A Role for Nuclear Phospholipase Cβ1 in Cell Cycle Control

Abstract

Phosphoinositide signaling resides in the nucleus, and among the enzymes of the cycle, phospholipase C (PLC) appears as the key element both in Saccharomyces cerevisiae and in mammalian cells. The yeast PLC pathway produces multiple inositol polyphosphates that modulate distinct nuclear processes. The mammalian PLC␤ 1 , which localizes in the nucleus, is activated in insulin-like growth factor 1-mediated mitogenesis and undergoes down-regulation during murine erythroleukemia differentiation. PLC␤ 1 exists as two polypeptides of 150 and 140 kDa generated from a single gene by alternative RNA splicing, both of them containing in the COOH-terminal tail a cluster of lysine residues responsible for nuclear localization. These clues prompted us to try to establish the critical nuclear target(s) of PLC␤ 1 subtypes in the control of cell cycle progression. The results reveal that the two subtypes of PLC␤ 1 that localize in the nucleus induce cell cycle progression in Friend erythroleukemia cells. In fact when they are overexpressed in the nucleus, cyclin D3, along with its kinase (cdk4) but not cyclin E is overexpressed even though cells are serum-starved. As a consequence of this enforced expression, retinoblastoma protein is phosphorylated and E2F-1 transcription factor is activated as well. On the whole the results reveal a direct effect of nuclear PLC␤ 1 signaling in G 1 progression by means of a specific target, i.e. cyclin D3/cdk4.It is demonstrated that an autonomous intranuclear inositide cycle exists and that nuclear PLC␤ 1 1 is a key enzyme for cell proliferation and differentiation (1). The enzymes of polyphosphoinositide turnover occur in the nucleus of mammalian cells and yeast as well (Ref. references therein),and there is evidence for phosphatidylinositol bisphosphate (PIP 2 ) synthesis and degradation in the nuclear matrix (3). The evidence obtained with confocal and electron microscope analysis indicates that enzymes required for the synthesis and hydrolysis of phosphoinositides are localized at ribonucleoprotein structures of the inner nuclear matrix involved in transcript processing within the interchromatin domains (4). Although phosphatidylinositol cycle is activated only in nuclei from HeLa cells in S phase (5), striking changes occur mainly in PLC␤ 1 activity a few minutes after growth factor stimulation (1). PLC␤ 1 is composed of two subtypes, 150-kDa PLC␤ 1 a and 140-kDa PLC␤ 1 b, that are derived from a single gene by alternative RNA splicing (6). The two forms of the PLC␤ 1 are detectable both in cytosolic and nuclear fractions although PLC␤ 1 b exists almost entirely in the nucleus (7), and the ␤ 1 a form localizes in equal amount in nuclei and plasma membrane (8). Previous investigations from our group have demonstrated that the nucleus-confined PLC␤ 1 is directly involved in maintaining the undifferentiated state of Friend erythroleukemia cells even in the presence of inducers of erythroid differentiation, possibly due to a continuous stimulation of the cell cycle (9). With the above in min...

Synergistic induction of PI-PLCβ1 signaling by azacitidine and valproic acid in high-risk myelodysplastic syndromes

“…19 Furthermore, our results showed that the treatment with AZA and VPA results in an increase in Cyclin D3 expression. Cyclin D3 is a downstream target of PI-PLCb1 signaling 22,23 and has a unique function in early hematopoietic differentiation. Indeed, the data presented here prove that AZA alone, but above all the combination of AZA and VPA, can increase the levels of Cyclin D3, thus hinting at an activation of PI-PLCb1 signaling in HL60 cells exposed to demethylating agents.…”

“…Given that PI-PLCb1 targets Cyclin D3, 22,23 the effects of the combination of AZA and VPA on Cyclin D3 expression were also investigated. As Figure 2 shows, gene expression analyses, western blot experiments and immunocytochemical investigations confirmed the specific induction of Cyclin D3 expression in HL60 cells treated with AZA, which was enhanced in cells receiving the combination of AZA and VPA, but not in HL60 cells exposed to VPA alone, nor in U937 cell line ( Figure 2).…”

“…18,19 Lipid signaling in disease is an important field of investigation, and phosphoinositides have been demonstrated to be involved in leukemogenesis. 20,21 Namely, phosphoinositide-phospholipase C (PI-PLC) b1 is a key enzyme acting on cell proliferation and differentiation, mainly targeting Cyclin D3, 22,23 which is involved in G1 progression, and, more importantly, has been demonstrated to be highly expressed in the early stages of the hematopoietic differentiation. 24,25 PI-PLCb1 has also been demonstrated to have a role both in the genetic 26 and in the epigenetic mechanisms of MDS progression to AML.…”

Nuclear phospholipid signaling: phosphatidylinositol-specific phospholipase C and phosphoinositide 3-kinase

“…In addition, there are two splice variants of the PI-PLC-β 1 isoform, PI-PLC-β 1a and PI-PLC-β 1b , and several studies demonstrate that b splicing variant is the one that is predominantly nuclear (4,25,40,47). The "classical" PI-PLC-β, i.e.…”

“…The majority of PI-PLC-β 1 is probably not located in the nuclear envelope as the activity of PI-PLC-β 1 is present in both whole nuclei (21,40) and nuclei treated with different concentrations of detergents in order to remove the nuclear membrane (21,25,48,68,75,77). In rat liver nuclei, fractionation experiments indicated that both PI-PLC-β 1 and PI-PLC-γ 1 persisted in nuclear matrix and lamina, obtained after nuclease digestion and extraction with high salt and detergent (6).…”