• 제목/요약/키워드: RNA splicing

검색결과 157건 처리시간 0.033초

UAP56- a key player with surprisingly diverse roles in pre-mRNA splicing and nuclear export

  • Shen, Hai-Hong
    • BMB Reports
    • /
    • 제42권4호
    • /
    • pp.185-188
    • /
    • 2009
  • Transcripts contain introns that are usually removed from premessenger RNA (MRNA) in the process of pre-mRNA splicing. After splicing, the mature RNA is exported from the nucleus to the cytoplasm. The splicing and export processes are coupled. UAP56 protein, which is ubiquitously present in organisms from yeasts to humans, is a DExD/H-box family RNA helicase that is an essential splicing factor with various functions in the prespliceosome assembly and mature spliceosome assembly. Collective evidence indicates that UAP56 has an essential role in mRNA nuclear export. This mini-review summarizes recent evidence for the role of UAP56 in pre-mRNA splicing and nuclear export.

Role of $Mg^{2+}$ in RNA splicing of T4 td intron

  • Sung, Jung-Suk;Shin, Sook;Park, In-Kook
    • Journal of Microbiology
    • /
    • 제33권2호
    • /
    • pp.160-164
    • /
    • 1995
  • The splicing activity of T4 phage td intron RNA has been examined with various Mg$^{2+}$ ions such as MGCl$_{2}$, MgS $O_{4}$ and magnesium acetate using various splicing conditions such as different incubation time and temperature. The maximum splicing of td intron RNA occurred at the concentration of 5 mM MgCl$_{2}$. Raising the Mg$^{2+}$ concentration up to 15 mM appeared to promote P2 delection mutant to overcome the loss of some splicing activity. In both wild type and mutant, a complete hydrolysis of RNA occurred at 30 mM MgCI$_{2}$ MgS $O_{4}$ and magnesium acetate exhibited the rate and pattern of RNA splicing identical to MGCI$_{2}$. The optimal splicing conditions involve the incubation of RNA with 5 mM MgCI$_{2}$ at 58 .deg.C for 15 min. The results suggest that Mg$^{2+}$ may play a key role in the catalytic mechanism of td intron RNA.n RNA.

  • PDF

Isolation and Characterization of Pre-$tRNA^{Val}$ Splicing Mutants of Schizosaccharomyces pombe

  • Hwang, Ku-Chan;Kim, Dae-Myung
    • Journal of Microbiology
    • /
    • 제35권4호
    • /
    • pp.334-340
    • /
    • 1997
  • A collection of 132 temperature sensitive (ts) mutants was generated by the chemical mutagenesis of Schizosaccharomyces pombe wild type strain and screened for tRNA splicing defects on Northern blots by hybridization with an oligonucleotide that recognizes the exon of the S. pombe tRNA^Val as a probe. We identidied 6 mutants which accumulate precursor $tRNA^{Val}$. Among them, 2 mutants exhibited remarkable morphological differences compared to wild type cells. One tRNA splicing mutant showed elongated cell shape in permissive as well as non-permissive cultures. The other mutant exhibited shortened cell morphology only in nonpermissive culture. The total RNA pattern in the splicing mutants appeared to be normal. Genetic analysis of four $tRNA^{Val}$ splicing mutants demonstrated that the mutation reside in different genes.

  • PDF

Tetrahymena thermophila의 group I intron에 의한 trans-splicing 반응에 미치는 표적 RNA 구조의 영향분석 (Effects of Substrate RNA Structure on the Trans-splicing Reaction by Group I Intron of Tetrahymena thermophila)

  • 이성욱
    • 미생물학회지
    • /
    • 제35권3호
    • /
    • pp.211-217
    • /
    • 1999
  • 표적 RNA 의 구조가 Tetrahymena thermophila 의 group I intron 에 의한 trans-splicing 반응에 미치는 영향을 분석하기 위해 강력한 stem-loop 형태의 안정된 구조를 갖고 있는 표적 RNA mapping 분석 방법을 이용한 결과 in vitro 뿐만 아니라 in vivo 에서도 stem 부위의 염기들에 반해 loop 부위의 염기들이 ribozyme 에 의해 잘 인지되었으며 이러한 결과는 그러한 부위들을 인지할 수 있는 ribozyme 들에 의한 trans-cleavage 그리고 trans-splicing 반응을 수행함으로써 검증하였다. 또한 이러한 trans-splicing 반응은 정확하게 일어남을 반응 산물의 염기서열 결정을 통해 확인하였다. 따라서 표적 RNA 의 구조가 in vitro 및 in vivo 에서의 ribozyme 활성에 매우 중요한 요인임을 확인하였다.

  • PDF

Effects of $K^+$ lon on in vitro RNA Splicing of T4 Phage Thymidylate Synthase Gene

  • Sung, Jung-Suk;Park, In-Kook
    • Journal of Microbiology
    • /
    • 제34권1호
    • /
    • pp.49-53
    • /
    • 1996
  • The effects of K$^{+}$ ion on the activity of RNA splicing of T4 phage thymidylate synthase gene have been investigated. The splicing activity was stimulated within the range of 5 to 20 mM concentration of KCI. When the concentration of KCI in the splicing reaction was brought to 100 or 200 mM a small amount of the exonl-intron product (1, 4 kb) was formed with large proportion of primary RNA transcript not undergoing splicing. This observation strongly suggests that there may exist come kinds of interferences with transesterification at the first step of splicing. Overall it can be concluded that K$^{+}$ ion exhibits very unique roles in RNA splicing of tdd gene depending on its concentration.ion.

  • PDF

Replacement of Thymidine Phosphorylase RNA with Group I Intron of Tetrahymena thermophila by Targeted Trans-Splicing

  • Park, Young-Hee;Jung, Heung-Su;Kwon, Byung-Su;Lee, Seong-Wook
    • Journal of Microbiology
    • /
    • 제41권4호
    • /
    • pp.340-344
    • /
    • 2003
  • The group I intron from Tetrahymena thermophila has been demonstrated to employ splicing reactions with its substrate RNA in the trans configuration. Moreover, we have recently shown that the transsplicing group I ribozyme can replace HCV-specific transcripts with a new RNA that exerts anti-viral activity. In this study, we explored the potential use of RNA replacement for cancer treatment by developing trans-splicing group I ribozymes, which could replace tumor-associated RNAs with the RNA sequence attached to the 3' end of the ribozymes. Thymidine phosphorylase (TP) RNA was chosen as a target RNA because it is known as a valid cancer prognostic factor. By performing an RNA mapping strategy that is based on a trans-splicing ribozyme library, we first determined which regions of the TP RNA are accessible to ribozymes, and found that the leader sequences upstream of the AUG start codon appeared to be particularly accessible. Next, we assessed the ribozyme activities by comparing trans-splicing activities of several ribozymes that targeted different regions of the TP RNA. This assessment was performed to verify if the target site predicted to be accessible is truly the most accessible. The ribozyme that could target the most accessible site, identified by mapping studies, was the most active with high fidelity in vitro. Moreover, the specific trans-splicing ribozyme reacted with and altered the TP transcripts by transferring an intended 3' exon tag sequence onto the targeted TP RNA in mammalian cells with high fidelity. These results suggest that the Tetrahymena ribozyme can be utilized to replace TP RNAs in tumors with a new RNA harboring anti-cancer activity, which would revert the malignant phenotype.

Pressure-Overload Cardiac Hypertrophy Is Associated with Distinct Alternative Splicing Due to Altered Expression of Splicing Factors

  • Kim, Taeyong;Kim, Jin Ock;Oh, Jae Gyun;Hong, Seong-Eui;Kim, Do Han
    • Molecules and Cells
    • /
    • 제37권1호
    • /
    • pp.81-87
    • /
    • 2014
  • Chronic pressure-overload cardiac hypertrophy is associated with an increased risk of morbidity/mortality, largely due to maladaptive remodeling and dilatation that progresses to dilated cardiomyopathy. Alternative splicing is an important biological mechanism that generates proteomic complexity and diversity. The recent development of next-generation RNA sequencing has improved our understanding of the qualitative signatures associated with alternative splicing in various biological conditions. However, the role of alternative splicing in cardiac hypertrophy is yet unknown. The present study employed RNA-Seq and a bioinformatic approach to detect the RNA splicing regulatory elements involved in alternative splicing during pressure-overload cardiac hypertrophy. We found GC-rich exonic motifs that regulate intron retention in 5' UTRs and AT-rich exonic motifs that are involved in exclusion of the AT-rich elements that cause mRNA instability in 3' UTRs. We also identified motifs in the intronic regions involved in exon exclusion and inclusion, which predicted splicing factors that bind to these motifs. We found, through Western blotting, that the expression levels of three splicing factors, ESRP1, PTB and SF2/ASF, were significantly altered during cardiac hypertrophy. Collectively, the present results suggest that chronic pressure-overload hypertrophy is closely associated with distinct alternative splicing due to altered expression of splicing factors.

Re-Engineering of Carcinoembryonic Antigen RNA with the Group I Intron of Tetrahymena thermophila by Targeted Trans-Splicing

  • JUNG HEUNG-SU;LEE SEONG-WOOK
    • Journal of Microbiology and Biotechnology
    • /
    • 제15권6호
    • /
    • pp.1408-1413
    • /
    • 2005
  • Elevated expression of carcinoembryonic antigen (CEA) has been implicated in various biological aspects of neoplasia such as tumor cell adhesion, metastasis, blocking of cellular immune mechanisms, and antiapoptosis function. Thus, the CEA could be an important target for anticancer therapy. In this study, we developed Tetrahymena group 1 intron-based trans-splicing ribozymes that can specifically target and replace CEA RNA. To this end, we first determined which regions of the CEA RNA were accessible to ribozymes by employing an RNA mapping strategy that was based on a trans-splicing ribozyme library. Next, we assessed the ribozyme activities by comparing the trans-splicing activities of several ribozymes that targeted different regions of the CEA RNA, and then the ribozyme that could target the most accessible site was observed to be the most active with high fidelity in vitro. Moreover, the specific trans-splicing ribozyme was found to react with and altered the target CEA transcripts in mammalian cells with high fidelity. These results suggest that the Tetrahymena ribozyme can be utilized to replace CEA RNAs in tumors with a new RNA-harboring anticancer activity, thereby hopefully reverting the malignant phenotype.

In Vivo Target RNA Specificity of Trans-Splicing Phenomena by the Group I Intron

  • Song, Min-Sun;Lee, Seong-Wook
    • Genomics & Informatics
    • /
    • 제6권2호
    • /
    • pp.84-86
    • /
    • 2008
  • The Tetrahymena group I intron has been shown to employ a trans-splicing reaction and has been modified to specifically target and replace human telomerase reverse transcriptase (hTERT) RNA with a suicide gene transcript, resulting in the induction of selective cytotoxicity in cancer cells that express the target RNA, in animal models as well as in cell cultures. In this study, we evaluated the target RNA specificity of trans-splicing phenomena by the group I intron in mice that were intraperitoneally inoculated with hTERT-expressing human cancer cells to validate the anti-cancer therapeutic applicability of the group I intron. To this end, an adenoviral vector that encoded for the hTERT-targeting group I intron was constructed and systemically injected into the animal. 5'-end RACE-PCR and sequencing analyses of the trans-spliced cDNA clones revealed that all of the analyzed products in the tumor tissue of the virus-infected mice resulted from reactions that were generated only with the targeted hTERT RNA. This study implies the in vivo target specificity of the trans-splicing group I intron and hence suggests that RNA replacement via a trans-splicing reaction by the group I intron is a potent anti-cancer genetic approach.

Suppressive Effects of Divalent Cations on Self-splicing Inhibition by Spectinomycin of Group 1 Intron RNA

  • Park, In-Kook
    • Journal of Microbiology
    • /
    • 제37권4호
    • /
    • pp.243-247
    • /
    • 1999
  • Effects of divalent cations on self-splicing inhibition by the antibiotic spectinomycin of the phage T4 thymidylate synthase intron (td) have been investigated. $Ca^{2+}$ ion at 1mM concentration suppressed splicing inhibition of spectinomycin by 10% and 50 ${\mu}M\;Co^{2+}$ ion also suppressed splicing inhibition of specinomycin by 10%. $Mg^{2+}$ ion at 6 mM concentration decreased splicing inhibition of spectinomycin by 42% while $Mn^{2+}$ ion decreased the splicing inhibition by 10%. $Zn^{2+}$ ion at 10 uM concentration lowered the splicing inhibition by spectinomycin of 15%. Of all divalent cations tested, $Mg^{2+}$ ion was the most effective in suppressing splicing inhibition by specinomycin whereas $Ca^{2+}$ ion was the least effective. The results suggest that spectinomycin may interact with specific and functional $Mg^{2+}$-binding sites within intron RNA that lead to a displacement of $Mg^{2+}$ essential for catalytic activity.

  • PDF