Deregulation of the actin cytoskeleton and macropinocytosis in response to phorbol ester by the mutant protein kinase C gamma that causes spinocerebellar ataxia type 14.
Deregulation of the actin cytoskeleton and macropinocytosis in response to phorbol ester by the mutant protein kinase C gamma that causes spinocerebellar ataxia type 14.
复制标题
DOI：
10.3389/fphys.2014.00126
复制
发表时间：
2014
影响因子：
4
通讯作者：
Sakai N
中科院分区：
文献类型：
--
作者：
Yamamoto K;Seki T;Yamamoto H;Adachi N;Tanaka S;Hide I;Saito N;Sakai N
蛋白激酶Cγ（γPKC）基因的几个错义突变已被发现导致脊髓小脑共济失调14型（SCA 14），这是一种常染色体显性遗传的神经退行性疾病。γPKC是经典PKC的神经元特异性成员，并且由于多种刺激（包括二酰基甘油合成、细胞内Ca 2+增加和佛波醇酯）而被激活并移位至亚细胞区域。我们通过用TPA（12-O-tetradecanoylpholbol 13-acetate）刺激HeLa细胞来研究SCA 14突变是否影响γ PKC相关功能。野生型（WT）γPKC-GFP在TPA刺激10 min内移位至质膜，随后以PKC激酶活性和微管依赖性方式移位至核周并导致细胞皱缩。另一方面，尽管SCA 14突变体γPKC-GFP表现出类似的向质膜的易位，但随后的核周易位和细胞收缩在对TPA的响应中显著受损。易位的WT γPKC与F-actin共定位，并在核周区域形成大的囊泡结构。TPA刺激可促进表达WT γPKC的细胞对巨胞饮标志物--FITC-dextran的吸收，并且FITC-dextran被γ PKC阳性囊泡包围。TPA还可诱导PKC的膜底物MARCKS磷酸化，使磷酸化的MARCKS转位到核周区，提示TPA通过激活γPKC诱导巨胞饮。然而，TPA不能激活巨胞饮作用，也不能触发表达SCA 14突变体γPKC的细胞中磷酸化MARCKS的易位。这些结果表明，γPKC参与调节HeLa细胞的肌动蛋白细胞骨架和巨胞饮，而SCA 14突变体γPKC由于其质膜激酶活性降低而不能调节这些过程。这一特性可能与SCA 14的发病机制有关。
Several missense mutations in the protein kinase Cγ (γPKC) gene have been found to cause spinocerebellar ataxia type 14 (SCA14), an autosomal dominant neurodegenerative disease. γPKC is a neuron-specific member of the classical PKCs and is activated and translocated to subcellular regions as a result of various stimuli, including diacylglycerol synthesis, increased intracellular Ca2+ and phorbol esters. We investigated whether SCA14 mutations affect the γPKC-related functions by stimulating HeLa cells with TPA (12-O-tetradecanoylpholbol 13-acetate), a type of phorbol ester. Wild-type (WT) γPKC-GFP was translocated to the plasma membrane within 10 min of TPA stimulation, followed by its perinuclear translocation and cell shrinkage, in a PKC kinase activity- and microtubule-dependent manner. On the other hand, although SCA14 mutant γPKC-GFP exhibited a similar translocation to the plasma membrane, the subsequent perinuclear translocation and cell shrinkage were significantly impaired in response to TPA. Translocated WT γPKC colocalized with F-actin and formed large vesicular structures in the perinuclear region. The uptake of FITC-dextran, a marker of macropinocytosis, was promoted by TPA stimulation in cells expressing WT γPKC, and FITC-dextran was surrounded by γPKC-positive vesicles. Moreover, TPA induced the phosphorylation of MARCKS, which is a membrane-substrate of PKC, resulting in the translocation of phosphorylated MARCKS to the perinuclear region, suggesting that TPA induces macropinocytosis via γPKC activation. However, TPA failed to activate macropinocytosis and trigger the translocation of phosphorylated MARCKS in cells expressing the SCA14 mutant γPKC. These findings suggest that γPKC is involved in the regulation of the actin cytoskeleton and macropinocytosis in HeLa cells, while SCA14 mutant γPKC fails to regulate these processes due to its reduced kinase activity at the plasma membrane. This property might be involved in pathogenesis of SCA14.
登录
查看更多内容
影响因子：
4.2
作者：
Abreu, Bento J.;Guirnares, Maila;Uliana, Livia C.;Vigh, Jozsef;von Gersdorff, Henrique;Prado, Marco A.;Guatimosim, Cristina
通讯作者：
Guatimosim, Cristina
影响因子：
--
作者：
Brkanac, Z;Bylenok, L;Bird, TD
通讯作者：
Bird, TD
影响因子：
9.8
作者：
Chen, DH;Brkanac, Z;Raskind, WH
通讯作者：
Raskind, WH
DOI：
10.1152/ajpcell.1999.277.1.c163
发表时间：
1999-07-01
影响因子：
5.5
作者：
Carballo, E;Pitterle, DM;Blackshear, PJ
通讯作者：
Blackshear, PJ
影响因子：
4.8
作者：
Idkowiak-Baldys, Jolanta;Becker, Kevin P.;Hannun, Yusuf A.
通讯作者：
Hannun, Yusuf A.