Subnuclear localization and differentiation-dependent increased expression of DGK-zeta in C2C12 mouse myoblasts.
Diacylglycerol kinases (DGKs) catalyze phosphorylation of diacylglycerol (DG) to yield phosphatidic acid (PA). Previous evidence has shown that the nucleus contains several DGK isoforms. In this study, we have analyzed the expression and subnuclear localization of DGK-zeta employing C2C12 mouse myoblasts. Immunocytochemistry coupled to confocal laser scanning microscopy showed that both endogenous and green fluorescent protein-tagged overexpressed DGK-zeta localized mostly to the nucleus. In contrast, overexpressed DGK-alpha, -beta, -delta, and -iota did not migrate to the nucleus. DGK-zeta was present in the nuclear speckle domains, as also revealed by immuno-electron microscopy analysis. Moreover, DGK-zeta co-localized and interacted with phosphoinositide-specific phospholipase Cbeta1 (PLCbeta1), that is involved in inositide-dependent signaling pathways important for the regulation of cell proliferation and differentiation. Furthermore, we report that DGK-zeta associated with nuclear matrix, the fundamental organizing principle of the nucleus where many cell functions take place, including DNA replication, gene expression, and protein phosphorylation. Nuclear DGK-zeta increased during myogenic differentiation of C2C12 cells, while DGK-zeta down-regulation by siRNA markedly impaired differentiation. Overall, our findings further support the importance of speckles and nuclear matrix in lipid-dependent signaling and suggest that nuclear DGK-zeta might play some fundamental role during myogenic differentiation of C2C12 cells.
Evangelisti
Camilla
C
Dipartimento di Scienze Anatomiche Umane e Fisiopatologia dell'Apparato Locomotore, Sezione di Anatomia, Università di Bologna, Bologna, Italy.
Riccio
Massimo
M
Faenza
Irene
I
Zini
Nicoletta
N
Hozumi
Yasukazu
Y
Goto
Kaoru
K
Cocco
L
L
Martelli
Alberto M
AM
D000546:Amanitins / Q000494:pharmacology; D000818:Animals; D001562:Benzimidazoles / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D019852:Diacylglycerol Kinase / Q000378:metabolism*; D015536:Down-Regulation / Q000187:drug effects; D006570:Heterochromatin / Q000648:ultrastructure; D007527:Isoenzymes / Q000378:metabolism; D051379:Mice; D024510:Muscle Development / Q000187:drug effects; D032448:Myoblasts, Skeletal / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D015530:Nuclear Matrix / Q000378:metabolism*; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D054799:Phospholipase C beta; D010770:Phosphotransferases / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D010738:Type C Phospholipases / Q000378:metabolism
C110962:5,6-dichlorobenzimidazole; D000546:Amanitins; D001562:Benzimidazoles; D006570:Heterochromatin; D007527:Isoenzymes; D019269:Phosphatidylinositol 4,5-Diphosphate; D010770:Phosphotransferases; D019852:Diacylglycerol Kinase; C512480:diacylglycerol kinase zeta, mouse; D010738:Type C Phospholipases; D054799:Phospholipase C beta