scholarly journals A long non-coding RNA is required for targeting centromeric protein A to the human centromere

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Delphine Quénet ◽  
Yamini Dalal
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Biochemical Characterization ◽  
Protein A ◽  
Human Centromere ◽  
Non Coding Rna ◽  
Histone H3 Variant ◽  
Centromeric Protein ◽  
Long Non Coding Rna

The centromere is a specialized chromatin region marked by the histone H3 variant CENP-A. Although active centromeric transcription has been documented for over a decade, the role of centromeric transcription or transcripts has been elusive. Here, we report that centromeric α-satellite transcription is dependent on RNA Polymerase II and occurs at late mitosis into early G1, concurrent with the timing of new CENP-A assembly. Inhibition of RNA Polymerase II-dependent transcription abrogates the recruitment of CENP-A and its chaperone HJURP to native human centromeres. Biochemical characterization of CENP-A associated RNAs reveals a 1.3 kb molecule that originates from centromeres, which physically interacts with the soluble pre-assembly HJURP/CENP-A complex in vivo, and whose down-regulation leads to the loss of CENP-A and HJURP at centromeres. This study describes a novel function for human centromeric long non-coding RNAs in the recruitment of HJURP and CENP-A, implicating RNA-based chaperone targeting in histone variant assembly.

scholarly journals Functional dissection of the catalytic mechanism of mammalian RNA polymerase II

2005 ◽  
Vol 83 (4) ◽  
pp. 497-504 ◽  
Author(s):  
Benoit Coulombe ◽  
Marie-France Langelier
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Catalytic Mechanism ◽  
Mutational Analysis ◽  
Structural Elements ◽  
Catalytic Mechanisms ◽  
Rnap Ii ◽  
Affinity Peptide

High resolution X-ray crystal structures of multisubunit RNA polymerases (RNAP) have contributed to our understanding of transcriptional mechanisms. They also provided a powerful guide for the design of experiments aimed at further characterizing the molecular stages of the transcription reaction. Our laboratory used tandem-affinity peptide purification in native conditions to isolate human RNAP II variants that had site-specific mutations in structural elements located strategically within the enzyme's catalytic center. Both in vitro and in vivo analyses of these mutants revealed novel features of the catalytic mechanisms involving this enzyme.Key words: RNA polymerase II, transcriptional mechanisms, mutational analysis, mRNA synthesis.

scholarly journals LncRNA PVT1 promotes cervical cancer progression by sponging miR-503 to upregulate ARL2 expression

2021 ◽  
Vol 16 (1) ◽  
pp. 1-13
Author(s):  
Weiwei Liu ◽  
Dongmei Yao ◽  
Bo Huang
Keyword(s):  
Cervical Cancer ◽  
Cancer Progression ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Nude Mouse Model ◽  
Western Blot Assay ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Abstract Cervical cancer (CC) is a huge threat to the health of women worldwide. Long non-coding RNA plasmacytoma variant translocation 1 gene (PVT1) was proved to be associated with the development of diverse human cancers, including CC. Nevertheless, the exact mechanism of PVT1 in CC progression remains unclear. Levels of PVT1, microRNA-503 (miR-503), and ADP ribosylation factor-like protein 2 (ARL2) were measured by quantitative reverse transcription-polymerase chain reaction or western blot assay. 3-(4,5)-Dimethylthiazole-2-y1)-2,5-biphenyl tetrazolium bromide (MTT) and flow cytometry were used to examine cell viability and apoptosis, respectively. For migration and invasion detection, transwell assay was performed. The interaction between miR-503 and PVT1 or ARL2 was shown by dual luciferase reporter assay. A nude mouse model was constructed to clarify the role of PVT1 in vivo. PVT1 and ARL2 expressions were increased, whereas miR-503 expression was decreased in CC tissues and cells. PVT1 was a sponge of miR-503, and miR-503 targeted ARL2. PVT1 knockdown suppressed proliferation, migration, and invasion of CC cells, which could be largely reverted by miR-503 inhibitor. In addition, upregulated ARL2 could attenuate si-PVT1-mediated anti-proliferation and anti-metastasis effects on CC cells. Silenced PVT1 also inhibited CC tumor growth in vivo. PVT1 knockdown exerted tumor suppressor role in CC progression via the miR-503/ARL2 axis, at least in part.

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