Zhang Y, Kunkel J G
Biology Department, University of Massachusetts, Amherst 01003.
Dev Biol. 1994 Feb;161(2):513-21. doi: 10.1006/dbio.1994.1049.
A high concentration of calmodulin (CaM) appears in mid- to late vitellogenic cockroach follicles which composes 1.5% of total protein. CaM levels during oogenesis were estimated by densitometric analysis of immunoblots using anti-Blattella germanica egg CaM antibody as a probe. CaM accumulates in the follicle throughout the yolk deposition phase maintaining its highest level of accumulation during the later 6-fold increase in oocyte volume. Evidence suggests that this later accumulated CaM is synthesized by the follicle cells and deposited in the oocyte. In vitro experiments with [35S]-met showed that the highly abundant CaM accumulating in vitellogenic follicles may not all be synthesized by the oocyte. Isolated follicle cells incorporate 13-fold more [35S]-met into CaM than the oocytes themselves but do not accumulate the product. The follicle cells are capable of producing all the CaM observed in newly ovulated eggs. No CaM is detectable in transit in the hemolymph of the female. These facts argue that CaM produced by follicle cells is the most likely source of CaM in the vitellogenic oocyte. Indirect immunofluorescent staining with anti-egg CaM demonstrated that in early- and midvitellogenic follicles CaM is localized in the cytoplasm of follicle cells and the cytoplasmic compartment surrounding yolk granules of oocytes but is excluded from yolk granules. Immunofluorescence was most intense in the cortex of the oocyte and outside the membranes of yolk granules. Transport of CaM into the cytoplasmic compartment of the oocyte is possible without invoking traditional adsorptive endocytosis.
在卵黄生成中期到后期的蟑螂卵泡中出现高浓度的钙调蛋白(CaM),其占总蛋白的1.5%。使用抗德国小蠊卵CaM抗体作为探针,通过免疫印迹的光密度分析来估计卵子发生过程中的CaM水平。在整个卵黄沉积阶段,CaM在卵泡中积累,在卵母细胞体积后期增加6倍期间保持其最高积累水平。有证据表明,这种后期积累的CaM是由卵泡细胞合成并沉积在卵母细胞中的。用[35S]-甲硫氨酸进行的体外实验表明,在卵黄生成卵泡中积累的大量CaM可能并非全部由卵母细胞合成。分离的卵泡细胞将[35S]-甲硫氨酸掺入CaM的量比卵母细胞本身多13倍,但不会积累产物。卵泡细胞能够产生新排卵子中观察到的所有CaM。在雌性的血淋巴中未检测到转运中的CaM。这些事实表明,卵泡细胞产生的CaM是卵黄生成卵母细胞中CaM最可能的来源。用抗卵CaM进行的间接免疫荧光染色表明,在卵黄生成早期和中期的卵泡中,CaM定位于卵泡细胞的细胞质以及围绕卵母细胞卵黄颗粒的细胞质区域,但不包括在卵黄颗粒中。免疫荧光在卵母细胞的皮质和卵黄颗粒膜外最为强烈。CaM有可能在不涉及传统吸附性内吞作用的情况下转运到卵母细胞的细胞质区域。