Immunocytochemical and molecular analyses reveal that the disassembly of the cell wall may be mediated by changes in the level and subcellular location of extensin protein and hemicelluloses during olive-fruit abscission. Although cell-wall modification is believed to underlie the changes in organ abscission, information concerning the changes in cell-wall proteins and hemicellulose polysaccharides is still limited. The aim of this work was to analyze the spatio–temporal patterns of the distribution of different extensin proteins and hemicelluloses in the abscission zone (AZ) during natural ripe-fruit abscission in olive (Olea europaea L.). In this study, we employed immunogold labeling in the ripe-fruit AZ during olive AZ cell separation, using an expanded set of monoclonal antibodies that recognize different types of hemicelluloses (LM11, LM15, and LM21), callose (anti-(1,3)-β-d-glucan) and extensin (JIM19) epitopes, and transmission electron microscopy imaging. Our data demonstrate that AZ cell separation was accompanied by a loss of the JIM19 extensin epitopes and a reduction in the detection of the LM15 xyloglucan epitopes in AZ cell walls, whereas AZ cells were found to be enriched with respect to the xylan and callose levels of the cell wall during olive ripe-fruit abscission. By contrast, AZ cell-wall polysaccharide remodeling did not involve mannans. Moreover, in ripe-fruit AZ, quantitative RT-PCR analysis revealed that OeEXT1, OeEXT2, OeXTH9, and OeXTH13 genes were downregulated during abscission, whereas the expression of OeXTH1, OeXTH5, and OeXTH14 genes increased during abscission. Taken together, the results indicate that AZ cell-wall dynamics during olive ripe-fruit abscission involves extensin protein and hemicellulose modifications, as well as related expressed genes. This is the first study available demonstrating temporal degradation of extensin protein and hemicelluloses in the AZ at the subcellular level.

中文翻译：

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橄榄果实脱落过程中伸长蛋白和半纤维素多糖的时空免疫定位

免疫细胞化学和分子分析表明，细胞壁的分解可能是由橄榄果实脱落过程中伸长蛋白和半纤维素的水平和亚细胞位置的变化介导的。尽管细胞壁修饰被认为是器官脱落变化的基础，但有关细胞壁蛋白和半纤维素多糖变化的信息仍然有限。这项工作的目的是分析橄榄（Olea europaea L.）自然成熟果实脱落过程中不同伸长蛋白和半纤维素在脱落区（AZ）分布的时空模式。在这项研究中，我们在橄榄 AZ 细胞分离过程中在成熟水果 AZ 中使用了免疫金标记，使用了一组扩展的单克隆抗体，可识别不同类型的半纤维素（LM11、LM15、和 LM21)、胼胝质 (抗 (1,3)-β-d-葡聚糖) 和伸长蛋白 (JIM19) 表位，以及透射电子显微镜成像。我们的数据表明，AZ 细胞分离伴随着 JIM19 延伸蛋白表位的丢失和 AZ 细胞壁中 LM15 木葡聚糖表位的检测减少，而发现 AZ 细胞在木聚糖和胼胝质水平方面富集。橄榄成熟果实脱落时的细胞壁。相比之下，AZ 细胞壁多糖重塑不涉及甘露聚糖。此外，在成熟果实 AZ 中，定量 RT-PCR 分析显示 OeEXT1、OeEXT2、OeXTH9 和 OeXTH13 基因在脱落过程中下调，而 OeXTH1、OeXTH5 和 OeXTH14 基因的表达在脱落过程中增加。综合起来，结果表明，橄榄成熟果实脱落过程中的AZ细胞壁动力学涉及伸长蛋白和半纤维素修饰，以及相关的表达基因。这是第一个在亚细胞水平上证明 AZ 中伸长蛋白和半纤维素随时间降解的研究。