Methylthioacetic acid, a derivative of aroma compounds from Cucumis melo var. conomon dose-dependently triggers differentiation and apoptosis of RCM-1 human colorectal cancer cells.

Methylthioacetic acid (MTA) is an acid-hydrolyzed derivative of a natural aroma compound, methylthioacetic acid ethyl ester isolated from Cucumis melo var. conomon (Katsura-uri, Japanese Picking Melon), and induces a villiform-like structure dome in RCM-1 human colorectal cancer cell culture. Thus far, the physiological and molecular properties of MTA-mediated dome formation remain unknown. Herein, MTA (not more than 2 mM) was demonstrated to differentiate the unorganized cell mass into the dome in RCM-1 cell culture by disclosing the correlation between dome formation and several intestinal differentiation markers such as alkaline phosphatase activity and the protein levels of dipeptidyl peptidase 4, villin, and Krüppel-like factor 4. Dome formation in RCM-1 cell culture was additively enhanced by the simultaneous administration of MTA and butyric acid (BA), suggesting that MTA directs the differentiation of RCM-1 cells, potentially through the same or similar pathway(s) shared with BA. Notably, a high dose of MTA (2 mM or more) elevated several apoptosis markers, such as DNA fragmentation, caspase-3/7 activity, and cleavage of poly(ADP-ribose) polymerase. Altogether, in addition to RCM-1 cell differentiation, MTA triggers apoptosis. These results indicate that MTA is a potential anticarcinogenic agent applicable in differentiation therapy and traditional chemotherapy against colorectal cancers.

Behavior and possible function of Arabidopsis BES1/BZR1 homolog 2 in brassinosteroid signaling.

Two key transcription factors (TFs) in brassinosteroid (BR) signaling BRASSINOSTEROID INSENSITIVE 1-EMS-SUPPRESSOR 1 (BES1) and BRASSINAZOLE RESISTANT 1 (BZR1), belong to a small family with four BES1/BZR1 homologs (BEH1-4). To date, in contrast to the wealth of knowledge regarding BES1 and BZR1, little is known about BEH1-4. Here, we show that BEH2 was expressed preferentially in the roots and leaf margins including serrations, which was quite different from another member BEH4, and that BRs downregulated BEH2 through a module containing GSK3-like kinases and BES1/BZR1 TFs, among which BES1, rather than BZR1, contributed to this process. In addition, BEH2 consistently existed in the nucleus, suggesting that its subcellular localization is not under BR-dependent nuclear-cytoplasmic shuttling control. Furthermore, gene ontology analysis on RNA-seq data indicated that BEH2 may be implicated in stress response and photosynthesis. These findings might assist in the future elucidation of the molecular mechanisms underlying BR signaling.

Chemical and molecular bases of dome formation in human colorectal cancer cells mediated by sulphur compounds from Cucumis melo var. conomon.

Colorectal cancer was the third most commonly diagnosed malignant tumor and the fourth leading cause of cancer deaths worldwide in 2012. A human colorectal cancer cell line, RCM-1, was established from a colon cancer tissue diagnosed as a well-differentiated rectum adenocarcinoma. RCM-1 cells spontaneously form 'domes' (formerly designated 'ducts') resembling villiform structures. Two sulphur-containing compounds from Cucumis melo var. conomon (Katsura-uri, or Japanese pickling melon), referred to as 3-methylthiopropionic acid ethyl ester (MTPE) and methylthioacetic acid ethyl ester (MTAE), can induce the differentiation of the unorganized cell mass of an RCM-1 human colorectal cancer cell culture into a dome. However, the underlying molecular mechanisms of such dome formation have not been previously reported. Here, we performed a structure-activity relationship analysis, which indicated that methylthioacetic acid (MTA) was the lowest molecular weight compound with the most potent dome-inducing activity among 37 MTPE and MTAE analogues, and the methylthio group was essential for this activity. According to our microarray analysis, MTA resulted in down-regulation of 537 genes and up-regulation of 117 genes. Furthermore, MTA caused down-regulation of many genes involved in cell-cycle control, with the cyclin E2 (CCNE2) and cell division cycle 25A (CDC25A) genes being the most significantly reduced. Pharmacological analysis showed that the administration of two cell-cycle inhibitors for inactivating CDC25A phosphatase (NSC95397) and the cyclin E2/cyclin-dependent kinase 2 complex (purvalanol A) increased the dome number independently of MTA. Altogether, our results indicate that MTA is the minimum unit required to induce dome formation, with the down-regulation of CDC25A and possibly CCNE2 being important steps in this process.

Expression profiles of four BES1/BZR1 homologous genes encoding bHLH transcription factors in Arabidopsis.

Arabidopsis bHLH-type transcription factors-BRASSINOSTEROID INSENSITIVE 1-EMS-SUPPRESSOR 1 (BES1) and BRASSINAZOLE RESISTANT 1 (BZR1)-play key roles in brassinosteroid (BR) signaling. By contrast, the functions of the other four BES1/BZR1 homologs (BEH1-4) remain unknown. Here, we describe the detailed expression profiles of the BES1/BZR1 family genes. Their expressions were distinct regarding growth-stage dependence and organ specificity but exhibited some overlaps as well. Furthermore, their mRNA levels mostly remained unchanged responding to seven non-BR phytohormones. However, BEH1 and BEH2 were downregulated by brassinolide, suggesting a close association with the BR function. Additionally, BEH4 was ubiquitously expressed throughout the life of the plant but displayed some expression preference. For instance, BEH4 expression was limited to guard cells and the adjacent pavement cells in the leaf epidermis and was induced during growth progression in very young seedlings, suggesting that BEH4 is specifically regulated in certain contexts, although it is almost constitutively controlled.