Intracellular Signal Transduction and Topological Change of Intranuclear DNA in the Differentiation of Human Leukemic HL-60 Cells.

人骨髓性HL-60白血病细胞可以被各种化学试剂诱导分化为单核细胞/巨噬细胞样细胞或粒细胞。其中包括1，25-二羟维生素D_3（1，25-（OH）_2D_3）和12-o-十四酰基佛波醇13-乙酸酯（TPA），它们可介导单核细胞分化;蛋白激酶C（PKC）的激活、转位及随后的下调是TPA诱导HL-60细胞向巨噬细胞样细胞分化的必要条件。使用PKC抑制剂（如H-7和二氢鞘氨醇）的药理学方法以及TPA耐药HL-60变体（HL-60 R）细胞的研究证明了这一点。在PKC同工酶中，PKC-β可能是TPA诱导HL-60细胞分化的关键信号，HL-60 R细胞对TPA诱导的抗性可能是由于胞浆PKC-β的选择性抑制。PKC活性和表达水平 ...更多信息 在DMSO和ATRA诱导的HL-60分化过程中，PKC-α、-β、-γ亚型的表达增加。1，25-（OH）_2D_3诱导的单核细胞分化还与PKC活性及PKC-α和-β表达的增加有关。这些结果表明，PKC的激活和/或其同工酶表达的调节在调节造血细胞对生长因子和细胞生长和分化的非生理诱导剂的反应中起关键作用，通过磷酸化调节蛋白质功能除了蛋白激酶外，还需要蛋白磷酸酶（PPs）。PPs可能参与HL-60细胞的分化过程。PP 1和PP 2A的有效抑制剂，calyculin-A（CAL-A）和冈田酸可以增强ATRA诱导的粒细胞分化，而TPA向巨噬细胞系的分化在CAL-A的存在下没有变化。ATRA处理HL-60细胞后，PP活性和PP 2A蛋白表达明显下降。ATRA处理后3 h内PP 2A mRNA水平明显下降。因此，PP 2A的下调可能参与了ATRA诱导HL-60细胞分化的过程。本研究通过观察分化诱导剂对拓扑异构酶II表达的调控，探讨蛋白磷酸化在HL-60细胞分化过程中对核DNA拓扑结构变化的调控作用。少

Human myelogenous HL-60 leukemia cells can be induced to differentiate into either monocyte/macrophage-like cells or into granulocytes by various chemical agents. These include 1,25-dihydroxyvitamin D_3(1,25-(OH)_2D_3) and 12-o-tetra-decanoylphorbol 13-acetate(TPA), which mediate monocytic differentiation, and all-trans retinoic acid(ATRA) for granulocytic differentiation. The activation, translocation and subsequent down-regulation of protein kinase C(PKC) appears to be essential for TPA-induced differentiation of HL-60 cells into macrophage-like cells. This is evidenced by pharmacological methods using PKC inhibitors such as H-7 and sphinganine, and investigations with TPA-resistant HL-60 variant(HL-60R) cells. Among PKC isozymes, PKC-beta may be crucial signal in the regulation of TPA-induced HL-60 cell differentiation, and the resistance of HL-60R cells to TPA induction may be due to the selective depression of cytosolic PKC-beta. The levels of both PKC activity and the expression … More of PKC-alpha, -beta, -gamma isoforms have been shown to increase during HL-60 differentiation induced by DMSO and ATRA. 1,25-(OH)_2D_3 induced monocytic differentiation is also associated with an in- crease in both PKC activity and the expression of PKC-alpha and -beta. These results suggest that PKC activation and/or modulation of its isoenzyme expression play key roles in regulating the responses of hematopoietic cells to both growth factors and non-physiological inducers of cell growth and differentiation.The regulation of protein function by phosphorylation necessarily requires protein phosphatases(PPs) in addition to protein kinases. It is conceivable that PPs may also be involved in the process of HL-60 cell differentiation. Potent inhibitors of PP1 and PP2A, calyculin-A(CAL-A) and okadaic acid could augment ATRA-induced granulocytic differentiation, whereas the differentiation toward macrophage lineage by TPA was unchanged in the presence of CAL-A. Treatment of HL-60 cells with ATRA led to a dramatic decrease in the activity of PP and PP2A protein expression. The mRNA level of PP2A was markedly decreased within 3 hrs after addition of ATRA. Thus, down regulation of PP2A may be involved in differentiation events of HL-60 cells induced by ATRA.We are now examining the modulation/expression of topoisomerase II induced by differentiation inducers in order to know the regulatory roles of protein phosphorylation in the topological changes of nuclear DNAs during HL-60 cell differentiation. Less