蛋白激酶C对鼻咽癌细胞P21^ras表达的影响

EFFECT OF PROTEIN KINASE C ON P21 ras EXPRESSION OF NASOPHARYNGEAL CARCINOMA CELL LINE

使用蛋白激酶Ｃ（ＰＫＣ）抑制剂与抗Ｐ２１Ｈ－ｒａｓ单抗，通过免疫细胞化学，双参数流式细胞仪等方法，探讨鼻咽癌细胞ＣＮＥ－２Ｚ中ＰＫＣ与Ｈ－ｒａｓ表达的关系。结果：１）Ｐ２１ｒａｓ蛋白在ＣＮＥ－２Ｚ细胞主要在胞膜（２７％）与胞浆（９３．７％）表达。作用于ＰＫＣ调节区的抑制剂ｓｐｈｉｎｇｏｓｉｎｅ（ＳＳ）（２×１０－５ｍｏｌ／Ｌ）与作用于催化区的ｓｔａｕｒｏｓｐｏｒｉｎｅ（ＳＴ）（２×１０－６ｍｏｌ／Ｌ）在明显抑制细胞生长的浓度使Ｐ２１ｒａｓ表达程度均减弱，胞膜阳性率明显降低，分别为５．６％与１．６％。２）蛋白质－ＤＮＡ双参数流式细胞法测定发现在Ｇ１期细胞群Ｒａｓ蛋白表达差异较大，提示Ｒａｓ蛋白主要在Ｇ１期合成逐渐增多；经ＰＫＣ抑制剂ＳＳ与ＳＴ处理后，Ｇ１期细胞表达的差异减小，说明ＰＫＣ抑制剂抑制了Ｇ１期Ｒａｓ蛋白合成。本研究结果提示：ＰＫＣ对Ｒａｓ蛋白向发挥作用的部位胞膜的转移具有一定的调控作用。Ｒａｓ蛋白的表达可能与ＣＮＥ－２Ｚ细胞Ｇ１／Ｓ期的过渡有关。

Protein kinase C (PKC) plays a key role in modulating the growth of human poorly-differentiated nasopharyngeal carcinoma (NPC) cell line (CNE 2Z). To further investigate its mechanism, the effect of PKC on the p21 ras expression of CNE 2Z was studied by using the potent inhibitors of PKC and monoclone antibody of p21 ras . The results showed that p21 ras located in cellular membrane (positive rate 27%) and cytosol (positive rate 93 7%), and the rate of expression in membrane decreased to 5 6% and 1 6% respectively after treated with sphingosine (SS) (2×10 5 mol/L) which is a potent inhibitor acting on the modulating domain of PKC, and staurosporine (ST) (2×10 -6 mol/L) which is a potent inhibitor acting on the catalytic domain of PKC. By using dual parameter flow cytometer to measure cellular DNA and protein content, the expression of p21 ras varied greatly in the cells of G1 phase, indicating that protein synthesis of p21 ras occurred mainly in G1 phase. The difference p21 ras expression among cells became smaller when treated with ST or SS at the concentration of inhibiting CNE 2Z cell growth. These findings suggest that PKC may modulate the translocation of p21 ras to the membrane and that the effect of PKC on the expression of p21 ras in G1 phase may be one of the mechanisms in which PKC modulates CNE 2Z cell growth.