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Inhibition of Rho‐kinase‐induced myristoylated alanine‐rich C kinase substrate (MARCKS) phosphorylation in human neuronal cells by H‐1152, a novel and specific Rho‐kinase inhibitor
Author(s) -
Ikenoya Mami,
Hidaka Hiroyoshi,
Hosoya Takamitsu,
Suzuki Masaaki,
Yamamoto Naoki,
Sasaki Yasuharu
Publication year - 2002
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2002.00801.x
Subject(s) - marcks , protein kinase c , rho associated protein kinase , phosphorylation , kinase , rho kinase inhibitor , microbiology and biotechnology , protein kinase a , biology , myristoylation , chemistry , biochemistry
Abstract The functions of small G protein Rho‐associated kinase (Rho‐kinase) have been determined in muscle and non‐muscle cells, but, particularly in neuronal cells, its effector(s) has not been well known. Recently, we preliminarily reported that Rho‐kinase phosphorylates the Ser159 residue in myristoylated alanine‐rich C kinase substrate (MARCKS) in vitro , but it remains obscure in vivo . To further clarify this point, we developed an isoquinolinesulfonamide derivative, H‐1152, that is a more specific, stronger and membrane‐permeable inhibitor of Rho‐kinase with a K i value of 1.6 n m , but poor inhibitor of other serine/threonine kinases. H‐1152 dose‐ dependently inhibited the phosphorylation of MARCKS in human neuroteratoma (NT‐2) cells stimulated by Rho‐activator lysophosphatidic acid (LPA), which was determined by phosphorylation site‐specific antibody against phospho‐Ser159 in MARCKS, whereas it hardly inhibited the phosphorylation stimulated by phorbol‐12,13‐dibutyrate (PDBu). In contrast, two other Rho‐kinase inhibitors, HA‐1077 at 30 µ m and Y‐27632 at 10–30 µ m , inhibited the phosphorylation of MARCKS in the cells stimulated by LPA and PDBu. A PKC inhibitor Ro‐31‐8220 selectively inhibited PDBu‐induced phosphorylation of MARCKS. Taken together with our previous results, the present findings strongly suggest that Rho/Rho‐kinase phosphorylates MARCKS at Ser159 residue in neuronal cells in response to LPA stimulation and that H‐1152 is a useful tool to confirm Rho‐kinase function(s) in cells and tissues.