Aguado-Velasco C, Véron M, Rambow J A, Kuczmarski E R
Department of Physiology and Biophysics, Finch University of Health Sciences, Chicago Medical School, Illinois 60064, USA.
Cell Motil Cytoskeleton. 1996;34(3):194-205. doi: 10.1002/(SICI)1097-0169(1996)34:3<194::AID-CM3>3.0.CO;2-A.
Extraction of Dictyostelium amoebae with Triton X-100 produces robust cytoskeletons composed mainly of actin and myosin II. These cytoskeletons rapidly contract when mixed with Mg-ATP in simple buffers. The Triton-soluble fraction was found to contain a GTP-dependent activity that prevented contraction by Mg-ATP. This activity was purified, and identified, as nucleoside diphosphate kinase (NDP kinase). The apparent inhibition resulted from pre-contraction of the cytoskeletons. Tightly bound cytoskeletal ADP was presumably phosphorylated, and the resulting ATP powered contraction. NDP kinase appeared to be unique in this capacity, since other regenerating systems did not cause pre-contraction. Reconstitution experiments demonstrated that the kinase must be in physical contact with the cytoskeleton. These results suggest that Dictyostelium NDP kinase is able to channel ATP to the myosin molecule, and this could play a role in directly regulating cytoskeletal contraction or in facilitating contraction under conditions where intracellular ATP concentrations are low. This ability to modulate cytoskeletal contraction could help to explain observations in other systems whereby defects in NDP kinase result in abnormal development or changes in the metastatic potential of cancer cells.
用Triton X - 100提取盘基网柄菌变形虫可产生主要由肌动蛋白和肌球蛋白II组成的坚固细胞骨架。这些细胞骨架在简单缓冲液中与Mg - ATP混合时会迅速收缩。发现Triton可溶部分含有一种GTP依赖性活性，可阻止Mg - ATP引起的收缩。该活性经纯化后被鉴定为核苷二磷酸激酶（NDP激酶）。这种明显的抑制作用是由细胞骨架的预收缩引起的。紧密结合的细胞骨架ADP大概被磷酸化，产生的ATP为收缩提供动力。NDP激酶在这种能力方面似乎是独特的，因为其他再生系统不会引起预收缩。重组实验表明，激酶必须与细胞骨架有物理接触。这些结果表明，盘基网柄菌NDP激酶能够将ATP输送到肌球蛋白分子，这可能在直接调节细胞骨架收缩或在细胞内ATP浓度低的条件下促进收缩中发挥作用。这种调节细胞骨架收缩的能力有助于解释在其他系统中的观察结果，即NDP激酶的缺陷会导致异常发育或癌细胞转移潜能的变化。
