Abstract
In order to characterize the effects of increasing phosphatidylinositol(4,5)bisphosphate (PtdIns(4,5)P(2)) on nuclear function, we expressed the human phosphatidylinositol (4)-phosphate 5-kinase (HsPIP5K) 1α in Nicotiana tabacum (NT) cells. The HsPIP5K-expressing (HK) cells had altered nuclear lipids and nuclear functions. HK cell nuclei had 2-fold increased PIP5K activity and increased steady state PtdIns(4,5)P(2). HK nuclear lipid classes showed significant changes compared to NT (wild type) nuclear lipid classes including increased phosphatidylserine (PtdSer) and phosphatidylcholine (PtdCho) and decreased lysolipids. Lipids isolated from protoplast plasma membranes (PM) were also analyzed and compared with nuclear lipids. The lipid profiles revealed similarities and differences in the plasma membrane and nuclei from the NT and transgenic HK cell lines. A notable characteristic of nuclear lipids from both cell types is that PtdIns accounts for a higher mol% of total lipids compared to that of the protoplast PM lipids. The lipid molecular species composition of each lipid class was also analyzed for nuclei and protoplast PM samples. To determine whether expression of HsPIP5K1α affected plant nuclear functions, we compared DNA replication, histone 3 lysine 9 acetylation (H3K9ac) and phosphorylation of the retinoblastoma protein (pRb) in NT and HK cells. The HK cells had a measurable decrease in DNA replication, histone H3K9 acetylation and pRB phosphorylation.