• BMB Reports
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• 제49권7호
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• pp.355-356
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• 2016

The THO/TREX complex consists of several conserved subunits and is required for mRNA export. In metazoans, THO/TREX binds a subset of mRNAs during RNA splicing, and facilitates their nuclear export. How THO/TREX selects RNA targets is, however, incompletely understood. In our recent study, we reported that THO is loaded onto Piwi-interacting RNA (piRNA) precursor transcripts independent of splicing, and facilitates convergent transcription in Drosophila ovary. The precursors are later processed into mature piRNAs, small noncoding RNAs that silence transposable elements (TEs). We observed that piRNAs originating from dual-strand clusters, where precursors are transcribed from both strands, were specifically affected by THO mutation. Analysis of THO-bound RNAs showed enrichment of dual-strand cluster transcripts. Interestingly, THO loading onto piRNA precursors was dependent on Cutoff (Cuff), which comprises the Rhino-Deadlock-Cutoff (RDC) complex that is recruited to dual-strand clusters by recognizing H3K9me3 and licenses convergent transcription from he cluster. We also found that THO mutation affected transcription from dual-strand clusters. Therefore, we concluded that THO/TREX is recruited to dual-strand piRNA clusters, independent of splicing events, via multi-protein interactions with chromatin structure. Then, it facilitates transcription likely by suppressing premature termination to ensure adequate expression of piRNA precursors.

• Genomics & Informatics
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• 제7권4호
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• pp.181-186
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• 2009

Myotonic dystrophy type 1 (DM1), which is a dominantly inherited neurodegenerative disorder, results from a CTG trinucleotide repeat expansion in the 3'-untranslated region (3'-UTR) of the myotonic dystrophy protein kinase (DMPK) gene. Retention of mutant DMPK (mDMPK) transcripts in the nuclei of affected cells has been known to be the main cause of pathogenesis of the disease. Thus, reducing the RNA toxicity through elimination of the mutant RNA has been suggested as one therapeutic strategy against DM1. In this study, we suggested RNA replacement with a trans -splicing ribozyme as an alternate genetic therapeutic approach for amelioration of DM1. To this end, we identified the regions of mDMPK 3'-UTR RNA that were accessible to ribozymes by using an RNA mapping strategy based on a trans-splicing ribozyme library. We found that particularly accessible sites were present not only upstream but also downstream of the expanded repeat sequence. Repair or replacement of the mDMPK transcript with the specific ribozyme will be useful for DM1 treatment through reduction of toxic mutant transcripts and simultaneously restore wild-type DMPK or release nucleus-entrapped mDMPK transcripts to the cytoplasm.

• BMB Reports
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• 제52권11호
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• pp.641-646
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• 2019

The Ron proto-oncogene is a human receptor for macrophage-stimulating protein (MSP). The exclusion of exon 11 in alternative splicing generates ${\Delta}RON$ protein that is constitutively activated. Heterogenous ribonucleaoprotein (hnRNP) $C_1/C_2$ is one of the most abundant proteins in cells. In this manuscript, we showed that both hnRNP $C_1$ and $C_2$ promoted exon 11 inclusion of Ron pre-mRNA and that hnRNP $C_1$ and hnRNP $C_2$ functioned independently but not cooperatively. Moreover, hnRNP $C_1$ stimulated exon 11 splicing through intron 10 activation but not through intron 11 splicing. Furthermore, we showed that, whereas the RRM domain was required for hnRNP $C_1$ function, the Asp/Glu domain was not. In conclusion, hnRNP $C_1/C_2$ promoted exon 11 splicing independently by stimulating intron 10 splicing through RRM but not through the Asp/Glu domain.

• BMB Reports
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• 제36권6호
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• pp.538-544
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• 2003

The hepatitis C virus (HCV) is a major causative agent of chronic hepatitis and hepatocellular carcinoma. The development of alternative antiviral therapies is warranted because current treatments for the HCV infection affect only a limited number of patients and lead to significant toxicities. The HCV genome is exclusively present in the RNA form; therefore, ribozyme strategies to target certain HCV sequences have been proposed as anti-HCV treatments. In this study, we determined which regions of the internal ribosome entry site (IRES) of HCV are accessible to ribozymes by employing an RNA mapping strategy that is based on a trans-splicing ribozyme library. We then discovered that the loop regions of the domain IIIb of HCV IRES appeared to be particularly accessible. Moreover, to verify if the target sites that were predicted to be accessible are truly the most accessible, we assessed the ribozyme activities by comparing not only the trans-splicing activities in vitro but also the trans-cleavage activities in cells of several ribozymes that targeted different sites. The ribozyme that could target the most accessible site identified by mapping studies was then the most active with high fidelity in cells as well as in vitro. These results demonstrate that the RNA mapping strategy represents an effective method to determine the accessible regions of target RNAs and have important implications for the development of various antiviral therapies which are based on RNA such as ribozyme, antisense, or siRNA.

• 한국콘텐츠학회논문지
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• 제10권10호
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• pp.102-110
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• 2010

선택 스플라이싱은 유전자 발현의 중요한 과정 중 하나이다. 선택 스플라이싱이 발생함에 따라, 돌연변가 발생하여, 질병을 일으킬 수 있다. 대부분의 선택 스플라이싱 연구는 EST(Expressed Sequence Tag)를 이용한다. 그러나, EST를 이용하여 선택 스플라싱을 예측하는 데는 몇 가지 단점이 있다. EST가 저장되어 있는 라이브러리가 잘 정돈되어 있지 않거나, 잘못 열거되어 있을 경우, 실험 시 EST를 잘못 선택할 수 있다. 또한, EST가 아직 발견되지 않은 유전 서열에서는 선택 스플라이싱을 찾을 방법이 없다. 이 논문에서는 이러한 EST 기반 연구의 약점을 개선하고, 선택 스플라이싱의 탐지 및 예측의 질을 높이기 위해서, pre-mRNA에서 One-leaf One-node Tree 알고리즘을 제안한다. 이 트리는 Arabidopsis thaliana의 각 염색체에 대해서 실험되었다. 실험 결과, 모든 염색체에서 codons에 따라 일반 스플라싱과 선택 스플라싱이 다른 패턴을 가지는 것으로 나타났다. 트리 알고리즘에서 도출된 패턴으로 부터, 아직 발견되지 않은 선택 스플라싱도 예측할 수 있다.

• Molecules and Cells
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• 제46권11호
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• pp.664-671
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• 2023

The proper maintenance of mRNA quality that is regulated by diverse surveillance pathways is essential for cellular homeostasis and is highly conserved among eukaryotes. Here, we review findings regarding the role of mRNA quality control in the aging and longevity of Caenorhabditis elegans, an outstanding model for aging research. We discuss the recently discovered functions of the proper regulation of nonsense-mediated mRNA decay, ribosome-associated quality control, and mRNA splicing in the aging of C. elegans. We describe how mRNA quality control contributes to longevity conferred by various regimens, including inhibition of insulin/insulin-like growth factor 1 (IGF-1) signaling, dietary restriction, and reduced mechanistic target of rapamycin signaling. This review provides valuable information regarding the relationship between the mRNA quality control and aging in C. elegans, which may lead to insights into healthy longevity in complex organisms, including humans.

• 생명과학회지
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• 제30권6호
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• pp.570-579
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• 2020

Pre-mRNA의 스플라이싱은 진핵생물 유전자의 적절한 발현에 매우 중요한 역할을 한다. 선택적 스플라이싱은 스플라이싱 위치가 서로 다르게 인식될 때 발생하며 동일한 pre-mRNA로부터 둘 이상의 전사체와 단백질을 생성할 수 있다. 스플라이싱 위치의 결정은 스플라이소솜과 SR 단백질, hnRNP, CBP 등의 스플라이싱 인자에 의해 조절된다. 고온, 저온, 고염, 건조, 저산소 등 다양한 환경 스트레스 조건에서 식물의 많은 스트레스 반응 유전자에 대해 선택적 스플라이싱이 일어나는 것이 알려져 있으며, 이러한 선택적 스플라이싱은 식물이 환경 변화에 적응하기 위한 중요한 기작 중 하나로 여겨진다. 저온, 고온, 고염, 건조 스트레스 조건에서는 스플라이싱 인자의 발현이 변하거나 또는 정상 조건에서와는 다른 스플라이싱 활성을 가짐으로써 선택적 스플라이싱이 일어난다. 환경 스트레스 반응 유전자의 스플라이싱 이소형은 각각 환경 스트레스에 대해 서로 다른 반응을 보이는데 생성되는 조직이 서로 다르기도 하고, 일부 이소형은 넌센스-매개 분해에 의해 분해되기도 한다. 스플라이싱 이소형의 단백질은 환경 스트레스 조건에서 정상 조건과 비교하여 세포 내 위치가 다르기도 하고, 전사인자 또는 효소로서 다른 활성을 가지기도 한다. 이러한 다양한 연구에도 불구하고 식물의 환경 스트레스 반응에서 선택적 스플라이싱에 대한 연구는 일부 스트레스와 유전자에 국한 되어 있고, 아직 분자 기전이 제대로 밝혀지지 않은 부분이 많아 앞으로 더 많은 연구가 필요하다.

• Genomics & Informatics
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• 제5권1호
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• pp.32-35
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• 2007

Telomerase reverse transcriptase (TERT) is an enzymatic ribonucleoprotein that prolongs the replicative life span of cells by maintaining protective structures at the ends of eukaryotic chromosomes. Telomerase activity is highly up-regulated in 85-90% of human cancers, and is predominately regulated by hTERT expression. In contrast, most normal somatic tissues in humans express low or undetectable levels of telomerase activity. This expression profile identifies TERT as a potential anticancer target. By using an RNA mapping strategy based on a trans-splicing ribozyme library, we identified the regions of mouse TERT (mTERT) RNA that were accessible to ribozymes. We found that particularly accessible sites were present downstream of the AUG start codon. This mTERTspecific ribozyme will be useful for validation of the RNA replacement as cancer gene therapy approach in mouse model with syngeneic tumors.

• Molecules and Cells
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• 제46권1호
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• pp.48-56
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• 2023

Genomic information stored in the DNA is transcribed to the mRNA and translated to proteins. The 3' untranslated regions (3'UTRs) of the mRNA serve pivotal roles in post-transcriptional gene expression, regulating mRNA stability, translation, and localization. Similar to DNA mutations producing aberrant proteins, RNA alterations expand the transcriptome landscape and change the cellular proteome. Recent global analyses reveal that many genes express various forms of altered RNAs, including 3'UTR length variants. Alternative polyadenylation and alternative splicing are involved in diversifying 3'UTRs, which could act as a hidden layer of eukaryotic gene expression control. In this review, we summarize the functions and regulations of 3'UTRs and elaborate on the generation and functional consequences of 3'UTR diversity. Given that dynamic 3'UTR length control contributes to phenotypic complexity, dysregulated 3'UTR diversity might be relevant to disease development, including cancers. Thus, 3'UTR diversity in cancer could open exciting new research areas and provide avenues for novel cancer theragnostics.

• Animal cells and systems
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• 제4권2호
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• pp.141-144
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• 2000

Effects of deamido-$\textrm{NAD}^{+}$on self-splicing of primary transcripts of the phage T4 thymidylate synthase gene (td) was investigated. The self-splicing was not affected by deamido-$\textrm{NAD}^{+}$- at concentrations up to 2 mM. However, it began to decrease at 5 mM and the formation of splicing products such as the linear intron, intron-exon 2 and exon 1-exon 2, was slightly reduced. At 20 mM the self-splicing activity was almost completely abolished. This analog of the coenzyme $\textrm{NAD}^{+}$- inhibits the self-splicing of td intron RNA although it does not possess a guanidine group in its structure. The analysis of inhibitory concentrations and structural examination suggests that the key structural features of deamido-$\textrm{NAD}^{+}$ responsible for the inhibition of splicing may be the ADP-ribose moiety.