The role of the C-terminus in functional expression and internalization of rat connexin46 (rCx46)
Tóm tắt
The C-terminus (CT) of rCx46 consists of 186 residues (H230-I416). Recent studies showed that rCx4628.2, truncated after H243, altered the formation of functional hemichannels when expressed in Xenopus oocytes, while rCx4637.7, truncated after A333 formed gap junction hemichannels similarly to rCx46wt. To analyze the role of the CT up to A333 in functional expression with cell imaging and dye-transfer techniques, different mutants were generated by C-terminal truncation between H243-A333, labeled with EGFP and expressed in HeLa cells. These rCx46 variants were characterized according to their compartmentalization in organelles, their presence in microscopic detectable vesicles and their ability to form gap junction plaques. rCx46 truncated after A311 (rCx4635.3) was compartmentalized, was found in vesicles and formed functional gap junction plaques similarly to rCx46wt. With a truncation after P284 (rCx4632.6), the protein was not compartmentalized and the amount of vesicles containing the protein were reduced; however, functional gap junction plaque formation was not affected as compared to rCx4635.3. rCx4628.2 did not form functional gap junction plaques; it was not found in vesicles or in cellular compartments. Live-cell imaging and detection of annular junctions for rCx4632.6 and rCx4635.3 revealed that the truncation after P284 reduced the frequency of vesicle budding from gap junction plaques and the formation of annular junctions. These results suggest that the C-terminal region of rCx46 up to A311 (rCx4635.3) is necessary for its correct compartmentalization and internalization in the form of annular junctions, while the H230-P284 C-terminal region (rCx4632.6) is sufficient for the formation of dye coupled gap junction channels.
Tài liệu tham khảo
citation_journal_title=Am J Physiol; citation_title=Phenotypic control of gap junctional communication by cultured alveolar epithelial cells; citation_author=V Abraham, ML Chou, KM DeBolt, M Koval; citation_volume=276; citation_publication_date=1999; citation_pages=L825-L834; citation_id=CR1
citation_journal_title=Biochem Biophys Res Commun; citation_title=Molecular cloning and functional analysis of a novel Cx43 partner protein CIP150; citation_author=M Akiyama, N Ishida, T Ogawa, K Yogo, T Takeya; citation_volume=335; citation_publication_date=2005; citation_pages=1264-1271; citation_doi=10.1016/j.bbrc.2005.08.019; citation_id=CR2
citation_journal_title=Life Sci; citation_title=Altered expression of connexins in urethane-induced mouse lung adenomas; citation_author=JL Avanzo, M Mesnil, FJ Hernandez-Blazquez, TC Silva, H Fukumasu, CM Mori, H Yamasaki, ML Dagli; citation_volume=79; citation_publication_date=2006; citation_pages=2202-2208; citation_doi=10.1016/j.lfs.2006.07.033; citation_id=CR3
citation_journal_title=Int J Cancer; citation_title=A novel role of gap junction connexin46 protein to protect breast tumors from hypoxia; citation_author=D Banerjee, G Gakhar, D Madgwick, A Hurt, D Takemoto, TA Nguyen; citation_volume=127; citation_publication_date=2010; citation_pages=839-848; citation_id=CR4
citation_journal_title=Antioxid Redox Signal; citation_title=Oxidative stress, lens gap junctions, and cataracts; citation_author=VM Berthoud, EC Beyer; citation_volume=11; citation_publication_date=2009; citation_pages=339-353; citation_doi=10.1089/ars.2008.2119; citation_id=CR5
citation_journal_title=Cardiovasc Res; citation_title=Pathways for degradation of connexins and gap junctions; citation_author=VM Berthoud, PJ Minogue, JG Laing, EC Beyer; citation_volume=62; citation_publication_date=2004; citation_pages=256-267; citation_doi=10.1016/j.cardiores.2003.12.021; citation_id=CR6
citation_journal_title=Mol Biol Cell; citation_title=Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels; citation_author=Z Chai, DA Goodenough, DL Paul; citation_volume=22; citation_publication_date=2011; citation_pages=4503-4512; citation_doi=10.1091/mbc.E11-05-0438; citation_id=CR7
citation_journal_title=Glia; citation_title=Nerve injury and inflammatory cytokines modulate gap junctions in the peripheral nervous system; citation_author=KJ Chandross; citation_volume=24; citation_publication_date=1998; citation_pages=21-31; citation_doi=10.1002/(SICI)1098-1136(199809)24:1<21::AID-GLIA3>3.0.CO;2-3; citation_id=CR8
citation_journal_title=Mol Biol Cell; citation_title=ERp29 restricts Connexin43 oligomerization in the endoplasmic reticulum; citation_author=S Das, TD Smith, JD Sarma, JD Ritzenthaler, J Maza, BE Kaplan, LA Cunningham, L Suaud, MJ Hubbard, RC Rubenstein, M Koval; citation_volume=20; citation_publication_date=2009; citation_pages=2593-2604; citation_doi=10.1091/mbc.E08-07-0790; citation_id=CR9
citation_journal_title=Cell Commun Adhes; citation_title=Identification of rab20 as a potential regulator of connexin 43 trafficking; citation_author=J Das Sarma, BE Kaplan, D Willemsen, M Koval; citation_volume=15; citation_publication_date=2008; citation_pages=65-74; citation_doi=10.1080/15419060802014305; citation_id=CR10
citation_journal_title=Cell Commun Signal; citation_title=Connexins: a myriad of functions extending beyond assembly of gap junction channels; citation_author=HA Dbouk, RM Mroue, ME El-Sabban, RS Talhouk; citation_volume=7; citation_publication_date=2009; citation_pages=4; citation_doi=10.1186/1478-811X-7-4; citation_id=CR11
citation_journal_title=Novartis Found Symp; citation_title=Trafficking pathways leading to the formation of gap junctions; citation_author=WH Evans, S Ahmad, J Diez, CH George, JM Kendall, PE Martin; citation_volume=219; citation_publication_date=1999; citation_pages=44-54; citation_id=CR12
citation_journal_title=J Cell Biol; citation_title=Membrane insertion of gap junction connexins: polytopic channel forming membrane proteins; citation_author=MM Falk, NM Kumar, NB Gilula; citation_volume=127; citation_publication_date=1994; citation_pages=343-355; citation_doi=10.1083/jcb.127.2.343; citation_id=CR13
citation_journal_title=Mol Biol Cell; citation_title=Gap junction turnover is achieved by the internalization of small endocytic double-membrane vesicles; citation_author=MM Falk, SM Baker, AM Gumpert, D Segretain, RW Buckheit; citation_volume=20; citation_publication_date=2009; citation_pages=3342-3352; citation_doi=10.1091/mbc.E09-04-0288; citation_id=CR14
citation_journal_title=Curr Biol; citation_title=The gap junction protein connexin43 interacts with the second PDZ domain of the zonula occludens-1 protein; citation_author=BN Giepmans, WH Moolenaar; citation_volume=8; citation_publication_date=1998; citation_pages=931-934; citation_doi=10.1016/S0960-9822(07)00375-2; citation_id=CR15
citation_journal_title=Dev Biol; citation_title=Modulation of cell junctions during differentiation of the chicken otocyst sensory epithelium; citation_author=RD Ginzberg, NB Gilula; citation_volume=68; citation_publication_date=1979; citation_pages=110-129; citation_doi=10.1016/0012-1606(79)90247-1; citation_id=CR16
citation_journal_title=Prog Biophys Mol Biol; citation_title=Gap junctional complexes: from partners to functions; citation_author=JC Hervé, N Bourmeyster, D Sarrouilhe, HS Duffy; citation_volume=94; citation_publication_date=2007; citation_pages=29-65; citation_doi=10.1016/j.pbiomolbio.2007.03.010; citation_id=CR17
citation_journal_title=Methods; citation_title=Identification of connexin-interacting proteins: application of the yeast two-hybrid screen; citation_author=C Jin, AF Lau, KD Martyn; citation_volume=20; citation_publication_date=2000; citation_pages=219-231; citation_doi=10.1006/meth.1999.0939; citation_id=CR18
citation_journal_title=Mol Biol Cell; citation_title=Trafficking, assembly, and function of a connexin43-green fluorescent protein chimera in live mammalian cells; citation_author=K Jordan, JL Solan, M Dominguez, M Sia, A Hand, PD Lampe, DW Laird; citation_volume=10; citation_publication_date=1999; citation_pages=2033-2050; citation_id=CR19
citation_journal_title=J Cell Sci; citation_title=The origin of annular junctions: a mechanism of gap junction internalization; citation_author=K Jordan, R Chodock, AR Hand, DW Laird; citation_volume=114; citation_publication_date=2001; citation_pages=763-773; citation_id=CR20
citation_journal_title=Trends Cell Biol; citation_title=Pathways and control of connexin oligomerization; citation_author=M Koval; citation_volume=16; citation_publication_date=2006; citation_pages=159-166; citation_doi=10.1016/j.tcb.2006.01.006; citation_id=CR21
citation_journal_title=J Cell Biol; citation_title=Connexin46 is retained as monomers in a trans-Golgi compartment of osteoblastic cells; citation_author=M Koval, JE Harley, E Hick, TH Steinberg; citation_volume=137; citation_publication_date=1997; citation_pages=847-857; citation_doi=10.1083/jcb.137.4.847; citation_id=CR22
citation_journal_title=Trends Cell Biol; citation_title=The gap junction proteome and its relationship to disease; citation_author=DW Laird; citation_volume=20; citation_publication_date=2010; citation_pages=92-101; citation_doi=10.1016/j.tcb.2009.11.001; citation_id=CR23
citation_journal_title=Biochem J; citation_title=Life cycle of connexins in health and disease; citation_author=DW Laird; citation_volume=394; citation_publication_date=2006; citation_pages=527-543; citation_doi=10.1042/BJ20051922; citation_id=CR24
citation_journal_title=Arch Biochem Biophys; citation_title=Regulation of gap junctions by phosphorylation of connexins; citation_author=PD Lampe, AF Lau; citation_volume=384; citation_publication_date=2000; citation_pages=205-215; citation_doi=10.1006/abbi.2000.2131; citation_id=CR25
citation_journal_title=Int J Biochem Cell Biol; citation_title=The effects of connexin phosphorylation on gap junctional communication; citation_author=PD Lampe, AF Lau; citation_volume=36; citation_publication_date=2004; citation_pages=1171-1186; citation_doi=10.1016/S1357-2725(03)00264-4; citation_id=CR26
citation_journal_title=Biochemistry; citation_title=Novel rab GAP-like protein, CIP85, interacts with connexin43 and induces its degradation; citation_author=Z Lan, WE Kurata, KD Martyn, C Jin, AF Lau; citation_volume=44; citation_publication_date=2005; citation_pages=2385-2396; citation_doi=10.1021/bi048306w; citation_id=CR27
citation_journal_title=J Cell Biol; citation_title=Evidence for the participation of actin microfilaments and bristle coats in the internalization of gap junction membrane; citation_author=WJ Larsen, HN Tung, SA Murray, CA Swenson; citation_volume=83; citation_publication_date=1979; citation_pages=576-587; citation_doi=10.1083/jcb.83.3.576; citation_id=CR28
citation_journal_title=J Cell Sci; citation_title=Ubiquitin protein ligase Nedd4 binds to connexin43 by a phosphorylation-modulated process; citation_author=K Leykauf, M Salek, J Bomke, M Frech, WD Lehmann, M Durst, A Alonso; citation_volume=119; citation_publication_date=2006; citation_pages=3634-3642; citation_doi=10.1242/jcs.03149; citation_id=CR29
citation_journal_title=Histochem Cell Biol; citation_title=Limiting transport steps and novel interactions of Connexin-43 along the secretory pathway; citation_author=IV Majoul, D Onichtchouk, E Butkevich, D Wenzel, LM Chailakhyan, R Duden; citation_volume=132; citation_publication_date=2009; citation_pages=263-280; citation_doi=10.1007/s00418-009-0617-x; citation_id=CR30
citation_journal_title=Biochem J; citation_title=Targeting motifs and functional parameters governing the assembly of connexins into gap junctions; citation_author=PE Martin, J Steggles, C Wilson, S Ahmad, WH Evans; citation_volume=350; citation_issue=Pt 3; citation_publication_date=2000; citation_pages=943; citation_doi=10.1042/0264-6021:3500943; citation_id=CR31
citation_journal_title=J Cell Sci; citation_title=Multiple pathways in the trafficking and assembly of connexin 26, 32 and 43 into gap junction intercellular communication channels; citation_author=PE Martin, G Blundell, S Ahmad, RJ Errington, WH Evans; citation_volume=114; citation_publication_date=2001; citation_pages=3845-3855; citation_id=CR32
citation_journal_title=Circ Res; citation_title=Fate of intercellular junctions in isolated adult rat cardiac cells; citation_author=F Mazet, BA Wittenberg, DC Spray; citation_volume=56; citation_publication_date=1985; citation_pages=195-204; citation_doi=10.1161/01.RES.56.2.195; citation_id=CR33
citation_journal_title=J Biol Chem; citation_title=An aberrant sequence in a connexin46 mutant underlies congenital cataracts; citation_author=PJ Minogue, X Liu, L Ebihara, EC Beyer, VM Berthoud; citation_volume=280; citation_publication_date=2005; citation_pages=40788-44095; citation_doi=10.1074/jbc.M504765200; citation_id=CR34
citation_journal_title=Endocr Res; citation_title=Endocytosis of connexin protein in adrenal cells; citation_author=SA Murray, BM Nickel, VL Gay; citation_volume=30; citation_publication_date=2004; citation_pages=647-654; citation_doi=10.1081/ERC-200043942; citation_id=CR35
citation_journal_title=J Cell Biol; citation_title=Biochemical analysis of connexin43 intracellular transport, phosphorylation, and assembly into gap junctional plaques; citation_author=LS Musil, DA Goodenough; citation_volume=115; citation_publication_date=1991; citation_pages=1357-1374; citation_doi=10.1083/jcb.115.5.1357; citation_id=CR36
citation_journal_title=Pflugers Arch; citation_title=Inactivation of expressed and conducting rCx46 hemichannels by phosphorylation; citation_author=A Ngezahayo, C Zeilinger, I Todt, I Marten, HA Kolb; citation_volume=436; citation_publication_date=1998; citation_pages=627-629; citation_doi=10.1007/s004240050681; citation_id=CR37
citation_journal_title=Biochem Biophys Res Commun; citation_title=Clathrin and Cx43 gap junction plaque endoexocytosis; citation_author=BM Nickel, BH DeFranco, VL Gay, SA Murray; citation_volume=374; citation_publication_date=2008; citation_pages=679-682; citation_doi=10.1016/j.bbrc.2008.07.108; citation_id=CR38
citation_journal_title=Cell Commun Adhes; citation_title=Characterization of the association of connexins and ZO-1 in the lens; citation_author=PA Nielsen, A Baruch, BN Giepmans, NM Kumar; citation_volume=8; citation_publication_date=2001; citation_pages=213-217; citation_doi=10.3109/15419060109080726; citation_id=CR39
citation_journal_title=Mol Biol Cell; citation_title=Lens connexins alpha3Cx46 and alpha8Cx50 interact with zonula occludens protein-1 (ZO-1); citation_author=PA Nielsen, A Baruch, VI Shestopalov, BN Giepmans, I Dunia, EL Benedetti, NM Kumar; citation_volume=14; citation_publication_date=2003; citation_pages=2470-2481; citation_doi=10.1091/mbc.E02-10-0637; citation_id=CR40
citation_journal_title=J Cell Biol; citation_title=Connexin46, a novel lens gap junction protein, induces voltage-gated currents in nonjunctional plasma membrane of Xenopus oocytes; citation_author=DL Paul, L Ebihara, LJ Takemoto, KI Swenson, DA Goodenough; citation_volume=115; citation_publication_date=1991; citation_pages=1077-1089; citation_doi=10.1083/jcb.115.4.1077; citation_id=CR41
citation_journal_title=Exp Cell Res; citation_title=The role of the cytoskeleton in the formation of gap junctions by Connexin 30; citation_author=C Qu, P Gardner, I Schrijver; citation_volume=315; citation_publication_date=2009; citation_pages=1683-1692; citation_doi=10.1016/j.yexcr.2009.03.001; citation_id=CR42
citation_journal_title=Adv Cardiol; citation_title=Life cycle of connexins: regulation of connexin synthesis and degradation; citation_author=A Salameh; citation_volume=42; citation_publication_date=2006; citation_pages=57-70; citation_doi=10.1159/000092562; citation_id=CR43
citation_journal_title=Biochemistry; citation_title=Connexin family members target to lipid raft domains and interact with caveolin-1; citation_author=AL Schubert, W Schubert, DC Spray, MP Lisanti; citation_volume=41; citation_publication_date=2002; citation_pages=5754-5764; citation_doi=10.1021/bi0121656; citation_id=CR44
citation_journal_title=Biochim Biophys Acta; citation_title=Regulation of connexin biosynthesis, assembly, gap junction formation, and removal; citation_author=D Segretain, MM Falk; citation_volume=1662; citation_publication_date=2004; citation_pages=3-21; citation_doi=10.1016/j.bbamem.2004.01.007; citation_id=CR45
citation_journal_title=Cell; citation_title=Microtubule plus-end-tracking proteins target gap junctions directly from the cell interior to adherens junctions; citation_author=RM Shaw, AJ Fay, MA Puthenveedu, M Zastrow, YN Jan, LY Jan; citation_volume=128; citation_publication_date=2007; citation_pages=547-560; citation_doi=10.1016/j.cell.2006.12.037; citation_id=CR46
citation_journal_title=J Neurocytol; citation_title=Porocytosis: secretion from small and medium-diameter vesicles and vesicle arrays without membrane fusion; citation_author=RB Silver, ME Kriebel, B Keller, GD Pappas; citation_volume=32; citation_publication_date=2003; citation_pages=277-291; citation_doi=10.1023/B:NEUR.0000010086.07636.df; citation_id=CR47
citation_journal_title=Cardiovasc Res; citation_title=Gap junctions and the connexin protein family; citation_author=G Sohl, K Willecke; citation_volume=62; citation_publication_date=2004; citation_pages=228-232; citation_doi=10.1016/j.cardiores.2003.11.013; citation_id=CR48
citation_journal_title=J Cell Sci; citation_title=A tyrosine-based sorting signal is involved in connexin43 stability and gap junction turnover; citation_author=MA Thomas, N Zosso, I Scerri, N Demaurex, M Chanson, O Staub; citation_volume=116; citation_publication_date=2003; citation_pages=2213-2222; citation_doi=10.1242/jcs.00440; citation_id=CR49
citation_journal_title=Nature; citation_title=Structure of the junction between communicating cells; citation_author=PN Unwin, G Zampighi; citation_volume=283; citation_publication_date=1980; citation_pages=545-549; citation_doi=10.1038/283545a0; citation_id=CR50
citation_journal_title=J Bioenerg Biomembr; citation_title=Phosphorylation in the C-terminus of the rat connexin46 (rCx46) and regulation of the conducting activity of the formed connexons; citation_author=WJ Walter, C Zeilinger, W Bintig, HA Kolb, A Ngezahayo; citation_volume=40; citation_publication_date=2008; citation_pages=397-405; citation_doi=10.1007/s10863-008-9151-0; citation_id=CR51
citation_journal_title=Biochem Biophys Res Commun; citation_title=The role of the Cx43 C-terminus in GJ plaque formation and internalization; citation_author=P Wayakanon, R Bhattacharjee, K Nakahama, I Morita; citation_volume=420; citation_publication_date=2012; citation_pages=456-461; citation_doi=10.1016/j.bbrc.2012.03.018; citation_id=CR52
citation_journal_title=J Biol Chem; citation_title=Connexin43 associated with an N-cadherin-containing multiprotein complex is required for gap junction formation in NIH3T3 cells; citation_author=CJ Wei, R Francis, X Xu, CW Lo; citation_volume=280; citation_publication_date=2005; citation_pages=19925-19936; citation_doi=10.1074/jbc.M412921200; citation_id=CR53
citation_journal_title=J Bioenerg Biomembr; citation_title=Multiple connexin proteins in single intercellular channels: connexin compatibility and functional consequences; citation_author=TW White, R Bruzzone; citation_volume=28; citation_publication_date=1996; citation_pages=339-350; citation_doi=10.1007/BF02110110; citation_id=CR54
citation_journal_title=Curr Opin Struct Biol; citation_title=Synthesis, assembly and structure of gap junction intercellular channels; citation_author=M Yeager, VM Unger, MM Falk; citation_volume=8; citation_publication_date=1998; citation_pages=517-524; citation_doi=10.1016/S0959-440X(98)80131-0; citation_id=CR55
citation_journal_title=Invest Ophthalmol Vis Sci; citation_title=Regulation of lens cell-to-cell communication by activation of PKCgamma and disassembly of Cx50 channels; citation_author=GA Zampighi, AM Planells, D Lin, D Takemoto; citation_volume=46; citation_publication_date=2005; citation_pages=3247-3255; citation_doi=10.1167/iovs.04-1504; citation_id=CR56
citation_journal_title=Mol Biol Cell; citation_title=Membrane integration of in vitro-translated gap junctional proteins: co- and post-translational mechanisms; citation_author=JT Zhang, M Chen, CI Foote, BJ Nicholson; citation_volume=7; citation_publication_date=1996; citation_pages=471-482; citation_id=CR57