BAHD acyltransferase induced by histone deacetylase inhibitor catalyzes 3-O-hydroxycinnamoylquinic acid formation in bamboo cells.

Suspension-cultured cells of a bamboo species (Bambusa multiplex; Bm) produce 3-O-feruloylquinic acid (3-FQA) and 3-O-p-coumaroylquinic acid (3-pCQA) by treatment with the histone deacetylase inhibitor suberoyl bis-hydroxamic acid (SBHA). Acyltransferases catalyzing the formation of 5-O-hydroxycinnamoylquinic acid esters by transesterification from hydroxycinnamoyl-CoAs to the C-5 hydroxy group of quinic acid (hydroxycinnamoyl-CoA:quinate hydroxycinnamoyltransferase, HQT) have been identified in the biosynthesis of chlorogenic acids and monolignols; however, an HQT that catalyzes the acylation of the C-3 hydroxy group of quinic acid has not been identified previously. In the present study, we purified a native HQT from SBHA-treated Bm cells. The purified enzyme preferentially accepted feruloyl-/p-coumaroyl-CoAs as acyl-donors and quinic acid as the acyl-acceptor, and the enzyme specifically formed 3-FQA and 3-pCQA but not 5-O-hydroxycinnamoylquinic acid esters or esters with shikimic acid. A cDNA (BmHQT1) encoding this HQT was isolated. Although BmHQT1 is a phylogenetically unique member of the BAHD acyltransferase superfamily that does not cluster with other HQTs, functional characterization of the recombinant enzyme verified that BmHQT1 catalyzes the regiospecific formation of 3-O-hydroxycinnamoylquinic acid esters. Transcript levels of BmHQT1 markedly increased in Bm cells cultured in the presence of SBHA. Moreover, elevated acetylation levels of histone H3 were observed in the coding region of BmHQT1 in the presence of SBHA, indicating that the induced accumulation of 3-FQA/3-pCQA by SBHA is caused by transcriptional activation of BmHQT1 by the action of SBHA as a histone deacetylase inhibitor. The results demonstrate the utility of HDAC inhibitors for discovery of cryptic secondary metabolites and unknown biosynthetic enzymes.

Anti-melanogenic effects of glucosylceramides and elasticamide derived from rice oil by-products in melanoma cells, melanocytes, and human skin.

Glucosylceramides (GlcCer), which are present in many edible plants, suppress melanin production in mouse melanocytes. Rice GlcCer consist of multiple molecules that comprise different types of sphingoid bases as well as diverse lengths and stereotypes of free fatty acids. Adjacent to the GlcCer fraction, there are free ceramides (Cer) as minor constituents. However, the anti-melanogenic activities of individual GlcCer and Cer remain unknown. Therefore, we herein isolated 13 GlcCer and elasticamide, a Cer [AP] from the gummy by-products of rice bran oil, and examined their anti-melanogenic activities. In theophylline-induced melanogenesis in B16 melanoma cells, GlcCer [d18:2(4E,8Z)/18:0], GlcCer [d18:2(4E,8Z)/20:0], and elasticamide significantly suppressed melanin production with IC50 values of 6.6, 5.2, and 3.9 µM, respectively. Elasticamide, but not GlcCer [d18:2 (4E,8Z)/20:0], suppressed melanogenesis in human 3D-cultured melanocytes and the expression of tyrosinase-related protein 1 in normal human melanocytes. Based on these results, we conducted a clinical trial on the effects of rice ceramide extract (Oryza ceramide®), containing 1.2 mg/day of GlcCer and 56 µg/day of elasticamide, on UV-B-induced skin pigmentation. The ingestion of Oryza ceramide® for 8 weeks significantly suppressed the accumulation of melanin 7 days after UV irradiation (1288 and 1546 mJ/cm2 ·S). Rice-derived GlcCer and elasticamide, which exhibited anti-melanogenic activities, were suggested to contribute to the suppressive effects of Oryza ceramide® on UV-induced skin pigmentation. Although the mechanisms underlying the anti-melanogenic activities of GlcCer remain unclear, elasticamide was identified as a promising Cer that exhibits anti-melanogenic activity. PRACTICAL APPLICATIONS: The anti-melanogenic activities of rice-derived GlcCer and elasticamide currently remain unclear. We herein demonstrated the inhibitory effects of individual GlcCer and elasticamide on melanogenesis in melanoma cells, melanocytes, and human skin.

Comparative Study on Epidermal Moisturizing Effects and Hydration Mechanisms of Rice-Derived Glucosylceramides and Ceramides.

Ceramide (Cer) plays an important role in skin barrier functions in the stratum corneum (SC). The ingestion of food-derived glucosylceramides (GlcCer) attenuates transepidermal water loss (TEWL). However, the moisturizing effects of single molecules of GlcCer and Cer remain unclear. Therefore, we herein purified 13 GlcCer and 6 Cer, including elasticamide, which has the same structure as human Cer[AP], from rice and compared their epidermal moisturizing effects in a reconstructed human epidermal keratinization model. The results obtained showed that 10 µM of 5 GlcCer[d18:2] with a 4E,8Z sphingadienine and C18 to C26 fatty acids and 10 µg/mL of 3 Cer with C23 or C24 fatty acids significantly reduced TEWL. The moisturizing effects of these GlcCer were dependent on the length of fatty acids. Furthermore, 10 µg/mL of elasticamide increased the SC Cer contents by promoting the expression of GlcCer synthase. Electron microscopic observations revealed that 1 µM of GlcCer[d18:2(4E,8Z)/26:0] increased the number of keratohyalin granules and desmosomes. Immunostaining and Western blotting indicated that 1 µM of GlcCer[d18:2(4E,8Z)/26:0] up-regulated the expression of filaggrin and corneodesmosin, which contribute to epidermal hydration. This comparative study on epidermal moisturization by GlcCer and Cer isolated from rice revealed differences in their hydration mechanisms.

Activation of Cryptic Secondary Metabolite Biosynthesis in Bamboo Suspension Cells by a Histone Deacetylase Inhibitor.

Plants have evolved a diverse array of secondary metabolite biosynthetic pathways. Undifferentiated plant cells, however, tend to biosynthesize secondary metabolites to a lesser extent and sometimes not at all. This phenomenon in cultured cells is associated with the transcriptional suppression of biosynthetic genes due to epigenetic alterations, such as low histone acetylation levels and/or high DNA methylation levels. Here, using cultured cells of bamboo (Bambusa multiplex; Bm) as a model system, we investigated the effect of histone deacetylase (HDAC) inhibitors on the activation of cryptic secondary metabolite biosynthesis. The Bm suspension cells cultured in the presence of an HDAC inhibitor, suberoyl bis-hydroxamic acid (SBHA), exhibited strong biosynthesis of some compounds that are inherently present at very low levels in Bm cells. Two major compounds induced by SBHA were isolated and were identified as 3-O-p-coumaroylquinic acid (1) and 3-O-feruloylquinic acid (2). Their productivities depended on the type of basal culture medium, initial cell density, and culture period, as well as the SBHA concentration. The biosynthesis of these two compounds was also induced by another HDAC inhibitor, trichostatin A. These results demonstrate the usefulness of HDAC inhibitors to activate cryptic secondary metabolite biosynthesis in cultured plant cells.