Related Experiment Video

Updated: Jun 26, 2026

Analysis of Spliceosomal snRNA Localization in Human Hela Cells Using Microinjection
07:35

Analysis of Spliceosomal snRNA Localization in Human Hela Cells Using Microinjection

Published on: August 6, 2019

Subnuclear compartmentalization of transiently expressed polyadenylated pri-microRNAs: processing at transcription

Jan M Pawlicki1, Joan A Steitz

  • 1Department of Pharmacology, Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, Connecticut 06536, USA.

Cell Cycle (Georgetown, Tex.)
|January 30, 2009
PubMed

Insights

Primary miRNA transcripts (pri-miRNAs) processing is coupled to transcription, not nuclear foci. Efficient processing requires co-transcriptional localization, indicating transcription-coupled miRNA biogenesis.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • MicroRNAs (miRNAs) are key regulators of gene expression.
  • Recent studies suggest primary miRNA transcripts (pri-miRNAs) processing is linked to transcription.
  • Pri-miRNAs and processing factors co-localize in nuclear foci.

Purpose of the Study:

  • To investigate the role of nuclear foci in pri-miRNA processing.
  • To determine if pri-miRNA processing is spatially coupled to transcription.
  • To elucidate the composition and function of pri-miRNA/SC35 nuclear foci.

Main Methods:

  • Utilized a self-cleaving hepatitis delta ribozyme to disrupt pri-miRNA processing and localization.
  • Analyzed pri-miRNA localization in nuclear foci using immunofluorescence.
  • Examined co-localization of pri-miRNAs with splicing factors (SC35) and Microprocessor components (Drosha, DGCR8).
  • Investigated the presence of other nuclear proteins within pri-miRNA/SC35 foci.

Main Results:

  • Ribozyme-mediated release of pri-miRNAs from transcription sites led to nucleoplasmic accumulation and inefficient processing.
  • Pri-miRNAs released from transcription sites did not localize to SC35 foci.
  • Cleaved and polyadenylated pri-miRNAs, with or without the pre-miRNA hairpin, localized to SC35 foci.
  • Pri-miRNA/SC35 foci contained splicing-associated proteins but not RNA polymerase II or PM/Scl-100.

Conclusions:

  • Pri-miRNA/SC35 nuclear foci are not primary sites for pri-miRNA processing.
  • Pri-miRNA processing is tightly coupled to transcription, not post-transcriptional events in nuclear foci.
  • Findings provide insights into miRNA biogenesis, mRNA metabolism, and nuclear gene expression organization.

More Related Videos

A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells06:48

A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells

Published on: June 16, 2022

Quantitative Immunofluorescence to Measure Global Localized Translation09:13

Quantitative Immunofluorescence to Measure Global Localized Translation

Published on: August 22, 2017

Related Concept Videos

Chromatin Structure Regulates pre-mRNA Processing02:41

Chromatin Structure Regulates pre-mRNA Processing

In eukaryotic cells, nascent mRNA transcripts need to undergo many post-transcriptional modifications to reach the cell cytoplasm and translate into functional proteins. For a long time, transcription and pre-mRNA processing were considered two independent events that occur sequentially in the cell. However, it has now been well established that transcription and pre-mRNA processing are two simultaneous processes that are precisely regulated inside the cell.
The chromatin structure, especially...
Regulation of Nuclear Protein Sorting01:45

Regulation of Nuclear Protein Sorting

Nuclear protein sorting regulates nucleus composition and gene expression, crucial for determining the fate of a eukaryotic cell. Hence, the entry and exit of molecules across the nuclear envelope is a tightly controlled process. Nuclear protein sorting can be inhibited by one of the following ways: 1) masking cargo signal sequences, 2) modifying the nuclear receptor's affinity for cargo, 3) controlling the nuclear pore size, 4) retaining the cargo during its transit to the cytosol or the...
Additional Subnuclear Structures02:10

Additional Subnuclear Structures

The eukaryotic nucleus is a double membrane-bound organelle that contains nearly all of the cell’s genetic material in the form of chromosomes. It is rightly called the “brain” of the cell as it shoulders the responsibility of responding to various physiological processes, stress, altered metabolic conditions, and other cellular signals. 
The nucleus contains many membrane-less subnuclear organelles or nuclear bodies, such as nucleoli, Cajal bodies, speckles, paraspeckles, etc. These nuclear...
Nuclear Export of mRNA02:31

Nuclear Export of mRNA

Before mRNAs are exported to the cytoplasm, it is crucial to check each mRNA for structural and functional integrity. Eukaryotic cells use several different mechanisms, collectively known as mRNA surveillance, to look for irregularities in mRNAs. Irregular or aberrant mRNA are rapidly degraded by various enzymes. If a defective mRNA escapes the surveillance, it would be translated into a protein which would either be non-functional or not function properly. One of the primary irregularities in...
Nuclear Export of mRNA02:31

Nuclear Export of mRNA

Before mRNAs are exported to the cytoplasm, it is crucial to check each mRNA for structural and functional integrity. Eukaryotic cells use several different mechanisms, collectively known as mRNA surveillance, to look for irregularities in mRNAs. Irregular or aberrant mRNA are rapidly degraded by various enzymes. If a defective mRNA escapes the surveillance, it would be translated into a protein which would either be non-functional or not function properly. One of the primary irregularities in...
Regulated mRNA Transport02:22

Regulated mRNA Transport

In eukaryotes, transcription and translation are compartmentalized; an mRNA is first synthesized in the nucleus and then selectively transported to the cytoplasm for protein synthesis. Before transport, a pre-mRNA undergoes several steps of post-transcriptional modifications including splicing, 5' capping, and the addition of a poly-adenine tail. Various proteins bind to the pre-mRNA during these modifications. The mRNA transport takes place with the help of multiple proteins playing specific...