Related Experiment Video

Updated: Jul 2, 2026

Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA
12:19

Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA

Published on: February 12, 2020

Knockdown endogenous CypA with siRNA in U2OS cells results in disruption of F-actin structure and alters tumor

Colonya C Calhoun1, Ying-Chun Lu, Jun Song

  • 1Dental Research Institute, UCLA School of Dentistry, Los Angeles, CA 90095, USA.

Molecular and Cellular Biochemistry
|August 16, 2008
PubMed

Insights

Cyclophilin A (CypA) stabilizes N-WASP, crucial for F-actin formation. Lacking CypA disrupts actin structure and cell growth, highlighting CypA

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Cyclophilin A (CypA) is known for its peptidyl-prolyl isomerase activity and role in immune responses.
  • Other molecular mechanisms of CypA-mediated biological events remain largely unexplored.
  • Understanding CypA's functions beyond immunity is critical for its broader biological significance.

Purpose of the Study:

  • To investigate the non-immune functions of Cyclophilin A (CypA).
  • To elucidate the molecular mechanisms underlying CypA's role in cellular processes.
  • To determine if CypA's isomerase activity is essential for maintaining normal cell phenotype.

Main Methods:

  • RNA interference (RNAi) was used to knock down CypA expression in U2OS cells.
  • Cellular phenotypes including F-actin structure, anchorage-independent growth, proliferation, and migration were assessed.
  • Cyclosporine A (CsA), a peptidyl-prolyl isomerase inhibitor, was used to evaluate the role of isomerase activity.
  • In vitro and in vivo binding assays were performed to identify CypA-interacting proteins.
  • Pulse-chase labeling was employed to study protein degradation rates.

Main Results:

  • Knockdown of CypA in U2OS cells led to F-actin disruption, decreased anchorage-independent growth, proliferation, and migration.
  • Treatment of wild-type cells with CsA mimicked the CypA knockdown phenotype, indicating the necessity of CypA's isomerase activity.
  • CypA was found to bind to N-WASP, a key regulator of actin nucleation via the Arp2/3 complex.
  • Cells lacking CypA exhibited enhanced degradation of N-WASP.
  • CypA is required for stabilizing N-WASP, facilitating the formation of the N-WASP/Arp2/3 complex for F-actin polymerization.

Conclusions:

  • CypA's peptidyl-prolyl isomerase activity is essential for maintaining normal cellular phenotype, including F-actin organization and cell proliferation.
  • CypA plays a critical role in actin dynamics by stabilizing N-WASP, thereby promoting the formation of the N-WASP/Arp2/3 complex.
  • This stabilization mechanism is crucial for the nucleation and initiation of F-actin polymerization, impacting cell growth and migration.

More Related Videos

Knockdown of FAM83A to Verify Its Role in Cervical Cancer Cell Growth and Cisplatin Sensitivity04:20

Knockdown of FAM83A to Verify Its Role in Cervical Cancer Cell Growth and Cisplatin Sensitivity

Published on: February 9, 2024

Long-term Silencing of Intersectin-1s in Mouse Lungs by Repeated Delivery of a Specific siRNA via Cationic Liposomes. Evaluation of Knockdown Effects by Electron Microscopy15:55

Long-term Silencing of Intersectin-1s in Mouse Lungs by Repeated Delivery of a Specific siRNA via Cationic Liposomes. Evaluation of Knockdown Effects by Electron Microscopy

Published on: June 21, 2013

Related Concept Videos

Experimental RNAi02:15

Experimental RNAi

RNA interference (RNAi) is a cellular mechanism that inhibits gene expression by suppressing its transcription or activating the RNA degradation process. The mechanism was discovered by Andrew Fire and Craig Mello in 1998 in plants. Today, it is observed in almost all eukaryotes, including protozoa, flies, nematodes, insects, parasites, and mammals. This precise cellular mechanism of gene silencing has been developed into a technique that provides an efficient way to identify and determine the...
Inhibition of Cdk Activity02:34

Inhibition of Cdk Activity

The orderly progression of the cell cycle depends on the activation of Cdk protein by binding to its cyclin partner. However, the cell cycle must be restricted when undergoing abnormal changes. Most cancers correlate to the deregulated cell cycle, and since Cdks are a central component of the cell cycle, Cdk inhibitors are extensively studied to develop anticancer agents. For instance, cyclin D associates with several Cdks, such as Cdk 4/6, to form an active complex. The cyclin D-Cdk4/6 complex...