Articles by Jeremy Z.R. Han in JoVE
Dissecting Multi-protein Signaling Complexes by Bimolecular Complementation Affinity Purification (BiCAP) Jordan F. Hastings1, Jeremy Z.R. Han1, Robert F. Shearer1,2, Sean P. Kennedy1,3, Mary Iconomou1,4, Darren N. Saunders5, David R. Croucher1,6,7 1The Kinghorn Cancer Centre, Garvan Institute of Medical Research, 2Ubiquitin Signaling Group, Protein Signaling Program, The Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, 3RCSI Molecular Medicine, Royal College of Surgeons in Ireland, 4Department of Epigenetics, Max Planck Institute of Immunobiology and Epigenetics, 5School of Medical Sciences, University of New South Wales, 6St Vincent's Hospital Clinical School, University of New South Wales, 7School of Medicine and Medical Science, University College Dublin This manuscript describes the protocol for Bimolecular Complementation Affinity Purification (BiCAP). This novel method facilitates the specific isolation and downstream proteomic characterization of any two interacting proteins, while excluding un-complexed individual proteins as well as complexes formed with competing binding partners.
Other articles by Jeremy Z.R. Han on PubMed
Bimolecular Complementation Affinity Purification (BiCAP) Reveals Dimer-specific Protein Interactions for ERBB2 Dimers Science Signaling. 07, 2016 | Pubmed ID: 27405979 The dynamic assembly of multiprotein complexes is a central mechanism of many cell signaling pathways. This process is key to maintaining the spatiotemporal specificity required for an accurate, yet adaptive, response to rapidly changing cellular conditions. We describe a technique for the specific isolation and downstream proteomic characterization of any two interacting proteins, to the exclusion of their individual moieties and competing binding partners. We termed the approach bimolecular complementation affinity purification (BiCAP) because it combines the use of conformation-specific nanobodies with a protein-fragment complementation assay with affinity purification. Using BiCAP, we characterized the specific interactome of the epidermal growth factor receptor (EGFR) family member ERBB2 when in the form of a homodimer or when in the form of a heterodimer with either EGFR or ERBB3. We identified dimer-specific interaction patterns for key adaptor proteins and identified a number of previously unknown interacting partners. Functional analysis for one of these newly identified partners revealed a noncanonical mechanism of extracellular signal-regulated kinase (ERK) activation that is specific to the ERBB2:ERBB3 heterodimer and acts through the adaptor protein FAM59A in breast cancer cells.
The Under-Appreciated Promiscuity of the Epidermal Growth Factor Receptor Family Frontiers in Cell and Developmental Biology. 2016 | Pubmed ID: 27597943 Each member of the epidermal growth factor receptor (EGFR) family plays a key role in normal development, homeostasis, and a variety of pathophysiological conditions, most notably in cancer. According to the prevailing dogma, these four receptor tyrosine kinases (RTKs; EGFR, ERBB2, ERBB3, and ERBB4) function exclusively through the formation of homodimers and heterodimers within the EGFR family. These combinatorial receptor interactions are known to generate increased interactome diversity and therefore influence signaling output, subcellular localization and function of the heterodimer. This molecular plasticity is also thought to play a role in the development of resistance toward targeted cancer therapies aimed at these known oncogenes. Interestingly, many studies now challenge this dogma and suggest that the potential for EGFR family receptors to interact with more distantly related RTKs is much greater than currently appreciated. Here we discuss how the promiscuity of these oncogenic receptors may lead to the formation of many unexpected receptor pairings and the significant implications for the efficiency of many targeted cancer therapies.