Previous investigations in gene expression changes in blood after radiation exposure have highlighted its potential to provide biomarkers of exposure. Here, FDXR transcriptional changes in blood were investigated in humans undergoing a range of external radiation exposure procedures covering several orders of magnitude (cardiac fluoroscopy, diagnostic computed tomography (CT)) and treatments (total body and local radiotherapy). Moreover, a method was developed to assess the dose to the blood using physical exposure parameters. FDXR expression was significantly up-regulated 24 hr after radiotherapy in most patients and continuously during the fractionated treatment. Significance was reached even after diagnostic CT 2 hours post-exposure. We further showed that no significant differences in expression were found between ex vivo and in vivo samples from the same patients. Moreover, potential confounding factors such as gender, infection status and anti-oxidants only affect moderately FDXR transcription. Finally, we provided a first in vivo dose-response showing dose-dependency even for very low doses or partial body exposure showing good correlation between physically and biologically assessed doses. In conclusion, we report the remarkable responsiveness of FDXR to ionising radiation at the transcriptional level which, when measured in the right time window, provides accurate in vivo dose estimates.

• MeSH
• biologické markery metabolismus   MeSH
• celotělové ozáření * MeSH
• dospělí MeSH
• ferredoxin-NADP-reduktasa genetika   metabolismus   MeSH
• kurkumin farmakologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• lipopolysacharidy farmakologie   MeSH
• mladý dospělý MeSH
• nádory metabolismus   radioterapie   MeSH
• počítačová rentgenová tomografie MeSH
• RNA krev   účinky léků   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• upregulace účinky léků   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

The prevalent c.903+469T>C mutation in MTRR causes the cblE type of homocystinuria by strengthening an SRSF1 binding site in an ESE leading to activation of a pseudoexon. We hypothesized that other splicing regulatory elements (SREs) are also critical for MTRR pseudoexon inclusion. We demonstrate that the MTRR pseudoexon is on the verge of being recognized and is therefore vulnerable to several point mutations that disrupt a fine-tuned balance between the different SREs. Normally, pseudoexon inclusion is suppressed by a hnRNP A1 binding exonic splicing silencer (ESS). When the c.903+469T>C mutation is present two ESEs abrogate the activity of the ESS and promote pseudoexon inclusion. Blocking the 3'splice site or the ESEs by SSOs is effective in restoring normal splicing of minigenes and endogenous MTRR transcripts in patient cells. By employing an SSO complementary to both ESEs, we were able to rescue MTRR enzymatic activity in patient cells to approximately 50% of that in controls. We show that several point mutations, individually, can activate a pseudoexon, illustrating that this mechanism can occur more frequently than previously expected. Moreover, we demonstrate that SSO blocking of critical ESEs is a promising strategy to treat the increasing number of activated pseudoexons.

• MeSH
• buněčné linie MeSH
• exony * MeSH
• ferredoxin-NADP-reduktasa genetika   metabolismus   MeSH
• HEK293 buňky MeSH
• homocystinurie enzymologie   genetika   MeSH
• kultivované buňky MeSH
• lidé MeSH
• megaloblastová anemie enzymologie   genetika   MeSH
• místa sestřihu RNA MeSH
• mutace * MeSH
• oligonukleotidy * MeSH
• regulační sekvence ribonukleových kyselin * MeSH
• sestřih RNA * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• ferredoxin-NADP-reduktasa genetika   MeSH
• fluoruracil terapeutické užití   MeSH
• genotyp MeSH
• kolorektální nádory farmakoterapie   genetika   MeSH
• lidé MeSH
• methylentetrahydrofolátreduktasa (NADPH2) genetika   MeSH
• polymorfismus genetický MeSH
• přežití bez známek nemoci MeSH
• protinádorové antimetabolity terapeutické užití   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• dopisy MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

Polymorphic variants in genes involved in one-carbon metabolism, in particular of dietary folate, may modulate the risk for colorectal cancer through aberrant DNA-methylation and altered nucleotide synthesis and repair. In the present study, we have assessed the association of six polymorphisms and relative haplotypes in the MTHFR gene (rs1801133 and rs1801131) and in the MTRR gene (rs1801394, rs1532268, rs162036, and rs10380) with the risk for colorectal cancer in 666 patients and 1377 controls from the Czech Republic. We found that the 677 C>T polymorphism in the MTHFR gene significantly decreased the risk for colorectal cancer in homozygous carriers of the variant allele (OR, 0.58; 95% CI, 0.39-0.87). Also, we noted a significantly different distribution of genotypes between cases and controls for the 66A>G polymorphism in the MTRR gene. In particular, homozygous carriers of the G-containing allele of this polymorphism were at an increased risk for colorectal cancer (OR, 1.39; 95% CI, 1.04-1.85). Haplotype analysis of the two MTHFR polymorphisms showed a moderate difference in the distribution of the TA haplotype between cases and controls. In comparison to the most common haplotype (CA), the TA haplotype was associated with a decreased risk for colorectal cancer (OR, 0.84; 95% CI, 0.71-0.99). No difference in the distribution between cases and controls was observed for the haplotypes based on the four polymorphisms in the MTRR gene. The present study suggests that the 677TT genotype and the TA haplotype in the MTHFR gene may also have a role in colorectal cancer risk in the Czech population, indicating the importance of genes involved in folate metabolism with respect to cancer risk. For MTRR, additional studies on larger populations are needed to clarify the possible role of variation in this gene in colorectal carcinogenesis.

• MeSH
• dospělí MeSH
• ferredoxin-NADP-reduktasa genetika   MeSH
• genetická predispozice k nemoci MeSH
• genotyp MeSH
• haplotypy MeSH
• jednonukleotidový polymorfismus MeSH
• kolorektální nádory genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• methylentetrahydrofolátreduktasa (NADPH2) genetika   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

Deep intronic mutations are often ignored as possible causes of human diseases. A deep intronic mutation in the MTRR gene, c.903+469T>C, is the most frequent mutation causing the cblE type of homocystinuria. It is well known to be associated with pre-mRNA mis-splicing, resulting in pseudoexon inclusion; however, the pathological mechanism remains unknown. We used minigenes to demonstrate that this mutation is the direct cause of MTRR pseudoexon inclusion, and that the pseudoexon is normally not recognized due to a suboptimal 5' splice site. Within the pseudoexon we identified an exonic splicing enhancer (ESE), which is activated by the mutation. Cotransfection and siRNA experiments showed that pseudoexon inclusion depends on the cellular amounts of SF2/ASF and in vitro RNA-binding assays showed dramatically increased SF2/ASF binding to the mutant MTRR ESE. The mutant MTRR ESE sequence is identical to an ESE of the alternatively spliced MST1R proto-oncogene, which suggests that this ESE could be frequently involved in splicing regulation. Our study conclusively demonstrates that an intronic single nucleotide change is sufficient to cause pseudoexon activation via creation of a functional ESE, which binds a specific splicing factor. We suggest that this mechanism may cause genetic disease much more frequently than previously reported.

• MeSH
• Cercopithecus aethiops MeSH
• COS buňky MeSH
• exony genetika   MeSH
• ferredoxin-NADP-reduktasa genetika   MeSH
• homocystinurie klasifikace   enzymologie   genetika   MeSH
• introny genetika   MeSH
• jaderné proteiny metabolismus   MeSH
• messenger RNA genetika   metabolismus   MeSH
• místa sestřihu RNA genetika   MeSH
• molekulární sekvence - údaje MeSH
• mutace genetika   MeSH
• mutantní proteiny genetika   MeSH
• proteiny vázající RNA metabolismus   MeSH
• sekvence nukleotidů MeSH
• sestřih RNA genetika   MeSH
• vazba proteinů MeSH
• vitamin B 12 metabolismus   MeSH
• výpočetní biologie MeSH
• zesilovače transkripce genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH