title: Role of ADAM10 and ADAM17 in CD16b shedding mediated by different stimulators.
abstract: OBJECTIVE
  To investigate the main proteinases responsible for CD16b shedding under different stimulators.
  
  
  METHODS
  HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then overexpressed with a disintegrin and metalloproteinase 10 (ADAM10) or ADAM17, suppressed with short hairpin RNA of ADAM10 or ADAM17, and reconstituted with ADAM10 or ADAM17, respectively. After each treatment, the cell line was stimulated with ionomycin or phorbol 12-myristate- 13-acetate (PMA) for 12 hours. The soluble CD16b released from cell membrane was detected by immunoprecipition and immunoblot. Quantitation was then implemented to compare the amount of soluble CD16b in cell supernatant after stimulation.
  
  
  RESULTS
  HEK293 cell line stably expressing CD16b was successfully established. When CD16b expressing cell line was overexpressed with ADAM10, shedding of CD16b was increased after stimulation with ionomycin but not PMA; when the cell line overexpressed with ADAM17, shedding of CD16b was increased after stimulation with PMA but not ionomycin. Similarly, when ADAM10 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with ionomycin; when ADAM17 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with PMA. The shedding of CD16b was increased again when CD16b expressing cell line was reconstituted with ADAM10 and stimulated by ionomycin or reconstituted with ADAM17 and stimulated by PMA.
  
  
  CONCLUSIONS
  Both ADAM10 and ADAM17 could shed CD16b, but they possess differed preferences. ADAM10 is the main sheddase under stimulation of ionomycin, while ADAM17 is the main sheddase under stimulation of PMA.
authors: Department of Immunology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China.
authors: Sha
authors: S
authors: Guo
authors: Min
authors: M
authors: Peng
authors: Qing
authors: Q
authors: Zhao
authors: Wei
authors: W
authors: Zhang
mesh_terms: D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000072197:ADAM10 Protein; D000072198:ADAM17 Protein; D053829:Amyloid Precursor Protein Secretases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D061207:Calcium Ionophores / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002478:Cells, Cultured; D004353:Drug Evaluation, Preclinical; D058851:GPI-Linked Proteins / Q000378:metabolism; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D059748:Proteolysis / Q000187:drug effects; D017452:Receptors, IgG / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
chemical_list: D061207:Calcium Ionophores; D002273:Carcinogens; C428200:FCGR3B protein, human; D058851:GPI-Linked Proteins; D008565:Membrane Proteins; D017452:Receptors, IgG; D015759:Ionomycin; D053829:Amyloid Precursor Protein Secretases; D051722:ADAM Proteins; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; D013755:Tetradecanoylphorbol Acetate