Rapamycin Inhibiting Jurkat T Cells Viability Through Changing MRNA Expression Of Serine/Threonine Protein Phosphatase2A

Introduction:Rapamycin is a relatively new immunosuppressant. It inhibits the mammalian target of rapamycin (mTOR) which is a central controller of cell proliferation, growth and survival. In response to growth factors and nutrients, mTOR regulates translation initiation and cell cycle by affecting translational regulators. Rapamycin could cause G1arrest of T cells and block G1to S phase transition in other cell types by inhibiting mTOR signaling.PP2A, a widespread serine/threonine (Ser/Thr) protein phosphatase, plays important roles in the control of cell cycle, proliferation, transformation, apoptosis and metabolic pathways. Its heterotrimeric holoenzyme is composed of a catalytic subunit (PP2Ac), structural A subunit (also termed PR65), and members of the regulatory B subunit families, such as B, B, B, and B. The phosphatase activity of PP2Ac is modulated by its association with PP2A-A, and-B regulatory subunits and tightly regulated at the translational or posttranslational level including phosphorylation and methylation. Inhibition of mTOR with rapamycin could induce the PP2A related phosphorylation/dephosphrylation alteration of S6K1,4E-BP1, JNK-1and Erk1/2. PP2A has been identified as a component of mTOR signaling pathway.The influence of rapamycin on PP2A enzyme activity has been a topic of great interest. In contrast, quite less attention has been given to the expression of PP2A. Now we want to define if rapamycin also has an effect on the transcriptional expressions of PP2A in the process of rapamycin acting on cell viability. The data derived from these studies have implications for understanding the regulation and function of PP2A during the mTOR signaling.Aims:In this study, we analyzed the mRNA expression of serine/threonine (Ser/Thr) protein phosphatase2A (PP2A) in the human leukemic T-cell line Jurkat cells treated with rapamycin, to determine whether rapamycin inhibiting cell viability is accompanied with the change of mRNA expression of PP2A.Methods and results:Jurkat cells were incubated with various concentrations of rapamycin and cultured for different hours. Cell viability was assessed by MTT assay. The mRNA expressions of PP2A subunits were measured by quantitative real-time polymerase chain reaction (PCR). We found that rapamycin had an inhibitory effect on cell viability. IC50was343.3nM at48h.We also found rapamycin had a dose and time-dependent effect on the gene expression of PP2A. When setting the concentration of rapamycin500nM, the mRNA expressions of PP2A subunits were declined significantly at48h. When treated with various concentrations of rapamycin for48h, the mRNA expressions of PP2A subunits were down-regulated in the range from10nM to500nM.Conclusions:rapamycin inhibiting Jurkat T cells viability may be related to the reduction of PP2A mRNA expressions.