Monitoring transcription initiation activities in rat and dog.

The promoter landscape of several non-human model organisms is far from complete. As a part of FANTOM5 data collection, we generated 13 profiles of transcription initiation activities in dog and rat aortic smooth muscle cells, mesenchymal stem cells and hepatocytes by employing CAGE (Cap Analysis of Gene Expression) technology combined with single molecule sequencing. Our analyses show that the CAGE profiles recapitulate known transcription start sites (TSSs) consistently, in addition to uncover novel TSSs. Our dataset can be thus used with high confidence to support gene annotation in dog and rat species. We identified 28,497 and 23,147 CAGE peaks, or promoter regions, for rat and dog respectively, and associated them to known genes. This approach could be seen as a standard method for improvement of existing gene models, as well as discovery of novel genes. Given that the FANTOM5 data collection includes dog and rat matched cell types in human and mouse as well, this data would also be useful for cross-species studies.

Linking FANTOM5 CAGE peaks to annotations with CAGEscan.

The FANTOM5 expression atlas is a quantitative measurement of the activity of nearly 200,000 promoter regions across nearly 2,000 different human primary cells, tissue types and cell lines. Generation of this atlas was made possible by the use of CAGE, an experimental approach to localise transcription start sites at single-nucleotide resolution by sequencing the 5' ends of capped RNAs after their conversion to cDNAs. While 50% of CAGE-defined promoter regions could be confidently associated to adjacent transcriptional units, nearly 100,000 promoter regions remained gene-orphan. To address this, we used the CAGEscan method, in which random-primed 5'-cDNAs are paired-end sequenced. Pairs starting in the same region are assembled in transcript models called CAGEscan clusters. Here, we present the production and quality control of CAGEscan libraries from 56 FANTOM5 RNA sources, which enhances the FANTOM5 expression atlas by providing experimental evidence associating core promoter regions with their cognate transcripts.

New spirostanol glycosides from Solanum nigrum and S. jasminoides.

A new characteristic steroidal glycoside possessing a hydroxyl group at C-23, inunigroside A (1), was isolated from the withered berries of Solanum nigrum L. On the basis of spectroscopic analysis, the structure of 1 was characterized as (5α,22S,23S,25R)-3ß,23-dihydroxyspirostane 3-O-ß-lycotetraoside. Next, a major steroidal sapogenol, (22R, 25S)-3ß,15α-dihydroxy-spirost-5-ene (3), was obtained from the acid hydrolysate of the methanolic extract of the aerial parts of Solanum jasminoides L. A new bisdesmoside, 3-O-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl (22R,25S)-3ß,15α-dihydroxyspirost-5-ene 15-O-α-L-rhamnopyranoside (4), named jasminoside A, was isolated from the methanolic extract of S. jasminoides.

New solanocapsine-type tomato glycoside from ripe fruit of Solanum lycopersicum.

A new solanocapsine-type tomato glycoside, a novel and interesting natural steroidal glycoside, was isolated from a mini tomato, Solanum lycopersicum L. The chemical structure of the new minor glycoside, esculeoside B-5 (3), was determined to be (5S,22R,23S,24R,25S)-22,26-epimino-16ß,23-epoxy-3ß,23,24-trihydroxycholestane 3-O-ß-lycotetraoside.

Efficient improved extraction of tomato saponin using shock waves.

In the conventional method of mixer blending extraction, the yields of the tomato-saponin, esculeoside A, in the mini and middy tomatoes were found to be 0.043% and 0.046%, respectively. In order to improve the yields, we attempted a more efficient extraction using shock waves. The yields of esculeoside A were 0.0987% in air after 1 shock, 0.0792% in air after two shots, 0.0648% in half water after 1 or 2 shocks, and 0.0599% in water after 1 or 2 shocks. The yields obtained by the proposed method were approximately twice those of the conventional mixer blending method; therefore, this method is regarded to be very efficient. Moreover, two spirosolane glycosides, tomatine and lycoperoside A, were obtained for the first time from the ripe tomato fruit in this method. To date, these compounds have not been obtained with the mixer blending method. However, whether these glycosides are produced from esculeoside A or are newly extracted from the plant organ by the shock wave is still unclear.