Type

Journal Article

Authors

Jochen H M Prehn
Michael Scannell
Beau J Fenner

Subjects

Biochemistry

Topics
nf kappa b signal transduction protein binding metabolism dna primers base sequence cell line i kappa b kinase polymerase chain reaction ikbkg protein human humans recombinant proteins transcriptional activation protein array analysis

Expanding the substantial interactome of NEMO using protein microarrays. (2009)

Abstract Signal transduction by the NF-kappaB pathway is a key regulator of a host of cellular responses to extracellular and intracellular messages. The NEMO adaptor protein lies at the top of this pathway and serves as a molecular conduit, connecting signals transmitted from upstream sensors to the downstream NF-kappaB transcription factor and subsequent gene activation. The position of NEMO within this pathway makes it an attractive target from which to search for new proteins that link NF-kappaB signaling to additional pathways and upstream effectors. In this work, we have used protein microarrays to identify novel NEMO interactors. A total of 112 protein interactors were identified, with the most statistically significant hit being the canonical NEMO interactor IKKbeta, with IKKalpha also being identified. Of the novel interactors, more than 30% were kinases, while at least 25% were involved in signal transduction. Binding of NEMO to several interactors, including CALB1, CDK2, SAG, SENP2 and SYT1, was confirmed using GST pulldown assays and coimmunoprecipitation, validating the initial screening approach. Overexpression of CALB1, CDK2 and SAG was found to stimulate transcriptional activation by NF-kappaB, while SYT1 overexpression repressed TNFalpha-dependent NF-kappaB transcriptional activation in human embryonic kidney cells. Corresponding with this finding, RNA silencing of CDK2, SAG and SENP2 reduced NF-kappaB transcriptional activation, supporting a positive role for these proteins in the NF-kappaB pathway. The identification of a host of new NEMO interactors opens up new research opportunities to improve understanding of this essential cell signaling pathway.
Collections Ireland -> Royal College of Surgeons in Ireland -> PubMed

Full list of authors on original publication

Jochen H M Prehn, Michael Scannell, Beau J Fenner

Experts in our system

1
Jochen H M Prehn
Royal College of Surgeons in Ireland
Total Publications: 206
 
2
Beau J Fenner
Royal College of Surgeons in Ireland
Total Publications: 9