Abstract
The primary cilium is a microtubule-based sensory organelle. The molecular mechanism that regulates ciliary dynamics remains elusive. Here, we report an unexpected finding that MLN4924, a small molecule inhibitor of NEDD8-activating enzyme (NAE), blocks primary ciliary formation by inhibiting synthesis/assembly and promoting disassembly. This is mainly mediated by MLN4924-induced phosphorylation of AKT1 at Ser473 under serum-starved, ciliary-promoting conditions. Indeed, pharmaceutical inhibition (by MK2206) or genetic depletion (via siRNA) of AKT1 rescues MLN4924 effect, indicating its causal role. Interestingly, pAKT1-Ser473 activity regulates both ciliary synthesis/assembly and disassembly in a MLN4924 dependent manner, whereas pAKT-Thr308 determines the ciliary length in MLN4924-independent but VHL-dependent manner. Finally, MLN4924 inhibits mouse hair regrowth, a process requires ciliogenesis. Collectively, our study demonstrates an unexpected role of a neddylation inhibitor in regulation of ciliogenesis via AKT1, and provides a proof-of-concept for potential utility of MLN4924 in the treatment of human diseases associated with abnormal ciliogenesis.
Highlights
The primary cilium, a microtubule-based organelle, that projects from the apical surface of most eukaryotic cells, has emerged as a key organelle involved in many physiological and developmental processes (Goetz and Anderson, 2010)
Previous studies have shown that ubiquitylation regulates cilia assembly and disassembly (Hossain and Tsang, 2018), and several ciliary-associated proteins such as CP110 (D’Angiolella et al, 2010) Aurora A (Kwon et al, 2012) are substrates of F-box containing complex (Cullin)-RING ligases (CRLs), suggesting that CRL may regulate ciliogenesis
We recently showed that MLN4924 could activate AKT1 upon EGF addition to serum-starved cells via inducing epidermal growth factor receptor (EGFR) dimerization (Zhou et al, 2016), and two studies reported that EGFR inhibitor could maintain ciliogenesis, and prevented smoke mediated ciliary loss in airways (Khan et al, 2016; Valencia-Gattas et al, 2016), suggesting that the EGFR/AKT1 signal may mediate MLN4924 effect on ciliogenesis
Summary
The primary cilium (immotile cilium), a microtubule-based organelle, that projects from the apical surface of most eukaryotic cells, has emerged as a key organelle involved in many physiological and developmental processes (Goetz and Anderson, 2010). Potential involvement of abnormal primary cilia in tumorigenesis was implicated, as evidenced by reduced or loss of primary cilia in human cancer cell lines derived from breast carcinoma, melanoma, clear cell renal cell carcinoma (Liu et al, 2018). Neddylation modification is a process of covalent attachment of NEDD8, a ubiquitin-like peptide, to a lysine residue of a given protein substrate, catalyzed by NEDD8activating enzyme (NAE, E1), NEDD8-conjugating enzyme (E2), and NEDD8 ligase (E3) (Zhao et al, 2014).
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