Related Experiment Video

Updated: Mar 10, 2026

Detection of Heterodimerization of Protein Isoforms Using an in Situ Proximity Ligation Assay
09:18

Detection of Heterodimerization of Protein Isoforms Using an in Situ Proximity Ligation Assay

Published on: October 20, 2018

8.0K

Visualization of RAS/MAPK Signaling In Situ by the Proximity Ligation Assay (PLA)

Zijian Tang1,2, Chengkai Dai3

  • 1The Jackson Laboratory, 600 Main Street, Bar Harbor, ME, 04609, USA.

Methods in Molecular Biology (Clifton, N.J.)
|December 8, 2016
PubMed

Insights

This study introduces a new method using Proximity Ligation Assay (PLA) to track RAS/MAPK pathway activation by observing kinase-substrate interactions in cells. This technique helps understand how this important signaling pathway functions and malfunctions in diseases like cancer.

Area of Science:

  • Cellular signaling pathways
  • Molecular biology
  • Cancer research

Background:

  • The RAS/MAPK signaling pathway is crucial for cellular functions and is often dysregulated in human diseases, particularly cancer.
  • Understanding the activation mechanisms of this pathway is vital for disease research and therapeutic development.

Purpose of the Study:

  • To present a novel methodology for studying the activation of the RAS/MAPK signaling pathway.
  • To demonstrate the application of Proximity Ligation Assay (PLA) for in situ monitoring of kinase-substrate interactions within this pathway.

Main Methods:

  • Utilized Proximity Ligation Assay (PLA) to visualize and quantify protein interactions.
  • Focused on monitoring kinase-substrate interactions, specifically between MEK1 and HSF1, and MEK1 and ERK1.
  • Applied the assay in situ to study interactions within their native cellular environment.

Main Results:

  • Successfully employed PLA to detect and study specific kinase-substrate interactions within the RAS/MAPK pathway.
  • Demonstrated the feasibility of using PLA to monitor MEK1 interactions with its substrates HSF1 and ERK1.
  • Provided a novel in situ method for assessing pathway activation.

Conclusions:

  • Proximity Ligation Assay (PLA) offers a powerful new tool for studying RAS/MAPK pathway activation.
  • This methodology enables detailed in situ analysis of kinase-substrate dynamics.
  • The developed technique has significant implications for cancer research and understanding signaling pathway dysregulation.
Keywords:
ERKFluorescence imagingHSF1MAPK signalingMEKPLA

More Related Videos

Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay13:10

Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay

Published on: June 9, 2017

10.7K
Detection of Signaling Effector-Complexes Downstream of BMP4 Using in situ PLA, a Proximity Ligation Assay12:52

Detection of Signaling Effector-Complexes Downstream of BMP4 Using in situ PLA, a Proximity Ligation Assay

Published on: March 3, 2011

32.2K

Related Concept Videos

MAPK Signaling Cascades01:07

MAPK Signaling Cascades

Mitogen-activated protein kinase, or MAPK pathway, activates three sequential kinases to regulate cellular responses such as proliferation, differentiation, survival, and apoptosis. The canonical MAPK pathway starts with a mitogen or growth factor binding to an RTK. The activated RTKs stimulate Ras, which recruits Raf or MAP3 Kinase (MAPKKK), the first kinase of the MAPK signaling cascade. Raf further phosphorylates and activates MEK or MAP2 Kinases (MAPKK), which in turn phosphorylates MAP...
9.0K
The Ras Gene02:38

The Ras Gene

The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a...
7.4K
Small GTPases - Ras and Rho01:24

Small GTPases - Ras and Rho

Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
5.6K