Baldan B, Guzzo F, Filippini F, Gasparian M, LoSchiavo F, Vitale A, de Vries S C, Mariani P, Terzi M
Dipartimento di Biologiá, Università, Padova, Italy.
Planta. 1997;203(3):381-9. doi: 10.1007/s004250050204.
The carrot cell variant ts11 is unable to form somatic embryos at the non-permissive temperature of 32 degrees C, but the block can be overcome by the addition of a 32-kDa acidic endochitinase to the medium. In this work we conducted a cyto-histological analysis of the blocked embryo forms. The morphology of the endomembrane system is altered; in particular, the ER is dilated and may show electron-dense precipitates and continuity with the plasma membrane. These morphological alterations do not occur in the presence of externally-added endochitinase. We also noticed modifications of the culture medium that are probably related to the morphological observations: the total amount of secreted proteins is reduced and pulse-chase experiments revealed that, compared with wild-type cells, the secretion of major polypeptides is reduced while new minor polypeptides are secreted. Western blot analysis revealed the presence of the binding protein BiP, a resident of the ER and of glutamine synthase, a cytosolic protein, in the medium of ts11 but not wild-type cells. These results indicate that ts11 is altered in the secretory pathway but do not clarify the role of endochitinase.
胡萝卜细胞变体ts11在32℃的非允许温度下无法形成体细胞胚，但通过向培养基中添加一种32 kDa的酸性内切几丁质酶可以克服这种障碍。在这项工作中，我们对受阻的胚形态进行了细胞组织学分析。内膜系统的形态发生了改变；特别是，内质网扩张，可能显示出电子致密沉淀物，并与质膜连续。在添加外源内切几丁质酶的情况下不会出现这些形态改变。我们还注意到培养基的变化，这可能与形态学观察有关：分泌蛋白的总量减少，脉冲追踪实验表明，与野生型细胞相比，主要多肽的分泌减少，同时分泌出新的次要多肽。蛋白质印迹分析显示，在ts11细胞的培养基中存在结合蛋白BiP（内质网驻留蛋白）和谷氨酰胺合成酶（一种胞质蛋白），而野生型细胞的培养基中则没有。这些结果表明ts11在分泌途径中发生了改变，但没有阐明内切几丁质酶的作用。
