ClC-3 chloride channels facilitate endosomal acidification and chloride accumulation

被引:150
作者
Hara-Chikuma, M
Yang, BX
Sonawane, ND
Sasaki, S
Uchida, S
Verkman, AS
机构
[1] Univ Calif San Francisco, Cardiovasc Res Inst, Dept Med, San Francisco, CA 94143 USA
[2] Univ Calif San Francisco, Cardiovasc Res Inst, Dept Physiol, San Francisco, CA 94143 USA
[3] Tokyo Med & Dent Univ, Grad Sch, Dept Nephrol, Tokyo 1138519, Japan
关键词
D O I
10.1074/jbc.M407030200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We investigated the involvement of ClC-3 chloride channels in endosomal acidification by measurement of endosomal pH and chloride concentration [Cl-] in control versus ClC-3-deficient hepatocytes and in control versus ClC-3-transfected Chinese hamster ovary cells. Endosomes were labeled with pH or [Cl-]-sensing fluorescent transferrin (Tf), which targets to early/recycling endosomes, or alpha(2)-macroglobulin (alpha(2)M), which targets to late endosomes. In pulse label-chase experiments, [Cl-] was 19 mM just after internalization in alpha(2)M-labeled endosomes in primary cultures of hepatocytes from wildtype mice, increasing to 58 mM over 45 min, whereas pH decreased from 7.1 to 5.4. Endosomal acidification and [Cl-] accumulation were significantly impaired in hepatocytes from ClC-3 knock-out mice, with [Cl-] increasing from 16 to 43 mM and pH decreasing from 7.1 to 6.0. Acidification and Cl- accumulation were blocked by bafilomycin. In Tf-labeled endosomes, [Cl-] was 46 mM in wild-type versus 35 mM in ClC-3-deficient hepatocytes at 15 min after internalization, with corresponding pH of 6.1 versus 6.5. Approximately 4-fold increased Cl- conductance was found in alpha(2)M-labeled endosomes isolated from hepatocytes of wild-type versus ClC-3 null mice. In contrast, Golgi acidification was not impaired in ClC-3-deficient hepatocytes. In transfected Chinese hamster ovary cells expressing ClC-3A, endosomal acidification and [Cl-] accumulation were enhanced. [Cl-] in alpha(2)M-labeled endosomes was 42 mM ( control) versus 53 mM (ClC-3A) at 45 min, with corresponding pH 5.8 versus 5.2; [Cl-] in Tf-labeled endosomes at 15 min was 37 mM (control) versus 49 mM (ClC-3A) with pH 6.3 versus 5.9. Our results provide direct evidence for involvement of ClC-3 in endosomal acidification by Cl- shunting of the interior-positive membrane potential created by the vacuolar H+ pump.
引用
收藏
页码:1241 / 1247
页数:7
相关论文
共 38 条
[31]   Disruption of CIC-3, a chloride channel expressed on synaptic vesicles, leads to a loss of the hippocampus [J].
Stobrawa, SM ;
Breiderhoff, T ;
Takamori, S ;
Engel, D ;
Schweizer, M ;
Zdebik, AA ;
Bösl, MR ;
Ruether, K ;
Jahn, H ;
Draguhn, A ;
Jahn, R ;
Jentsch, TJ .
NEURON, 2001, 29 (01) :185-196
[32]   Human ClC-3 is not the swelling-activated chloride channel involved in cell volume regulation [J].
Weylandt, KH ;
Valverde, MA ;
Nobles, M ;
Raguz, S ;
Amey, JS ;
Diaz, M ;
Nastrucci, C ;
Higgins, CF ;
Sardini, A .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (20) :17461-17467
[33]   KINETICS OF ALPHA-2-MACROGLOBULIN ENDOCYTOSIS AND DEGRADATION IN MUTANT AND WILD-TYPE CHINESE-HAMSTER OVARY CELLS [J].
YAMASHIRO, DJ ;
BORDEN, LA ;
MAXFIELD, FR .
JOURNAL OF CELLULAR PHYSIOLOGY, 1989, 139 (02) :377-382
[34]   KINETICS OF ENDOSOME ACIDIFICATION IN MUTANT AND WILD-TYPE CHINESE-HAMSTER OVARY CELLS [J].
YAMASHIRO, DJ ;
MAXFIELD, FR .
JOURNAL OF CELL BIOLOGY, 1987, 105 (06) :2713-2721
[35]   SEGREGATION OF TRANSFERRIN TO A MILDLY ACIDIC (PH 6.5) PARA-GOLGI COMPARTMENT IN THE RECYCLING PATHWAY [J].
YAMASHIRO, DJ ;
TYCKO, B ;
FLUSS, SR ;
MAXFIELD, FR .
CELL, 1984, 37 (03) :789-800
[36]   Water and glycerol permeabilities of aquaporins 1-5 and MIP determined quantitatively by expression of epitope-tagged constructs in Xenopus oocytes [J].
Yang, BX ;
Verkman, AS .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1997, 272 (26) :16140-16146
[37]   CLC-3 deficiency leads to phenotypes similar to human neuronal ceroid lipofuscinosis [J].
Yoshikawa, M ;
Uchida, S ;
Ezaki, J ;
Rai, T ;
Hayama, A ;
Kobayashi, K ;
Kida, Y ;
Noda, M ;
Koike, M ;
Uchiyama, Y ;
Marumo, F ;
Kominami, E ;
Sasaki, S .
GENES TO CELLS, 2002, 7 (06) :597-605
[38]   2ND MESSENGERS REGULATE ENDOSOMAL ACIDIFICATION IN SWISS 3T3 FIBROBLASTS [J].
ZEN, K ;
BIWERSI, J ;
PERIASAMY, N ;
VERKMAN, AS .
JOURNAL OF CELL BIOLOGY, 1992, 119 (01) :99-110