Cellular localization of activated N-WASP using a conformation-sensitive antibody

Cell Motility and the Cytoskeleton
P SukumvanichJohn Condeelis

Abstract

The main regulators of Arp2/3 activity appear to be N-WASP and the other members of the Scar/WAVE family of proteins. We show here that after EGF stimulation, N-WASP is recruited to the nucleation zone of the dynamic leading edge compartment of carcinoma cells, with maximal recruitment of N-WASP within 1 min after EGF stimulation. The timing of N-WASP recruitment mirrors the timing of barbed-end formation at the leading edge. To determine the cellular activation of N-WASP after EGF stimulation, we made a conformation-sensitive antibody (CSA) against the CRIB domain of N-WASP that is predicted to recognize N-WASP in its open, active conformation, but not in its closed, inactive conformation. The ability of CSA to detect only active N-WASP was demonstrated by in vitro experiments using immunoprecipitation of active N-WASP from EGF-stimulated cells and Cdc42 activation of N-WASP activity. In cell staining experiments, N-WASP is maximally accessible to CSA 40 sec after EGF stimulation and this activated N-WASP is in the nucleation zone. These results indicate that active N-WASP is present at the leading edge of lamellipods, an unexpected finding given its reported involvement in filopod formation. This work establishes the feasibil...Continue Reading

References

Apr 13, 1978·Nature·J V SmallJ E Celis
Sep 1, 1992·The Journal of Experimental Medicine·I J MolinaE Remold-O'Donnell
Aug 1, 1991·The Journal of Cell Biology·M H Symons, T J Mitchison
Jul 1, 1995·Critical Reviews in Oncology/hematology·D S SalomonN Normanno
Jul 21, 1995·The Journal of Biological Chemistry·B J AgnewJ R Bamburg
Jan 1, 1994·Breast Cancer Research and Treatment·S A Chrysogelos, R B Dickson
Jan 15, 1994·International Journal of Cancer. Journal International Du Cancer·P H PedersenO D Laerum
Mar 13, 1998·Annals of Oncology : Official Journal of the European Society for Medical Oncology·B R VoldborgH S Poulsen
May 30, 1998·Proceedings of the National Academy of Sciences of the United States of America·R D MullinsT D Pollard
Dec 19, 1998·Microscopy Research and Technique·M BaillyJ E Segall
Mar 31, 1999·Proceedings of the National Academy of Sciences of the United States of America·L M MacheskyT D Pollard
Nov 5, 1999·Advances in Cancer Research·A Wells
Apr 1, 2000·Cell Motility and the Cytoskeleton·I IchetovkinJ S Condeelis
Sep 20, 2000·The Journal of Cell Biology·R RohatgiM W Kirschner
Nov 10, 2000·Molecular Biology of the Cell·M BaillyJ E Segall
Feb 24, 2001·Ultramicroscopy·C RotschM Radmacher
Mar 27, 2001·Current Biology : CB·A M WeaverJ A Cooper
May 2, 2001·Nature Cell Biology·N Martinez-QuilesN Ramesh
Aug 30, 2001·Current Biology : CB·L BlanchoinS E Hitchcock-DeGregori
Jul 31, 2002·The Journal of Biological Chemistry·Stefanie BeneschKlemens Rottner
Nov 5, 2002·Journal of Cell Science·Vera DesMaraisSarah E Hitchcock-DeGregori
Mar 21, 2003·Current Biology : CB·Fanny S ChangKendall J Blumer

❮ Previous
Next ❯

Citations

Jul 4, 2007·Proceedings of the National Academy of Sciences of the United States of America·Shandiz TehraniJohn A Cooper
Apr 5, 2014·Human Molecular Genetics·Ana Franco-VillanuevaMaría Dolores Ledesma
Jul 24, 2010·Journal of Oncology·Jessie ZhongGeraldine M O'Neill
Sep 1, 2004·The Journal of Cell Biology·Ghassan MouneimneJohn Condeelis
Mar 26, 2008·The Journal of Cell Biology·Corina SarmientoJohn Condeelis
Jan 23, 2008·The New Phytologist·Wing-Yen Francisca LiHon-Ming Lam
Jun 3, 2006·European Journal of Cell Biology·Atsuo T Sasaki, Richard A Firtel
Sep 7, 2016·The Journal of Clinical Investigation·Erin JanssenRaif S Geha
Nov 10, 2005·The Journal of Biological Chemistry·Carlos A BuscagliaVictor Nussenzweig

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.