Mature-fruit abscission (MFA) is a genetically controlled process, through poorly characterized mechanisms in fleshy fruit that include extensive transcriptional changes. While global transcriptome analyses have used to immature-fruit abscission in fleshy-fruit, no global gene-expression changes specific to MFA have been described. Here we use pyrosequencing to characterize the transcriptomes of the olive abscission-zone (AZ) during cell separation in order to understand MFA control at a stage of AZ-activation. Analysis of gene expression from these AZs reveal that membrane microdomains involving sterols/sphingolipids and remorins together with signaling proteins are potentially involved in MFA. This is accompanied by gene activity related to sphingolipid turnover, suggesting potentially the involvement of long-chain base metabolism in regulating MFA. Activation of vesicle trafficking involving small-GTPases is probably required for cell-wall modifications during abscission. Analysis of transcription-factors indicates that most members of MYB and bZIP families are abundantly represented in fruit-AZ, and it is consistent with a model by which most of key transcription-factors during abscission may regulate downstream processes mostly related to abscisic acid. The data provide the first thorough analysis available for a comprehensive picture of the array of cellular responses controlled by gene-expression that lead to MFA in fleshy-fruit.