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cDNAs encoding Hypericum sampsonii benzophenone synthase (HsBPS) and chalcone synthase (HsCHS) were isolated and functionally characterized. Differential expressions of HsBPS and HsCHS were monitored using quantitative polymerase chain reaction (PCR). In the vegetative stage, HsBPS was highly expressed in the roots; its transcript level was approx. 100 times higher than that of HsCHS. Relatively high transcript amounts of HsBPS were also detected in older leaves, whereas the youngest leaves contained higher transcript amounts of HsCHS. In the reproductive stage, maximum HsCHS expression was detected in flowers, the transcript level being approx. 5 times higher than that of HsBPS. The inversed situation with a 10-fold difference in the expression levels was observed with fruits. High transcript amounts for both proteins were found in roots.

Abstract 68 Several groups have shown that that the BCR-ABL1 transcript level measured at 3 or 6 months after starting TKI therapy strongly predicts for the achievement of cytogenetic and molecular responses and for PFS and OS. In particular, we have shown that CML patients treated with imatinib who at 3 months have a transcript level lower than 9.8% on the international scale or lower than 1.67% at 6 months fare significantly better. We have also shown that the molecular assessment made at 3 months on imatinib therapy is a better predictor of the prognosis of patients than the analysis of BCR-ABL1 transcripts at 6 months. Here we investigate whether it is possible to improve the prognostic accuracy of early measurement of the transcript level by combining the 3 and 6 month results. Between June 2000 and December 2010 282 consecutive adult patients with CML in CP seen at our institution received imatinib 400 mg daily as first line therapy. The median follow-up was 69 months (range 17–131). During follow-up 118 patients discontinued imatinib and received nilotinib (n=37), dasatinib (n=72) or an allogeneic stem cell transplant (n=9). BCR-ABL1 transcripts were measured in the blood at 6 to 12 week intervals using RQ-PCR and results were expressed as percent ratios relative to an ABL1 internal control with original laboratory values converted to the international scale. Two hundred and seventy-four patients were still alive in chronic phase and receiving imatinib at 6 months. We classified these patients according to their transcript levels at 3 months (lower or higher than 9.8%) and 6 months (lower or higher than 1.67%). 181 (66%) patients had low transcripts both at 3 and 6 months; these patients had an excellent outcome with a OS of 93.5% and a 100% cumulative incidence (CI) of CCyR. Fifty-seven (21%) of the patients had high transcript levels on both occasions; these patients had a significantly worse outcome than the previous cohort, namely an OS of 55.6% (p The measurement of the transcript level at 6 months adds very little prognostic discrimination to the measurement already taken at 3 months. The 11% of patients who met the three month milestone but failed the 6 month milestone had an OS and PFS identical to the patients who achieved both milestones. The CI of CCyR was also similar (although slower, median time to CCyR 12 months vs 6 months for the patients who met both milestones, p=0.001). The 2% of patients who failed the first milestone at 3 months but who met the second one at 6 months also seemed to fare well, although this group is too small to clearly establish whether patients with high 3 month transcript levels are ‘rescued’ by meeting the 6 month milestone. In summary, the prognosis of patients can be accurately established by assessing the transcript level purely at 3 months, although the analysis at 6 months may improve the prognostic classification of 2% of the patients. Disclosures: Goldman: Novartis, Bristol Myers-Squibb, and Amgen: Honoraria. Marin:BMS: Research Funding; Novartis: Research Funding.

The Sanger Institute in the UK has an ongoing initiative termed the “Cancer Genome Project” to systematically characterize the genetics and genomics of 770 cancer cell lines. Similarly, the Broad Institute in the US collaborates on the “Cancer Cell Line Encyclopedia” or CCLE project, entailing the eventual characterization of approximately 1000 cancer cell lines. At present, there are over 400 cell lines shared by these two initiatives. As part of a number of computational efforts in cancer biology we have used data from these two programs and have observed some unexpected differences in cell lines at the DNA and/or transcript level as well as some phenotypic differences, and we present here an analysis of these results including implications to the use of cancer cell lines in drug discovery and the larger question of genomic stability in cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5109. doi:1538-7445.AM2012-5109

Soduim butyrate decreases the c-myc mRNA and increases the c-fos transcript level in HTC cells. This effect is independent of the cell-cycle phase. Actinomycin D suppresses the effect on c-fos. Cycloheximide increases both mRNA levels. Sodium butyrate suppresses the effect on c-myc and potentializes the effect on c-fos mRNAs. This suggests that sodium butyrate acts at the transcriptional level and that its effect does not result from the arrest of the cells at the G1 phase.

In September 2003, the French CML study group (Fi-LMC) activated the four armed randomized SPIRIT trial comparing imatinib 400mg with imatinib 600mg, imatinib 400mg + ara-C and imatinib 400mg + pegylated interferon in newly diagnosed CML in chronic phase. The molecular monitoring was centralized and performed, according to the EAC protocol, in duplicate on the same BCR-ABL cDNA among two French laboratories (Bordeaux and Lille). Data were expressed as BCR-ABL/ABLx100 on the new internationally agreed scale (IS) using a conversion factor of 1.33 and 0.80 for Bordeaux and Lille respectively. A good correlation between the two laboratories was observed. During the first year of follow-up the quantification was performed each three months (M0 to M12). In May 2006, 297 of the 492 enrolled pts had a follow up of more than 12 months, and 263 pts were analysed at M12. Using the IS, the median values of the BCR-ABL/ABL normalized ratios were 90%, 6,4%, 0.68%, 0.32% and 0.16% at M0, M3, M6, M9, and M12 respectively. At M12, 15%, 29%, 27%, 17% and 12% of the pts presented a BCR-ABL/ABL ratio lower than 0.01%, 0.1%, 1%, 10%, and 100% respectively. Overall, 44% of the pts reached the Major Molecular Response (MMR). Results were also expressed using individual M12/M0 ratio. At M12 13%, 29%, 28%, 17% and 13% of the pts presented a 4 log, 3 log, 2 log, 1 log and 0 log reduction respectively. Overall, 42% of the pts presented an individual 3 log reduction. Despite very similar results between the two means of expression, the pts distribution was different due to unlike individual BCR-ABL transcript level at MO. Furthermore, the analysis of 75 paired samples before (at diagnosis) and after (at inclusion) hydroxyurea treatment (5 days to 1.2 months) revealed BCR-ABL/ABL ratios of 129% and 108% respectively, confirming the 0.2 log difference previously reported by A. Hochhaus et al. In conclusion, expression of the data using the new IS or the individual M12/M0 ratio was very similar in our experience regarding the percentage of each subgroup but the pts distribution was different and a longer follow up would be useful to define the best method. However, the good correlation observed in the IRIS trial between MMR and progression, the influence of hydroxyurea treatment on BCR-ABL level, the good correlation observed among laboratories and the need of harmonisation of expressed results influenced our choice for the new IS. The application of this recommendation is in progress in the great majority of the molecular French laboratories.

Previous studies in our laboratory both in pine needles and potato leaves have shown evidence of an increase in 18: 2 (linoleate) in the purified plasma membrane fraction during cold acclimation. This increase was reversible on deacclimation, thereby suggesting a link between the accumulation of 18: 2 and acquisition of freezing tolerance. These studies suggest that the activity of specific desaturases may be modulated during cold acclimation. This study was aimed at studying the possible involvement of stearoyl-ACP desaturase (delta9) in potato cold acclimation response. Our approach was to study the induction of delta9 desaturase at the transcript level by using potato delta9 desaturase gene specific primers and reverse transcriptase. For this purpose, mRNA from S. tuberosum (cold sensitive, unable to acclimate) and S. commersonii (cold tolerant, able to cold acclimate) was extracted before and after acclimation. Sequence analysis confirmed that the amplified band was delta9 desaturase. Our results show that there is an increase in delta9 desaturase gene transcripts during cold acclimation and that this increase is associated with the cold acclimation response in potato. These results together with previous reports on the increase in 18: 2 in the plasma membrane during cold acclimation give more evidence toward the involvement of stearoyl-ACP desaturase (delta9) in the potato cold response.

It has been a matter of controversy as to whether c-myc gene expression is activated in human hepatocellular carcinoma (HCC). We observed that the c-myc mRNA level in HCC was similar to that of the adjacent non-cancerous portion, as determined in freshly obtained specimens after a partial liver resection. However, the c-myc transcript was at a low level in non-cancerous tissue which was biopsied prior to surgery, whereas it was still at a high level in HCC obtained without performing hepatectomy. These results suggest that the high transcript level observed in the non-cancerous tissue from the excised liver relates to liver resection itself, and that the c-myc gene expression is enhanced in the HCC.

Background Invertase Suc2 was recently identified as a key hydrolase for inulin catabolism in Saccharomyces cerevisiae, whereas the Suc2 activity degrading inulin varies greatly in different S. cerevisiae strains. The molecular mechanism causing such variation remained obscure. The aim of this study is to investigate how Suc2 activity is regulated in S. cerevisiae. Results The effect of SUC2 expression level on inulin hydrolysis was investigated by introducing different SUC2 genes or their corresponding promoters in S. cerevisiae strain BY4741 that can only weakly catabolize inulin. Both inulinase and invertase activities were increased with the rising SUC2 expression level. Variation in the promoter sequence has an obvious effect on the transcript level of the SUC2 gene. It was also found that the high expression level of SUC2 was beneficial to inulin degradation and ethanol yield. Conclusions Suc2-mediated inulin catabolism is regulated at transcript level in S. cerevisiae. Our work should be valuable for engineering advanced yeast strains in application of inulin for ethanol production. Electronic supplementary material The online version of this article (doi:10.1186/s12934-015-0243-3) contains supplementary material, which is available to authorized users.

The discovery of genes that can be used to increase plant tolerance to environmental stress has practical implications for agriculture, since knowledge at the molecular level can potentially be translated from model plants to crops or from tolerant to sensitive cultivars. For more than a decade, researchers have attempted to identify transcriptional and metabolic pathways involved in stress tolerance using functional genomics tools. In some cases, promising results were obtained when a clear causal link was found between transcripts and tolerance/sensitivity to stress. However, recent reports question the global translational power of functional genomics for biotechnological applications, as one of the main limitations seems to be the large variability in gene expression. Transcript-level variability under stress has not been considered of interest in the scientific literature because it is intuitively obvious, but most reports seem to naively overlook the consequences. Here, three case situations are reviewed (variability between genotypes, variability due to environmental interactions and variability between stressors) in support of the concept that inherent transcript-level variation in biological systems may limit our knowledge of environmental plant tolerance and of functional genomics in molecular stress physiology.

Background Natural accessions of Arabidopsis thaliana are a well-known system to measure levels of intraspecific genetic variation. Leaf starch content correlates negatively with biomass. Starch is synthesized by the coordinated action of many (iso)enzymes. Quantitatively dominant is the repetitive transfer of glucosyl residues to the non-reducing ends of α-glucans as mediated by starch synthases. In the genome of A. thaliana, there are five classes of starch synthases, designated as soluble starch synthases (SSI, SSII, SSIII, and SSIV) and granule-bound synthase (GBSS). Each class is represented by a single gene. The five genes are homologous in functional domains due to their common origin, but have evolved individual features as well. Here, we analyze the extent of genetic variation in these fundamental protein classes as well as possible functional implications on transcript and protein levels. Findings Intraspecific sequence variation of the five starch synthases was determined by sequencing the entire loci including promoter regions from 30 worldwide distributed accessions of A. thaliana. In all genes, a considerable number of nucleotide polymorphisms was observed, both in non-coding and coding regions, and several amino acid substitutions were identified in functional domains. Furthermore, promoters possess numerous polymorphisms in potentially regulatory cis-acting regions. By realtime experiments performed with selected accessions, we demonstrate that DNA sequence divergence correlates with significant differences in transcript levels. Conclusions Except for AtSSII, all starch synthase classes clustered into two or three groups of haplotypes, respectively. Significant difference in transcript levels among haplotype clusters in AtSSIV provides evidence for cis-regulation. By contrast, no such correlation was found for AtSSI, AtSSII, AtSSIII, and AtGBSS, suggesting trans-regulation. The expression data presented here point to a regulation by common trans-regulatory transcription factors which ensures a coordinated action of the products of these four genes during starch granule biosynthesis. The apparent cis-regulation of AtSSIV might be related to its role in the initiation of de novo biosynthesis of granules.