pPKCδ activates SC35 splicing factor during H9c2 myoblastic differentiation.
Although Protein Kinase C (PKC) isoforms' role in the neonatal and adult cardiac tissue development and ageing has been widely described "in vivo", the interaction of such enzymes with specific nuclear substrates needs to be investigated. The aim of our research has been the study of the expression, localization and interaction with the splicing factor SC35 of PKC isoforms (α, δ, ε, ζ) and their potential role in modulating the transcription machinery. H9c2 cells induced to myoblast differentiation in the presence of 1% Horse Serum (HS) have represented our experimental model. The expression of PKC isoforms, their distribution and interaction with SC35 have been evaluated by western blotting, co-immunoprecipitation and double gold immunolabeling for transmission and scanning electron microscopy. Our results show PKCδ as the most expressed isoform in differentiated cells. Surprisingly, the distribution of PKCδ and SC35 does not show any significant modification between 10%FBS and 1%HS treated samples and no co-localization is observed. Moreover the interaction between the phosphorylated form of PKCδ (pPKCδ) and SC35 increases, is distributed and co-localizes within the nucleus of differentiated H9c2. These data represent reasonable evidence of pPKCδ mediated SC35 splicing factor activation, suggesting its direct effect on transcription via interaction with the transcription machinery. Furthermore, this co-localization represents a crucial event resulting in downstream changes in transcription of components which determine the morphological modifications related to cardiomyoblast differentiated phenotype.
Department of Drug Sciences, Faculty of Pharmacy, University G d'Annunzio, Chieti-Pescara, Italy. s.zara@unich.it
Susi
S
Zara
Mirella
M
Falconi
Monica
M
Rapino
Michela
M
Zago
Giovanna
G
Orsini
Giovanni
G
Mazzotti
Amelia
A
Cataldi
Gabriella
G
Teti
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D047468:Immunoprecipitation; D016253:Microscopy, Immunoelectron; D032386:Myoblasts, Cardiac / Q000166:cytology / Q000378:metabolism*; D032448:Myoblasts, Skeletal / Q000166:cytology / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012326:RNA Splicing; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D012261:Ribonucleoproteins / Q000378:metabolism
D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D012261:Ribonucleoproteins; D051745:Protein Kinase C-delta