Binding of JAB1/CSN5 to MIF is mediated by the MPN domain but is independent of the JAMM motif

Abstract

Macrophage migration inhibitory factor (MIF) binds to c-Jun activation domain binding protein-1 (JAB1)/subunit 5 of COP9 signalosome (CSN5) and modulates cell signaling and the cell cycle through JAB1. The binding domain of JAB1 responsible for binding to MIF is unknown. We hypothesized that the conserved Mpr1p Pad1p N-terminal (MPN) domain of JAB1 may mediate binding to MIF. In fact, yeast two hybrid (YTH) and in vitro translation/coimmunoprecipitation (CoIP) analysis showed that a core MPN domain, which did not cover the functional JAB1/MPN/Mov34 metalloenzyme (JAMM) deneddylase sequence, binds to MIF comparable to full-length JAB1. YTH and pull-down analysis in conjunction with nanobead affinity matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry demonstrated that MIF(50-65) and MPN are sufficient to mediate MIF-JAB1 interaction, respectively. Finally, endogenous CoIP of MIF-CSN6 complexes from mammalian cells demonstrated that MPN is responsible for MIF-JAB1 binding in vivo, and, as CSN6 does not contain a functional JAMM motif, confirmed that the interaction does not require JAMM.

The Subunit CSN6 of the COP9 Signalosome Is Cleaved during Apoptosis

“…It can associate either as a whole complex or as individual CSN subunits. For instance, p27 kip1 , protein kinase D, migration inhibitory factor, p53, and c-Jun have all been reported to be substrates for the COP9 complex (9,14,33,34). Here we report for the first time that Nod1 specifically binds to some components of the COP9 complex through its CARD domain.…”

Macrophage migration inhibitory factor (MIF) promotes cell survival by activation of the Akt pathway and role for CSN5/JAB1 in the control of autocrine MIF activity

“…MIF promotes cell proliferation and blockade of MIF by antibodies or genetic deletion leads to reduced cellular proliferation and inhibition of tumor growth and angiogenesis (Takahashi et al, 1998;Chesney et al, 1999;Hudson et al, 1999;Shimizu et al, 1999;Mitchell and Bucala, 2000;Bando et al, 2002;Amin et al, 2003;Nishihira et al, 2003). These effects may involve MIF-mediated regulation of CD74-dependent ERK mitogen-activated protein kinase (MAPK) signaling and activation of cytosolic phospholipase A2 (cPLA2) (Mitchell et al, 1999;Leng et al, 2003;Lue et al, 2006), modulation of activities of the tumor-associated protein c-Jun activation domain-binding protein-1 (JAB1) and signaling through the COP9 signalosome (CSN) (Kleemann et al, 2000;Burger-Kentischer et al, 2005). JAB1 is CSN5 of the CSN and functions as coactivator of activator protein-1 (AP-1)-driven gene expression and participates in CSN-mediated activation of SCF-E3 ligase-dependent proteasomal degradation of cell cycle regulators such as p27 or p53 (Chamovitz and Segal, 2001; Bech-Otschir et al, 2002; Wolf et al, 2003).…”

NAC1 Regulates the Recruitment of the Proteasome Complex into Dendritic Spines

“…A yeast 2-hybrid (Y2H) assay was used to screen a GAL4-AD/cDNA library constructed from adult mouse brain using NAC1-GAL4-BD fusion protein as bait. Of the 3 ϫ 10 6 library clones screened, positive clones were found to encode Cul3, a cullin protein that is a critical subunit in cullin-based E3 ubiquitin ligases (Glickman and Ciechanover, 2002), and Mov34, a 19S ATPase regulatory subunit in the 26S proteasome (Mason et al, 1998;Ambroggio et al, 2004;Burger-Kentischer et al, 2005). To verify the interaction between NAC1 with Cul3 and Mov34, isolated prey plasmids were retransformed into AH109 and only Cul3 or Mov34/Gal4BD ϫ lNac1/Gal4AD cotransformed yeast cells proliferated on the selecting medium.…”