Related Experiment Video

Updated: Jan 21, 2026

Single Molecule Fluorescence In Situ Hybridization smFISH Analysis in Budding Yeast Vegetative Growth and Meiosis
09:28

Single Molecule Fluorescence In Situ Hybridization smFISH Analysis in Budding Yeast Vegetative Growth and Meiosis

Published on: May 25, 2018

20.2K

Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization

Gal Haimovich1, Jeffrey E Gerst2

  • 1Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel. gal.haimovich@weizmann.ac.il.

Methods in Molecular Biology (Clifton, N.J.)
|August 14, 2019
PubMed

Insights

Researchers developed a method to track mRNA transfer between eukaryotic cells using single molecule fluorescent in situ hybridization (smFISH). This technique distinguishes between transfer via membrane nanotubes and extracellular vesicles (EVs).

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Genetics

Background:

  • Eukaryotic cells can transfer messenger RNA (mRNA) molecules between cells.
  • This intercellular mRNA transfer is thought to occur via membrane nanotubes or extracellular vesicles (EVs).
  • Understanding mRNA transfer mechanisms is crucial for comprehending cell-to-cell communication.

Purpose of the Study:

  • To describe a novel method for detecting and quantifying transferred mRNA in co-cultured cells.
  • To differentiate between mRNA transfer mediated by membrane nanotubes and extracellular vesicles (EVs).
  • To provide a protocol adaptable for studying most mRNA transcripts in various co-culture systems.

Main Methods:

  • Utilized single molecule fluorescent in situ hybridization (smFISH) for accurate mRNA detection and localization.
  • Developed a co-culture system with two distinct cell types to observe intercellular transfer.
  • Optimized smFISH procedures to preserve membrane nanotube structures.

Main Results:

  • Successfully detected and quantified transferred mRNA molecules between co-cultured cells.
  • Differentiated between nanotube-mediated and EV-mediated mRNA transfer pathways.
  • Demonstrated the method's applicability to various mRNA transcripts and cell types.

Conclusions:

  • The described smFISH-based method provides a robust tool for investigating mRNA transfer mechanisms.
  • This technique facilitates the study of cell-to-cell communication through mRNA exchange.
  • The protocol advancements enhance the preservation of cellular structures critical for nanotube-mediated transfer analysis.
Keywords:
ExosomesExtracellular vesiclesGlutaraldehydeMembrane nanotubesSingle-molecule fluorescent in situ hybridizationmRNAβ-Actin

More Related Videos

Probing mRNA Kinetics in Space and Time in Escherichia coli using Two-Color Single-Molecule Fluorescence In Situ Hybridization10:01

Probing mRNA Kinetics in Space and Time in Escherichia coli using Two-Color Single-Molecule Fluorescence In Situ Hybridization

Published on: July 30, 2020

7.8K
Visualization and Analysis of mRNA Molecules Using Fluorescence In Situ Hybridization in Saccharomyces cerevisiae07:00

Visualization and Analysis of mRNA Molecules Using Fluorescence In Situ Hybridization in Saccharomyces cerevisiae

Published on: June 14, 2013

35.5K

Related Concept Videos

FISH - Fluorescent In-situ Hybridization02:07

FISH - Fluorescent In-situ Hybridization

Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...
23.9K
In-situ Hybridization02:31

In-situ Hybridization

In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
Types of probes and labels
A probe is a complementary strand of DNA or RNA that binds to corresponding nucleotide sequences in a cell. Many...
10.4K
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...
57.2K
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...
8.7K
Nonsense-mediated mRNA Decay02:27

Nonsense-mediated mRNA Decay

The Upf proteins that carry out nonsense-mediated decay (NMD) are found in all eukaryotic organisms, including humans. Each protein has an individual role, but they need to work in collaboration. Upf1 is an ATP-dependent RNA helicase that unwinds the RNA helix. Because Upf1 can unwind any RNA, Upf2 and Upf3 are required to help Upf1 discriminate between nonsense and normal mRNAs.
Usually, Upf3 binds to an Exon Junction Complex (EJC) at mRNA splice sites. If a ribosome fully translates the mRNA,...
11.7K
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...
6.9K