Proteomic analysis of the cullin 4B interactome using proximity‐dependent biotinylation in living cells. Issue 8 (April 2017)

• Record Type: Journal Article
• Authors: Zhang, Hailong; Li, Shupeng; Liu, Pingting; Lee, Frankie H. F.; Wong, Albert H. C.; Liu, Fang
• Is Part Of: Proteomics. Volume 17:Issue 8(2017)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2017-04
• Subjects: Biotinylation -- Cellular stimuli -- CRLs proximal labeling -- Interactants -- Protein dynamics; Proteins -- Separation -- Periodicals; Bioinformatics -- Periodicals; Proteomics -- Periodicals; Genomes -- Periodicals; Molecular genetics -- Periodicals; 572.605
• DOI: 10.1002/pmic.201600163
• Languages: English
• ISSNs: 1615-9853

Abstract

Abstract : Cullin 4B (CUL4B) mutations have been implicated in mental retardation and dopamine‐related behaviors due to disruptions in their interaction with cullin‐RING E3 ligases (CRLs). Thus, further identification of CUL4B substrates can increase the knowledge of protein homeostasis and illuminate the role of CUL4B in neuropsychiatric disease. However, the transient nature of the coupling between CUL4B and its substrates is difficult to detect in vivo using current approaches, thus hampers efforts to investigate functions of CRLs within unperturbed living systems. In this study, we sought to discover CUL4B interactants with or without dopamine stimulation. BirA (118G) proximity‐dependent biotin labeling combined with LC‐MS was employed to biotinylate and identify transient and weak interactants of CUL4B. After purification with streptavidin beads and identified by LC‐MS, a total of 150 biotinylated proteins were identified at baseline condition, 53 of which are well‐known CUL4B interactants. After dopamine stimulation, 29 proteins disappeared and were replaced by 21 different protein interactants. The altered CUL4B interactants suggest that CUL4B regulates protein turnover and homeostasis in response to dopamine stimulation. Our results demonstrate the potential of this approach to identify novel CUL4B‐related molecules in respond to cellular stimuli, which may be applied to other types of signaling pathways.