Temesvari L A, Bush J M, Peterson M D, Novak K D, Titus M A, Cardelli J A
Department of Microbiology and Immunology, Louisiana State University Medical Center, Shreveport 71130, USA.
J Cell Sci. 1996 Mar;109 ( Pt 3):663-73. doi: 10.1242/jcs.109.3.663.
The role of myosin Is in endosomal trafficking and the lysosomal system was investigated in a Dictyostelium discoideum myosin I double mutant myoB-/C-, that has been previously shown to exhibit defects in fluid-phase endocytosis during growth in suspension culture (Novak et al., 1995). Various properties of the endosomal pathway in the myoB-/C- double mutant as well as in the myoB- and myoC- single mutants, including intravesicular pH, and intracellular retention time and exocytosis of a fluid phase marker, were found to be indistinguishable from wild-type parental cells. The intimate connection between the contractile vacuole complex and the endocytic pathway in Dictyostelium, and the localization of a myosin I to the contractile vacuole in Acanthamoeba, led us to also examine the structure and function of this organelle in the three myosin I mutants. No alteration in contractile vacuole structure or function was observed in the myoB-, myoC- or myoB-/C- cell lines. The transport, processing, and localization of a lysosomal enzyme, alpha-mannosidase, were also unaltered in all three mutants. However, the myoB- and myoB-/C- cell lines, but not the myoC- cell line, were found to oversecrete the lysosomal enzymes alpha-mannosidase and acid phosphatase, during growth and starvation. None of the mutants oversecreted proteins following the constitutive secretory pathway. Two additional myosin I mutants, myoA- and myoA-/B-, were also found to oversecrete the lysosomally localized enzymes alpha-mannosidase and acid phosphatase. Taken together, these results suggest that these myosins do not play a role in the intracellular movement of vesicles, but that they may participate in controlling events that occur at the actin-rich cortical region of the cell. While no direct evidence has been found for the association of myosin Is with lysosomes, we predict that the integrity of the lysosomal system is tied to the fidelity of the actin cortex, and changes in cortical organization could influence lysosomal-related membrane events such as internalization or transit of vesicles to the cell surface.
在盘基网柄菌肌球蛋白I双突变体myoB - /C - 中研究了肌球蛋白在胞内体运输和溶酶体系统中的作用，先前已表明该突变体在悬浮培养生长期间的液相内吞作用中表现出缺陷（诺瓦克等人，1995年）。发现myoB - /C - 双突变体以及myoB - 和myoC - 单突变体中胞内体途径的各种特性，包括囊泡内pH值、液相标记物的细胞内保留时间和胞吐作用，与野生型亲代细胞没有区别。盘基网柄菌中收缩泡复合体与内吞途径之间的紧密联系，以及肌球蛋白I在棘阿米巴中定位于收缩泡，促使我们也研究了这三种肌球蛋白I突变体中该细胞器的结构和功能。在myoB - 、myoC - 或myoB - /C - 细胞系中未观察到收缩泡结构或功能的改变。溶酶体酶α - 甘露糖苷酶的运输、加工和定位在所有三个突变体中也未改变。然而，发现myoB - 和myoB - /C - 细胞系，但不是myoC - 细胞系，在生长和饥饿期间会过度分泌溶酶体酶α - 甘露糖苷酶和酸性磷酸酶。没有一个突变体通过组成型分泌途径过度分泌蛋白质。另外两个肌球蛋白I突变体myoA - 和myoA - /B - 也被发现过度分泌定位于溶酶体的酶α - 甘露糖苷酶和酸性磷酸酶。综上所述，这些结果表明这些肌球蛋白在囊泡的细胞内运动中不发挥作用，但它们可能参与控制发生在细胞富含肌动蛋白的皮质区域的事件。虽然尚未找到肌球蛋白I与溶酶体相关联的直接证据，但我们预测溶酶体系统的完整性与肌动蛋白皮质的保真度相关，皮质组织的变化可能影响与溶酶体相关的膜事件，如囊泡内化或向细胞表面的转运。
