials and Biosystems

Article Title: Engineered extracellular vesicles antagonize SARS-CoV-2 infection by inhibiting mTOR signaling

doi: 10.1016/j.bbiosy.2022.100042

Figure Lengend Snippet: Gene expression assays.

Article Snippet: adam10 , Human , , Hs00153853_m1.

Techniques: Gene Expression

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: TSPAN15, a new ADAM10 interaction partner. a-I, II: Split-ubiquitin yeast two-hybrid system (modified from [50]). ADAM10 C-terminally fused to the C-terminal part of ubiquitin (Cub) and an artificial transcription factor (LexA-VP16) coexpressed in yeast together with a N-terminal NubG-tagged murine brain library. The close proximity between the ADAM10 bait protein and an interaction partner leads to the reconstitution of “split ubiquitin” to ubiquitin, which is recognized by cellular ubiquitin proteases, which in turn release the artificial transcription factor from the membrane. This enables yeast to grow on selective media [without leucine (-leu), tryptophane (-trp), histidine (-his)] plates. C-terminal part of ubiquitin (Cub, C), N-terminal part of ubiquitin (NubG, N), LexA (L). b ADAM10 bait protein coexpressed with the identified TSPAN15 prey protein and controls in NMY51 yeast. Transfection of both bait and prey protein is verified on Leu/Trp-lacking selective media plates, and interaction of ADAM10 and TSPAN15 is monitored under selective pressure on Leu/Trp/His-lacking media plates in comparison to controls (“+” and “−”). c-I: Mammalian expression constructs of murine ADAM10 and murine TSPAN15-myc were transiently coexpressed in HeLa cells. TSPAN15-myc (35–37 kDa) was precipitated using an anti-myc antibody, and coprecipitation of ADAM10 was detected with an anti-ADAM10 antibody. II: ADAM10 and TSPAN15-myc were transiently coexpressed in HeLa cells, and ADAM10 was precipitated using an anti-ADAM10 antibody. Coprecipitation was analyzed by Western blot using an anti-myc antibody. Single transfections of TSPAN15-myc and ADAM10 served as specificity controls for the antibodies used for immunoprecipitation. III: Quantification of band intensities of pro and mature form of ADAM10 in lysates and CoIP fractions. Ratios of band intensities of pro/(pro + mature) forms of ADAM10 were calculated (%, n = 5). Student’s t test was performed (***p < 0.005). [pro(p) ADAM10, 95 kDa, mature(m) ADAM10, 75 kDa]. Abbreviations: untransfected (Ø), vector (mock) transfected (V), murine ADAM10 (A10), murine TSPAN15-myc (T15), asterisk marks immunoglobulin signals

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Ubiquitin Proteomics, Modification, Membrane, Transfection, Comparison, Expressing, Construct, Western Blot, Immunoprecipitation, Plasmid Preparation

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: TSPAN15-myc influences ADAM10 maturation. a Murine ADAM10 transiently coexpressed with either EGFP or murine TSPAN15-myc in Cos7 cells. After immunoblotting ADAM10 was detected using an ADAM10 specific C-terminal antibody (B42.1) and TSPAN15-myc was detected using an anti-myc antibody. b TSPAN15-myc or EGFP was transiently expressed in N2A cells. After lysis proteins were immunoblotted, and endogenous ADAM10 and TSPAN15-myc were detected. c Murine ADAM17 or murine ADAM10 was transiently expressed either with EGFP or TSPAN15-myc in HeLa cells. Presence of ADAM17 [pro(p) ADAM17, closed arrowhead, 120 kDa; putative mature ADAM17, open arrowhead] was analyzed using a C-terminal-specific antibody, and TSPAN15-myc and ADAM10 expressions were monitored as mentioned above. Actin served as protein loading control. Asterisks mark unspecific antibody binding

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Western Blot, Lysis, Control, Binding Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: TSPAN15-myc influences ADAM10 localization. a-I, II: Murine ADAM10 and ADAM10/TSPAN15-myc expressed in Cos7 cells. III, IV: Murine ADAM17 and ADAM17/TSPAN15-myc expressed in Cos7 cells. Confocal immunofluorescence pictures were taken using an anti-KDEL antibody, an anti-ADAM10 antibody, and an anti-ADAM17 antibody, respectively. Scale bar 100 μm. V, VI: SHSY cells were transiently transfected with a C-terminal EGFP-tagged variant of human TSPAN15. Endogenous ADAM10 was stained using an anti-ADAM10 antibody (11G2), and an anti-PDI antibody was used as ER marker. Asterisks mark untransfected cells. Scale bar 10 μm. b N2A cells were biotinylated after transfection with either EGFP or murine TSPAN15-myc. Following cell lysis total protein samples were taken, and after precipitation of biotin-labeled proteins immunoblotting of total lysates and bound fractions was performed. TSPAN15-myc and ADAM10 were detected. The detection of the transferrin receptor (TFR) was included as a control for biotinylated surface proteins, and antibodies against the intracellular glycerin-aldehyd-3-phosphate dehydrogenase (GAPDH) were used as a negative control. c-I: N2A cells were transfected with ADAM10, TSPAN15-myc and ADAM10/TSPAN15-myc; ADAM10 cell surface expression was determined through FACS analysis using an N-terminal anti-ADAM10 antibody. RFUs were determined. Statistical significance was determined using Student’s t test; values are provided as highly significant (**p < 0.01). Abbreviation: relative fluorescence units (RFU). II: Representative overlay of the FACS analysis performed

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Immunofluorescence, Transfection, Variant Assay, Staining, Marker, Lysis, Labeling, Western Blot, Control, Negative Control, Expressing, Fluorescence

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: Activity analysis of ADAM10. a-I: murine (m)ADAM10 expressed or coexpressed with TSPAN15-myc in Cos7 cells. Activity of ADAM10 was studied through the analysis of ADAM10-specific shedding events using immunoblot detection of N-cadherin [full-length (Fl) N-cadherin (130 kDa), C-terminal fragment (CTF) N-cadherin (37 kDa)] applying a C-terminal-specific N-cadherin antibody. II: EGFP and TSPAN15-myc were transiently expressed in Cos7 cells, and the processing of N-cadherin was analyzed by Western blot. b N2A cells were transfected with pcDNA3.1 or TSPAN15-myc and APP/pcDNA3.1 or APP/TSPAN15-myc. Processing of APP was analyzed using a C-terminal-specific anti-APP antibody. Actin served as protein loading control. c Cell culture supernatants of N2A cells transfected with EGFP or TSPAN15-myc and sAPPalpha content were determined using sandwich ELISAs. d HEK293 cells were transfected with human TSPAN15-EGFP or vector control, and endogenous APP processing was analyzed. APP (FL) APP full-length protein; APP (CTF) APP C-terminal C83 fragment; overex overexposed image of the upper panel. ADAM10, TSPAN15 and actin expression analysis was included

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Activity Assay, Western Blot, Transfection, Control, Cell Culture, Plasmid Preparation, Expressing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: Knockdown of TSPAN15. a Knockdown of TSPAN15 in N2A cells was verified through qRT-PCR analysis. b N2A cells were transfected with siRNA against TSPAN15 and control siRNA. After lysis and immunoblot analysis, ADAM10 was detected. Actin served as protein-loading control. c-I: Surface expression of ADAM10 was analyzed through FACS analysis using a N-terminal anti ADAM10 antibody. Remaining surface expression of ADAM10 was determined (%). II: Representative overlay of FACS analysis of ADAM10 surface protein after TSPAN15 knockdown

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Knockdown, Quantitative RT-PCR, Transfection, Control, Lysis, Western Blot, Expressing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: Early interaction of TSPAN15 with ADAM10. a Murine ADAM10 was either expressed with TSPAN15-myc or TSPAN15-ER-myc. Cells were lysed, and myc-tagged proteins were precipitated using an anti-myc antibody. After immunoblotting coprecipitated ADAM10 was detected using a C-terminal-specific antibody (B42.1). b-I, II: Cos7 cells were transfected with TSPAN15-ER-myc or TSPAN15-ER-myc/ADAM10. Confocal immunofluorescence pictures were taken using an anti-myc antibody, an anti-KDEL antibody and an N-terminal-specific ADAM10 antibody and adequate secondary antibody pairs. Scale bar 100 μm. c N2A cells were transfected with EGFP or TSPAN15-myc, pulsed for 1 h with 35S methionine/cysteine and chased for 0, 2, 6, 18 and 30 h. After cell lysis equal amounts of protein were used, and ADAM10 was precipitated using a C-terminal-specific anti-ADAM10 antibody, subjected to SDS-PAGE and analyzed by fluorographics. TSPAN15-myc expression was analyzed by Western blot, and actin served as protein-loading control for the lysates

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Western Blot, Transfection, Immunofluorescence, Lysis, SDS Page, Expressing, Control

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Tetraspanin15 regulates cellular trafficking and activity of the ectodomain sheddase ADAM10

doi: 10.1007/s00018-012-0960-2

Figure Lengend Snippet: Model of TSPAN15 function. (1) ADAM10 is synthesized as an inactive precursor (pro-ADAM10) in the ER, and TSPAN15 accelerates its ER exit. (2) After ER exit the immature ADAM10 is activated through removal of the inhibitory prodomain by furin or proprotein convertase (PC7), and is then transported to the plasma membrane together with TSPAN15. (3) At the plasma membrane TSPAN15 facilitates the integration of ADAM10 in the tetraspanin web. (4, 5) ADAM10 is stabilized, and the web composition enables ADAM10 to get access to its substrates and subsequent cleavage events in the juxtamembrane regions of the substrates take place. (6) Afterwards remaining membrane-bound fragments are cleaved through ripping proteases (γ-secretase complex/SPPLs) in the transmembrane regions, thereby liberating soluble intracellular domains (ICD), which might have signaling functions

Article Snippet: ADAM10 (Mm00545742_m1, Applied Biosystems, Foster City, CA, USA), TSPAN15 (Mm01150417_m1, Applied Biosystems) and GAPDH (Mm99999915_g1, Applied Biosystems) expression were determined by real-time PCR analysis of 0.5 μl cDNA on a 7900HT Fast Real-time PCR System (Applied Biosystems) in 10-μl reaction volume in 384-well plates in duplicate.

Techniques: Synthesized, Clinical Proteomics, Membrane

Journal: Oncotarget

Article Title: Serological immune response against ADAM10 pro-domain is associated with favourable prognosis in stage III colorectal cancer patients

doi: 10.18632/oncotarget.11181

Figure Lengend Snippet: A. Serological reactivity of Crc patient pool of sera and control pool of sera against the biotinylated protein spot (Av-HRP) that, from the preparative 2DE gel (Coomassie blue), yielded the MS identification of ADAM10; the identity was confirmed by reactivity of an anti-ADAM10 Ab with the same spot. B. Surface expression of ADAM10 in the LS180 Crc cell line. Anti-ADAM10 immunofluorescence reactivity is present on both permeabilized and non-permeabilized cells. Anti-HLA-class I and anti- ß-actin reactivities were used as controls for surface and intracellular expressed proteins, respectively. C. Reactivity of Crc patients and control subjects (Cn) sera against purified ADAM10; anti-ADAM10 Ab reactivity was used for signal normalization. D. - E. Quantitative analysis of serological reactivity reported as normalized OD (mean +/− SEM of 3 experiments in duplicate). D. Testing cohorts Crc1, n = 57; Cn1, n = 39; Crc1-stage I n = 8, stage II n = 17, stage III n = 26, stage IV n = 6. E. Validation cohorts Crc2, n = 49; Cn2, n = 52; Crc2-stage I n = 13, stage II n = 13, stage III n = 13; stage IV n = 10. Statistical analysis was performed by either student-t (S-t) test or Mann-Whitney (M-W) test and non parametric analysis of variance by Kruskal-Wallis (K-W). (*** = p < 0.0001; ** = p < 0.01).

Article Snippet: Protein identity was confirmed on LS180-biotinylated material by 2DE-WB using an anti-ADAM10 Ab (AB936, R&D Systems).

Techniques: Control, Expressing, Immunofluorescence, Purification, MANN-WHITNEY

Journal: The Journal of Biological Chemistry

Article Title: ADAM17 mediates ectodomain shedding of the soluble VLDL receptor fragment in the retinal epithelium

doi: 10.1016/j.jbc.2021.101185

Figure Lengend Snippet: Effect of various proteinase inhibitors on VLDLR shedding. Primary human RPE cells were transduced with adenovirus (MOI = 25) expressing VLDLRII (Ad-VLDLRII) or with Ad-RFP as control for 24 h. Then, culture media were replaced by serum-free DMEM containing various proteinase inhibitors (10 μM) for another 24 h. After that, the culture media and cell lysates were harvested for Western blot analysis. A , representative images of Western blotting for soluble ectodomain of VLDLR (sVLDLR) in culture media and the full-length VLDLR in cell lysates. PMA (a PKC activator) and proteinase inhibitors including TAPI-1 (tumor necrosis factor-α proteinase inhibitor), GM6001 (pan-MMP inhibitor), GI254023x (ADAM10 inhibitor), and GW280264x (ADAM17 inhibitor) were used. The same volume of DMSO (vehicle) was used as control. B , densitometry analysis of sVLDLR in culture media normalized by the full-length VLDLR levels in cell lysates in ( A ) (n = 3). Cells with Ad-VLDLRII transduction and DMSO treatment were used as control. C , representative images of Western blotting for sVLDLR and VLDLR from primary human RPE cells treated with indicated doses of GW280264x or DMSO as control for 24 h. D , densitometry of sVLDLR in culture media normalized by VLDLR in cell lysates in ( C ) (n = 3). E , ADAM17 activity in GW280264x-treated cells were measured using an ADAM17 activity assay kit (n = 3). Data were presented as mean ± SD. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001. ADAM17, a disintegrin and metalloprotease 17; DMEM, Dulbecco's modified Eagle's medium; DMSO, dimethyl sulfoxide; MOI, multiplicity of infection; PKC, protein kinase C; PMA, phorbol 12-myristate 13-acetate; RFP, red fluorescent protein; RPE, retinal pigment epithelium; VLDLR, very low-density lipoprotein receptor; VLDLRII, VLDLR variant II.

Article Snippet: Expression vectors for human ADAM10 (Addgene; plasmid #31717) and human ADAM17 (Addgene; plasmid #31713) were gifts from Dr Rik Derynck ( ).

Techniques: Transduction, Expressing, Control, Western Blot, Activity Assay, Modification, Infection, Variant Assay

Journal: The Journal of Biological Chemistry

Article Title: ADAM17 mediates ectodomain shedding of the soluble VLDL receptor fragment in the retinal epithelium

doi: 10.1016/j.jbc.2021.101185

Figure Lengend Snippet: Effect of siRNA knockdown and plasmid overexpression of ADAM10 and ADAM17 on VLDLR shedding. Primary human RPE cells were transiently transfected with indicated siRNA (50 nM) or plasmid (1 μg/ml) for 24 h. Cells were subsequently infected with Ad-VLDLRII (MOI = 25) for 24 h. Then, culture media were replaced with serum-free DMEM for another 24 h. Finally, culture media and cell lysates were collected for Western blot analysis. A , representative images of Western blotting for sVLDLR in culture media and full-length VLDLR, ADAM10, and ADAM17 in cell lysates. The RPE cells were transfected with the siRNA for ADAM10, ADAM17, or control siRNA. B , quantification of densitometry of sVLDLR in culture media normalized by VLDLR in cell lysates in ( A ) (n = 3). C , representative images of Western blotting for sVLDLR in culture media, and VLDLR, ADAM10, and ADAM17 in cells transfected with plasmid overexpressing human ADAM10, human ADAM17, or RFP as control. D , protein levels of sVLDLR in the media in ( C ) were quantified and normalized by the full-length VLDLR in cell lysates (n = 3). E , representative images of Western blotting of sVLDLR in culture media, VLDLR, ADAM10, and ADAM17 in human primary RPE cells, which were transfected with siRNA knocking down human ADAM17 (siADAM17) or plasmid overexpressing human ADAM17 (pADAM17). F , quantification of densitometry of sVLDLR in culture media normalized by VLDLR in cell lysates in ( E ) (n = 3). In A , C , and E , p and m indicated precursor and mature forms of ADAM10 and ADAM17, respectively. Data were presented as mean ± SD. ∗∗ p < 0.01. ADAM10, a disintegrin and metalloprotease 10; ADAM17, a disintegrin and metalloprotease 17; DMEM, Dulbecco's modified Eagle's medium; MOI, multiplicity of infection; RFP, red fluorescent protein; RPE, retinal pigment epithelium; sVLDLR, soluble ectodomain of VLDLR; VLDLR, very low-density lipoprotein receptor; VLDLRII, VLDLR variant II.

Article Snippet: Expression vectors for human ADAM10 (Addgene; plasmid #31717) and human ADAM17 (Addgene; plasmid #31713) were gifts from Dr Rik Derynck ( ).

Techniques: Knockdown, Plasmid Preparation, Over Expression, Transfection, Infection, Western Blot, Control, Modification, Variant Assay

Journal: The Journal of Biological Chemistry

Article Title: ADAM17 mediates ectodomain shedding of the soluble VLDL receptor fragment in the retinal epithelium

doi: 10.1016/j.jbc.2021.101185

Figure Lengend Snippet: CRISPR/Cas9-mediated ADAM17 KO blocked VLDLR shedding and its effect on Wnt signaling. A , representative images of Western blotting of sVLDLR in the media, and VLDLR, ADAM10, and ADAM17 in cell lysates of Ad-VLDLRII-infected WT and ADAM17 KO ARPE-19 cells. WT cells and ADAM17 KO cells were transduced with Ad-VLDLRII (MOI = 25) for 24 h. Then, culture media were replaced with serum-free DMEM for another 24 h. After that, the conditioned media and cell lysates were harvested for Western blot analysis. B , quantification of densitometry of sVLDLR in culture media normalized by VLDLR in cell lysates in ( A ) (n = 3). C , representative images of Western blotting of sVLDLR in culture media, and VLDLR, ADAM10, and ADAM17 in cell lysates in ADAM17 KO cells that were transfected with plasmid overexpressing ADAM17 (pADAM17) or RFP as control. D , quantification of densitometry of sVLDLR in culture media normalized by VLDLR in cell lysates in ( C ) (n = 3). In A and C , p and m indicated precursor and mature forms of ADAM10 and ADAM17, respectively. Data were presented as mean ± SD. ∗∗∗ p < 0.001. ADAM10, a disintegrin and metalloprotease 10; ADAM17, a disintegrin and metalloprotease 17; DMEM, Dulbecco's modified Eagle's medium; MOI, multiplicity of infection; RFP, red fluorescent protein; sVLDLR, soluble ectodomain of VLDLR; VLDLR, very low-density lipoprotein receptor.

Article Snippet: Expression vectors for human ADAM10 (Addgene; plasmid #31717) and human ADAM17 (Addgene; plasmid #31713) were gifts from Dr Rik Derynck ( ).

Techniques: CRISPR, Western Blot, Infection, Transduction, Transfection, Plasmid Preparation, Control, Modification

Journal: The Journal of Biological Chemistry

Article Title: ADAM17 mediates ectodomain shedding of the soluble VLDL receptor fragment in the retinal epithelium

doi: 10.1016/j.jbc.2021.101185

Figure Lengend Snippet: ADAM17 cleaved VLDLR in a cis manner, which was blocked by O-glycosylation. Primary human RPE cells were transfected with a plasmid expressing human ADAM17 (pADAM17) or RFP as control for 24 h. Cells were then infected with Ad-VLDLRII (MOI = 25) or RFP as control for another 24 h. After that, the cells were equally split into new 10-cm dishes for comparing cis and trans manner. A , representative images of Western blotting of sVLDLR in culture media, VLDLR, ADAM10, and ADAM17 in ARPE-19 cell lysates. Cis indicated that ADAM17 and VLDLR were overexpressed in same cells. Trans indicated that ADAM17 and VLDLR were overexpressed in different cells. B , densitometry analysis of sVLDLR in culture media normalized by the full-length VLDLR in cell lysates in ( A ) (n = 3). C , representative images of Western blotting of sVLDLR in culture media, and VLDLR and ADAM17 in O-glycosylation–deficient cells (ldlD). ldlD cells were transfected with the ADAM17 expression plasmid (pADAM17) or RFP as control for 24 h. Then, cells were transduced with adenovirus overexpressing VLDLRI, the full-length VLDLR with O-glycosylation domain (MOI = 25) or RFP as control for another 24 h. After that, culture media were replaced with a serum-free media with or without 10 μM d -(+)-galactose and 100 μM N -acetyl- d -galactosamine for 24 h. D , densitometry analysis of sVLDLR in culture media normalized by VLDLR in cell lysates in ( C ) (n = 3). Gal: d -(+)-galactose and N -acetyl- d -galactosamine. In A and C , p and m indicated precursor and mature forms of ADAM10 and ADAM17, respectively. Data were presented as mean ± SD. ∗ p < 0.05, ∗∗∗ p < 0.001. ADAM10, a disintegrin and metalloprotease 10; ADAM17, a disintegrin and metalloprotease 17; MOI, multiplicity of infection; RFP, red fluorescent protein; RPE, retinal pigment epithelium; VLDLR, very low-density lipoprotein receptor; VLDLRI, VLDLR variant I; VLDLRII, VLDLR variant II.

Article Snippet: Expression vectors for human ADAM10 (Addgene; plasmid #31717) and human ADAM17 (Addgene; plasmid #31713) were gifts from Dr Rik Derynck ( ).

Techniques: Glycoproteomics, Transfection, Plasmid Preparation, Expressing, Control, Infection, Western Blot, Transduction, Variant Assay