• Animal cells and systems
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• 제1권2호
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• pp.363-370
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• 1997

RNase MRP is a ribonucleoprotein complex with a site-specific endonuclease activity. Its original substrate for cleavage is the small mitochondrial RNA near the mitochondrial DNA replication origin, thus it was proposed to generate the primer for mtDNA replication. Recently, it has been shown to have another substrate in the nucleus, such as pre-S.8S ribosomal RNA in nucleolus. The gene for the RNA component of RNase MRP (MRP RNA) was found to be encoded by the nucleus genome, suggesting an interesting intracellular trafficking of MRP RNA to both mitochondria and nucleolus after transcription in nucleus. In this study, genomic DNA encoding MRP RNA was microinjected into the nucleus of Xenopus oocytes, to analyze promoter regions involved in the transcription. It showed that the proximal sequence element and TATA box are important for basal level transcription; octamer motif and Sp1 binding sites are for elevated level transcription. Most of Xenopus MRP RNA was exported out to the cytoplasm following transcription in the nucleus. Utilizing various hybrid constructs, export of MRP RNA was found to be regulated by the promoter and the 5' half of the coding region of the gene. Interestingly, the transcription in nucleus seems to be coupled to the export of MRP RNA to cytoplasm. Intracellular transport of injected MRP RNA can be easily visualized by whole-mount in situ hybridization following microinjection; it also shows possible intra-nuclear sites for transcription and export of MRP RNA.

• Journal of Microbiology
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• 제44권1호
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• pp.11-22
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• 2006

Escherichia coli protein Rho is required for the factor-dependent transcription termination by an RNA polymerase and is essential for the viability of the cell. It is a homohexameric protein that recognizes and binds preferably to C-rich sites in the transcribed RNA. Once bound to RNA, it utilizes RNA-dependent ATPase activity and subsequently ATPase-dependent helicase activity to unwind RNA-DNA hybrids and release RNA from a transcribing elongation complex. Studies over the past few decades have highlighted Rho as a molecule and have revealed much of its mechanistic properties. The recently solved crystal structure could explain many of its physiological functions in terms of its structure. Despite all these efforts, many of the fundamental questions pertaining to Rho recognition sites, differential ATPase activity in response to different RNAs, translocation of Rho along the nascent transcript, interactions with elongation complex and finally unwinding and release of RNA remain obscure. In the present review we have attempted to summarize 'the knowns' and 'the unknowns' of the Rho protein revealed by the recent developments in this field. An attempt has also been made to understand the physiology of Rho in the light of its phylogeny.

• Journal of Microbiology
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• 제45권6호
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• pp.534-540
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• 2007

The Streptomyces coelicolor A3(2) genome contains six operons (rrnA to F) for ribosomal RNA synthesis. Transcription from rrnD occurs from four promoters (p1 to p4). We found that transcripts from the p1 and p3 promoters were most abundant in vivo in the early exponential phase. However, at later phases of exponential and stationary growth, transcripts from the p1 promoter decreased drastically, with the p3 and p4 transcripts constituting the major forms. Partially purified RNA polymerase supported transcription from the p3 and p4 promoters, whereas pure reconstituted RNA polymerase with core enzyme (E) and the major vegetative sigma factor ${\sigma}^{HrdB}$ ($E{\cdot}{\sigma}^{HrdB}$) did not. In order to assess any potential requirement for additional factor(s) that allow transcription from the p3 and p4 promoters, we fractionated a partially purified RNA polymerase preparation by denaturing gel filtration chromatography. We found that transcription from the p3 and p4 promoters required factor(s) of about 30-35 kDa in addition to RNAP holoenzyme ($E{\cdot}{\sigma}^{HrdB}$). Therefore, transcription from the p3 and p4 promoters, which contain a consensus -10 region but no -35 for ${\sigma}^{HrdB}$ recognition, are likely to be regulated by transcription factor(s) that modulate RNA polymerase holoenzyme activity in S. coelicolor.

• 미생물과산업
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• 제14권1호
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• pp.7-11
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• 1988

Efficient in vitro RNA synthesis can be easily accomplished from cloned DNA using bactrio-phage SP6, T7 or T3 RNA polymerase. Despite its popularity as in vitro transcription system, molecular mechanisms of bacteriophage transcription has not been studied, although physical and catalytic properties of several phage RNA polymerases have well been documented (1). Only recently the T7 promoter has been physically mapped by footprinting of the T7 RNA polymerase (2,3). These simple phage systems, however, could be useful for detailed molecular studies of transcription.

• 자연과학논문집
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• 제14권1호
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• pp.39-50
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• 2004

박테리오 파아지 T7 RNA 중합효소는 다른 RNA 중합효소와 비교하여 볼 때 보조인자 없이 전사를 진행하는 하나의 subunit로 구성된 RNA 중합효소이다. 전사 진행 단계 중에서 T7 RNA 중합효소의 전사연장을 연구하기 위해 biotin이 결합된 DNA 주형을 streptavidin bead로 고정시킴으로서 T7 RNA 중합효소의 진행과정을 관찰할 수 있었고, 이러한 기작을 이용하여 일련의 활성을 가지는 가장 안정한 전사연장복합체들을 얻을 수 있었다. 전사 연장체들은 16번 염기 위치로부터 18번 염기의 위치까지 방사선 동위원소가 표지되어 있으며 이들 표지된 전사연장복합체들은 단계별로 합성하여 22-40개 핵산잔기들이 합성된 전사연장복합체들을 얻을 수 있었다. 이와 같은 전사연장복합체들을 PTH 전사종결 부위가 있는 주형으로 사용하여 야생형 및 R173C 돌연변이 RNA 중합효소를 이용하여 전사연장복합체를 제조하여 비교한 결과 PTH 전사종결에 둔감한 R173C 돌연변이 중합효소의 경우 야생형에 비해 PTH 전사종결부위를 지난 위치에서도 전사연장복합체가 생성되었다.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 심포지엄
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• pp.37-37
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• 2003

Enzymes involved in catecholamine synthesis are present in the highest concentration in the adrenal medulla, however they were found also in other, mainly nervous tissues. Increased transcription of genes for catecholamine biosynthetic enzymes is an important mechanism to increase the capacity for epineprine/norepinephrine biosynthesis with stress. Gastrodia elata(Chinese name: Tienma), are very important Chinese herbal medicines used for the medical treatment of headaches, migraine, dizziness, epilepsy, rheumatism, neuralgia, paralysis and other neuralgic and nervous disorders. Immobilize stressed rat markedly increased tyrosine hydroxylase (TH) mRNA and dopamine-$\beta$-hydroxylase (DBH) mRNA transcription level more than control group. But treated Gastrodia elata extracts in immobilized stressed rat slightly increased TH mRNA and DBH mRNA transcription level more than normal group. In addition, we are obtained identical results in PC12 cell line. Decrease of transcription level of TH mRNA and DBH mRNA is indicating that Gastrodia elata have a anti-stress effects which decrease the transcription level of TH and DBH mRNA on catecholamine biosynthesis pathway.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2005년도 춘계학술대회
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• pp.5-14
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• 2005

Recently, data from several groups have raised the concept of 'checkpoint' in transcription. As capping of nascent RNA transcript is tightly coupled to RNA polymerase II transcription, we seek to obtain direct evidence that transcripiton checkpoint via capping enzyme functions in this early regulatory step. One of temperature sensitive (ts) alleles of ceg1, a guanylyltransferase subunit of the Saccharomyces cerevisiaecapping enzyme, showed 6-azauracil (6AU) sensitivity at the permissive growth temperature, which is a phenotype that is correlated with a transcription elongational defect. This ts allele, ceg1-63 also has an impaired ability to induce PUR5 in response to a 6AU treatment. However, this cellular and molecular defect is not due to the preferential degradation of the transcript attributed from a lack of guanylyltransferase activity. On the contrary, the data suggests that the guanylyltransferase subunit of the capping enzyme plays a role in transcription elongation. First, in addition to the 6AU sensitivity, ceg1-63is synthetically lethal with elongation defective mutations of the largest subunit of RNA polymerase II. Secondly, it exhibited a lower GAL1 mRNA turn-over after glucoseshut off. Third, it decreased the transcription read through a tandem array of promoter proximal pause sites in an orientation dependent manner. Interestingly, this mutant also showed lower pass through a pause site located further downstream of the promoter. Taken together, these results suggest that the capping enzyme plays the role of an early transcription checkpoint possibly in the step of the reversion of repression by stimulating polymerase to escape from the promoter proximal arrest once RNA becomes appropriately capped.

• 미생물학회지
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• 제29권4호
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• pp.221-225
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• 1991

The RNA transcripts produced from in vitro transcription reaction of BTV core were analyzed on agarose-urea gel. Fast migrating abortive RNAs, in addition to full length species of RNA, were observed. Fast migrating RNAs extracted from agarose-urea gel were hybridized to all 10 segments of genomic ds RNA, while solw migrating RNAs extracted from agarose-urea gel were hybridized only to the large and medium size genomic ds RNA. These results indicate that fast migrating RNA transcripts are most likely the products of abortive transcription.

• Journal of Microbiology
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• 제39권1호
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• pp.42-48
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• 2001

The regulation of actin gene expression during the differentiation of Naegleria gruberi was examined. Actin mRNA concentration was maximal in amoebae and decreased rapidly after the initiation of differentiation. At 20 min after initiation, the concentration of actin mRNA decreased to 55% of the maximal value. The actin mRNA concentration decreased to the minimum at 80 min (15% of the maximum), and then began to increase slightly at the end of differentiation. This decrease of actin mRNA concentration was regulated by the repression of actin gene transcription based on nuclear run-on transcription experiments. The rates of transcription of actin gene in nuclei prepared at 40 and 80 min after the initiation of differentiation were 50 and 28% of that of nuclei prepared at the beginning of differentiation, respectively. The addition of cycloheximide at the initiation of differentiation inhibited both the rapid decrease in the concentration of actin mRNA and the repression of actin gene transcription. These results suggest that the rapid decrease in the concentration of actin mRNA during the differentiation of N. gruberi is accomplished by the repression of actin gene transcription and this transcriptional regulation requires continuous protein synthesis during the differentiation.

• Journal of Microbiology and Biotechnology
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• 제4권3호
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• pp.183-190
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• 1994

Cell-free extracts prepared from cultured insect cells, Spodoptera. frugiperda, were analyzed for activation of early gene transcription of an insect baculovirus, Autographa californica nuclear polyhedrosis virus (AcNPV). The template DNA used for in vitro transcription assays contained promoter sites for the baculovirus genes that have been classified as immediate early ($\alpha$) or early genes. These genes are located in the HindIII-K/Q region of the AcNPV genome. Nuclei isolated from the AcNPV-infected Spodoptera frugiperda cells were also used for in vitro transcription analysis by RNase-mapping the labeled RNA synthesized from in vitro run-on reaction in the isolated nuclei. The genes studied by this technique were p26 and pl0 genes which were classified as delayed early and late gene, respectively. We found that transcription of the genes from the HindIII-K region was accurately initiated and unique in the whole cell extract obtained from uninfected cells, although abundance of the in vitro transcripts was reverse to that of in vivo RNA. With isolated nuclei transcription of the p26 gene was inhibited by $\alpha$-amanitin suggesting that the p26 gene was transcribed by host RNA polymerase II. However, transcription of the pl0 gene in isolated nuclei was not inhibited by $\alpha$-amanitin, but rather stimulated by the inhibitor. We also found that the synthesis of $\alpha$-amanitin-resistant RNA polymerase was begun before 6 hr p.i., the time point at which the onset of viral DNA replication as well as the appearance of a-amanitin-resistant viral transcripts were detected. These studies give us strong evidence to support the previous data that early genes of AcNPV were transcribed by host RNA polymerease III, while transcription of late genes was mediated at least by a novel $\alpha$-amanitin-resistant RNA polymerase.