Nucleolar localization of phosphatidylinositol 4-kinase PI4K230 in various mammalian cells.
BACKGROUND
Previous immunohistochemical investigations could not detect PI4K230, an isoform of mammalian phosphatidylinositol 4-kinases (also called type III alpha), in the nucleus and nucleolus of cells in spite of its predicted nuclear localization signals.


METHODS
Immunofluorescent detection of PI4K230 and other PI4K isoforms was performed on formaldehyde (PFA) or ethanol fixed cells and rat brain cryosections. Costaining with nucleolin and the effect of siRNA, Triton X-100, DNase, and RNase treatments were also tested to determine the localization of PI4K230.


RESULTS
PI4K230 gives a prominent signal in the nucleolus of ethanol fixed rat brain cryosections and of several cell types in addition to its presence in the nucleus and cytoplasm. The PI4K230 immunoreactivity of the nucleolus is masked in PFA fixed cells, but it can be restored by treatment of PFA fixed cells with hot wet citrate buffer or by washing the cryosections with PBS prior to PFA fixation. Nucleolar PI4K230 occurs in a Triton X-100 resistant complex. Treatment of COS-7 cells with siRNA targeting PI4K230 and permeabilized B50 cells with DNase or RNase results in the loss of PI4K230 signal from the nucleolus.


CONCLUSIONS
These experiments suggest the participation of PI4K230 in a DNase and RNase sensitive complex with a unique localization and function in the nucleolus.
Kakuk
Annamária
A
Department of Medical Chemistry, Faculty of Medicine, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary.
Friedländer
Elza
E
Vereb
György
G
Kása
Anita
A
Balla
András
A
Balla
Tamás
T
Heilmeyer
Ludwig M G
LM
Gergely
Pál
P
Vereb
György
G
D019870:1-Phosphatidylinositol 4-Kinase / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000166:cytology / Q000201:enzymology; D002460:Cell Line; D002466:Cell Nucleolus / Q000201:enzymology*; D002467:Cell Nucleus / Q000201:enzymology; D003593:Cytoplasm / Q000378:metabolism; D003851:Deoxyribonucleases / Q000737:chemistry; D000431:Ethanol / Q000737:chemistry; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D018613:Microscopy, Confocal; D009474:Neurons / Q000201:enzymology*; D017830:Octoxynol / Q000737:chemistry; D011108:Polymers / Q000737:chemistry; D034741:RNA, Small Interfering / Q000378:metabolism*; D051381:Rats; D012260:Ribonucleases / Q000737:chemistry; D013116:Spinal Cord / Q000166:cytology / Q000201:enzymology; D014162:Transfection
D011108:Polymers; D034741:RNA, Small Interfering; D005557:Formaldehyde; D000431:Ethanol; D017830:Octoxynol; D019870:1-Phosphatidylinositol 4-Kinase; D003851:Deoxyribonucleases; D012260:Ribonucleases; C003043:paraform