Bacterial RNA polymerase (RNAP) is an essential multisubunit protein complex required for gene expression. Here, we characterize YvgS (HelD) from Bacillus subtilis, a novel binding partner of RNAP. We show that HelD interacts with RNAP-core between the secondary channel of RNAP and the alpha subunits. Importantly, we demonstrate that HelD stimulates transcription in an ATP-dependent manner by enhancing transcriptional cycling and elongation. We demonstrate that the stimulatory effect of HelD can be amplified by a small subunit of RNAP, delta. In vivo, HelD is not essential but it is required for timely adaptations of the cell to changing environment. In summary, this study establishes HelD as a valid component of the bacterial transcription machinery.
- MeSH
- adenosintrifosfát metabolismus MeSH
- Bacillus subtilis enzymologie genetika MeSH
- bakteriální proteiny izolace a purifikace metabolismus MeSH
- DNA řízené RNA-polymerasy chemie izolace a purifikace metabolismus MeSH
- DNA metabolismus MeSH
- elongace genetické transkripce MeSH
- fenotyp MeSH
- genetická transkripce * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. Mutations in genes encoding regulators of the alternative complement pathway (CFH, MCP, C3, CFI, CFB, THBD, and CFHR1-5) are connected with this disease. Polymorphisms (SNPs) in these genes might also influence the manifestation of aHUS. We have analyzed the genes of CFH, CFI, MCP, and C3 in a cohort of 10 unrelated Czech patients with clinically diagnosed familial aHUS. Surprisingly, 4 patients had mutations only in MCP, without mutations in any of the other genes that cause aHUS. Mutations, as yet unpublished, were widely distributed over the gene (SCR2 domain, signal peptide, and cytoplasmic region). The phenotype of the patients and their close relatives (14 individuals) was also investigated. Functional examination of MCP was also provided and proved lower expression on granulocytes in all mutations. Severity of disease varied, but onset was never earlier than 5 years of age. Penetrance of disease was 50% among carriers. We found that the severity and recurrence of the disease within families varied and might also be dependent on SNPs. Mutations in the MCP gene seems to be a common etiology of aHUS in Czech patients.
- MeSH
- antigeny CD46 genetika metabolismus MeSH
- dítě MeSH
- dospělí MeSH
- fenotyp MeSH
- genetická predispozice k nemoci MeSH
- granulocyty imunologie MeSH
- hemolyticko-uremický syndrom genetika imunologie terapie MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mutace MeSH
- mutační analýza DNA MeSH
- penetrance MeSH
- prognóza MeSH
- recidiva MeSH
- rodokmen MeSH
- stupeň závažnosti nemoci MeSH
- věk při počátku nemoci MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
Závěrečná zpráva o řešení grantu Interní grantové agentury MZ ČR
24 l. : il., tab. ; 31 cm
In our study we will focus on DNA and protein analysis and correlation between mutations and clinical manifestations in patients with MYH9-related disorders.
V této studii se zaměříme na DNA a proteinovou analýzu a na souvislost mezi mutacemi v genu MYH9 a klinickými projevy u pacientů s "MYH9 onemocnením".
- MeSH
- klinické lékařství MeSH
- mutační analýza DNA MeSH
- myosiny genetika MeSH
- trombocytopatie MeSH
- trombocytopenie etiologie MeSH
- Konspekt
- Patologie. Klinická medicína
- NLK Obory
- hematologie a transfuzní lékařství
- onkologie
- biologie
- NLK Publikační typ
- závěrečné zprávy o řešení grantu IGA MZ ČR
Currently, the May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FTNS) and Epstein (EPS) syndrome are considered to be distinct clinical manifestations of a single disease caused by mutations of the MYH9 gene encoding the heavy chain of non-muscle myosin IIA (NMMHC-IIA). Manifestations of these disorders include giant platelets, thrombocytopenia and combinations of the presence of granulocyte inclusions, deafness, cataracts and renal failure. We examined 15 patients from 10 unrelated families on whom we performed immunostaining of NMMHC-IIA in blood samples. Polymerase chain reaction (PCR) analysis of selected exons of the MYH9 gene revealed mutations in nine samples with one novel mutation. Results of fluorescence and mutational analysis were compared with clinical manifestations of the MYH9 disorder. We also determined the number of glycoprotein sites on the surface of platelets. Most patients had an increased number of glycoproteins, which could be due to platelet size.
- MeSH
- buněčná inkluze MeSH
- exony MeSH
- genetické nemoci vrozené genetika krev patologie MeSH
- glykoproteiny MeSH
- granulocyty patologie MeSH
- lidé MeSH
- molekulární motory genetika MeSH
- mutace MeSH
- mutační analýza DNA MeSH
- polymerázová řetězová reakce MeSH
- rodina MeSH
- syndrom MeSH
- těžké řetězce myosinu genetika MeSH
- trombocytopenie MeSH
- trombocyty patologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FTNS) and Epstein (EPS) syndromes are rare autosomal dominant disorders with giant platelets and thrombocytopenia. Other manifestations of these disorders are combinations of the presence of granulocyte inclusions and deafness, cataracts and renal failure. Currently, MHA, SBS, FTNS and EPS are considered to be distinct clinical manifestation of a single illness caused by mutations of the MYH9 gene encoding the heavy chain of non-muscle myosin IIA (NMMHC-IIA). As the MYH9 gene has a high number of exons, it takes much time and material to use this method for the detection of MYH9 mutations. Recently, a new method has been introduced for scanning DNA mutations without the need for direct sequencing: high-resolution melting analysis (HRMA). Mutation detection with HRMA relies on the intercalation of the specific dye (LC Green plus) in double-strand DNA and fluorescence monitoring of PCR product melting profiles. In our study, we optimized the conditions and used HRMA for rapid screening of mutations in all MYH9 exons in seven affected individuals from four unrelated families with suspected MYH9 disorders. Samples identified by HRMA as positive for the mutation were analysed by direct sequencing. HRMA saved us over 85% of redundant sequencing.
- MeSH
- lidé MeSH
- missense mutace MeSH
- molekulární motory genetika chemie MeSH
- mutační analýza DNA metody MeSH
- těžké řetězce myosinu genetika chemie MeSH
- tranzitní teplota MeSH
- trombocytopenie genetika krev MeSH
- trombocyty patologie MeSH
- velikost buňky MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Východisko. Trombotická trombocytopenická purpura je těžké multisystémové onemocnění charakterizované konzumpční trombocytopenií, mikroangiopatickou hemolýzou s mikrotromby v mikrocirkulaci, fluktujícím neurologickým nálezem, případně poruchou renálních funkcí. Podobně se projevuje i hemolyticko-uremický syndrom a jiná onemocnění. Nedávné studie prokázaly, že onemocnění způsobuje defekt metaloproteázy štěpící von Willebrandův faktor do menších multimerů. Jako člen ADAMTS rodiny metaloproteáz získala název ADAMTS13. Cílem naší práce bylo charakterizovat mutace v genu pro ADAMTS13 u vrozené formy trombotické trombocytopenické purpury. Metody a výsledky. Mutační analýzu jsme prováděli u 9 pacientů a jejich 12 rodinných příslušníků. Aktivita ADAMTS13 byla měřena modifikovanou metodou kvantitativního imunoblotingu degradovaných multimerů von Willebrandova faktoru. Mutační analýza byla prováděna sekvenováním všech 29 exonů a jejich přilehlých intronových oblastí. Nalezli jsme pět rozdílných mutací. Tři z nich dosud nebyly publikovány. Závěry. Mutační analýza odhalila zajímavý výsledek. Nalezli jsme inzertní mutaci 4143insA v 8 z 9 pacientů. Tato často se vyskytující mutace představuje výhodu pro diferenciální diagnózu vrozené formy trombotické trombocytopenické purpury, protože klinická manifestace u dětí je velmi heterogenní a rychlá detekce mutace je přínosem v prevenci léčby rekurencí.
Background. Thrombotic thrombocytopenic purpura is characterized by microvascular platelet clumping resulting in thrombocytopenia, microangiopathic hemolysis, neurological abnormality, and renal dysfunction. Similar manifestations also occur in patients with the hemolytic uremic syndrome or other types of disorders. Recent studies demonstrate that severe deficiency of the von Willebrand factor cleaving metalloprotease, ADAMTS13, causes thrombotic thrombocytopenic purpura. Aim of our study was to characterize gene defects causing inherited type of disease. Methods and Results. We investigated nine patients with recurrent type of disease with familiar origin and twelve relatives. Samples were taken in a remission of disease. We measured activity of ADAMTS13 (vWF-CP) with modified method of the quantitative immunoblotting of degraded vWF multimers. Mutation screening was carried out by sequencing all 29 exons and flanking intron regions of the ADAMTS13 gene. Five distinct mutations were found. Three of them are novel. Conclusions. Mutation analysis of the ADAMTS13 gene brought interesting results in eight patients. We found a one single base frameshift insertion, 4143insA in 8 of 9 unrelated individuals. This investigation represents an advantage in the differential diagnosis of disease since the thrombotic thrombocytopenic purpura phenotype in childhood can be variable and rapid detection of mutation is helpful for the recurrence prevention.
- MeSH
- diagnostické techniky molekulární metody využití MeSH
- finanční podpora výzkumu jako téma MeSH
- kohortové studie MeSH
- lidé MeSH
- tkáňové inhibitory metaloproteinas genetika MeSH
- trombotická trombocytopenická purpura epidemiologie patofyziologie vrozené MeSH
- von Willebrandův faktor fyziologie MeSH
- Check Tag
- lidé MeSH
