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Related Concept Videos

pre-mRNA Processing02:01

pre-mRNA Processing

In eukaryotic cells, transcripts made by RNA polymerase are modified and processed before exiting the nucleus. Unprocessed RNA is called precursor mRNA or pre-mRNA to distinguish it from mature mRNA.
Once about 20-40 ribonucleotides have been joined together by RNA polymerase, a group of enzymes adds a “cap” to the 5’ end of the growing transcript. In this process, a 5’ phosphate is replaced by modified guanosine that has a methyl group attached to it (7-Methyl guanosine). This 5’ cap helps the...
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MicroRNA (miRNA) are short, regulatory RNA transcribed from introns—non-coding regions of a gene—or intergenic regions—stretches of DNA present between genes. Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA ends...
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MicroRNAs

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In eukaryotic cells, transcripts made by RNA polymerase are modified and processed before exiting the nucleus. Unprocessed RNA is called precursor mRNA or pre-mRNA to distinguish it from mature mRNA.
Once about 20-40 ribonucleotides have been joined together by RNA polymerase, a group of enzymes adds a cap to the 5' end of the growing transcript. In this process, a 5' phosphate is replaced by modified guanosine that has a methyl group attached (7-methyl guanosine). This 5' cap helps the cell...
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First step in pre-miRNAs processing by human Dicer.

Carlos Fabián Flores-Jasso1, Catalina Arenas-Huertero, Jose Luis Reyes

  • 1Departamento de Biología Celular, Instituto de Fisiología Celular, UNAM Apartado Postal 70-600, Circuito exterior s/n Ciudad Universitaria, Delegación Coyoacan, CP 04510, México DF.

Acta Pharmacologica Sinica
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PubMed
Summary

Human Dicer enzyme shows strand preference during the initial cleavage of pre-microRNAs (miRNAs). This specific cleavage bias was observed for both recombinant Dicer and in cellular extracts, impacting miRNA biogenesis.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • RNA Biology

Background:

  • MicroRNAs (miRNAs) are key regulators of gene expression.
  • Dicer is a critical enzyme in miRNA biogenesis, responsible for processing precursor miRNAs (pre-miRNAs) into mature miRNAs.
  • Understanding the initial cleavage events by Dicer is crucial for elucidating miRNA processing pathways.

Purpose of the Study:

  • To investigate the strand preference of the initial cleavage of human pre-miRNAs by human Dicer.
  • To determine if this cleavage preference is conserved between recombinant Dicer and cellular extracts.

Main Methods:

  • In vitro transcription of pre-miRNAs labeled at the 5' or 3' ends.
  • Cleavage assays using recombinant human Dicer and HeLa cytoplasmic S100 extracts.
  • Purification of pre-miRNA stem-loop structures and resolution of cleavage products by denaturing polyacrylamide gel electrophoresis (PAGE).

Main Results:

  • Recombinant human Dicer exhibited a biased initial cleavage of the 3' strand for pre-let-7b and hsa-pre-miR-17.
  • This strand-biased cleavage was also observed when using HeLa S100 cytoplasmic extracts.
  • The observed bias was specific to the stem-loop structure of the pre-miRNAs.

Conclusions:

  • Human Dicer demonstrates strand preference during the initial cleavage of specific pre-miRNAs.
  • This differential first-nick is dependent on the substrate being a stem-loop structure.
  • The findings suggest a regulated step in miRNA processing that may influence strand selection for the guide miRNA.