PACSIN3 binds ADAM12/meltrin alpha and up-regulates ectodomain shedding of heparin-binding epidermal growth factor-like growth factor.
A disintegrin and metalloprotease 12 (ADAM12/meltrin alpha) is a key enzyme implicated in the ectodomain shedding of membrane-anchored heparin-binding epidermal growth factor (EGF)-like growth factor (proHB-EGF)-dependent epidermal growth factor receptor (EGFR) transactivation. However, the activation mechanisms of ADAM12 are obscure. To determine how ADAM12 is activated, we screened proteins that bind to the cytoplasmic domain of ADAM12 using a yeast two-hybrid system and identified a protein called PACSIN3 that contains a Src homology 3 domain. An analysis of interactions between ADAM12 and PACSIN3 using glutathione S-transferase fusion protein revealed that a proline-rich region (amino acid residues 829-840) of ADAM12 was required to bind PACSIN3. Furthermore, co-immunoprecipitation and co-localization analyses of ADAM12 and PACSIN3 proteins also revealed their interaction in mammalian cells expressing both of them. The overexpression of PACSIN3 in HT1080 cells enhanced 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced proHB-EGF shedding. Furthermore, knockdown of endogenous PACSIN3 by small interfering RNA in HT1080 cells significantly attenuated the shedding of proHB-EGF induced by TPA and angiotensin II. Our data indicate that PACSIN3 has a novel function as an up-regulator in the signaling of proHB-EGF shedding induced by TPA and angiotensin II.
Mori
Seiji
S
Department of Molecular Pathology, School of Allied Health Science, Osaka University Faculty of Medicine, 1-7 Yamadaoka, Suita, Osaka 565-0871, USA.
Tanaka
Motonari
M
Nanba
Daisuke
D
Nishiwaki
Eiji
E
Ishiguro
Hiroshi
H
Higashiyama
Shigeki
S
Matsuura
Nariaki
N
D051722:ADAM Proteins; D000072199:ADAM12 Protein; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000737:chemistry; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins; D004815:Epidermal Growth Factor / Q000737:chemistry* / Q000378:metabolism; D017353:Gene Deletion; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000378:metabolism; D066257:Heparin-binding EGF-like Growth Factor; D006801:Humans; D015151:Immunoblotting; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism / Q000502:physiology*; D011233:Precipitin Tests; D011392:Proline / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate; D015533:Transcriptional Activation; D014162:Transfection; D020798:Two-Hybrid System Techniques; D015854:Up-Regulation
D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; C587883:HBEGF protein, human; C587884:Hbegf protein, mouse; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009124:Muscle Proteins; C421762:PACSIN3 protein, human; C486019:Pacsin3 protein, mouse; D010750:Phosphoproteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D000804:Angiotensin II; D004815:Epidermal Growth Factor; D011392:Proline; D005982:Glutathione Transferase; D051722:ADAM Proteins; D000072199:ADAM12 Protein; C000606480:ADAM12 protein, human; C096115:Adam12 protein, mouse; D008666:Metalloendopeptidases; D013755:Tetradecanoylphorbol Acetate