Bnip3 Mediates the Hypoxia-induced Inhibition on Mammalian Target of Rapamycin by Interacting with Rheb

Abstract

The mammalian target of rapamycin (mTOR) is a central controller of cell growth, and it regulates translation, cell size, cell viability, and cell morphology. mTOR integrates a wide range of extracellular and intracellular signals, including growth factors, nutrients, energy levels, and stress conditions. Rheb, a Ras-related small GTPase, is a key upstream activator of mTOR. In this study, we found that Bnip3, a hypoxia-inducible Bcl-2 homology 3 domain-containing protein, directly binds Rheb and inhibits the mTOR pathway. Bnip3 decreases Rheb GTP levels in a manner depending on the binding to Rheb and the presence of the N-terminal domain. Both knockdown and overexpression experiments show that Bnip3 plays an important role in mTOR inactivation in response to hypoxia. Moreover, Bnip3 inhibits cell growth in vivo by suppressing the mTOR pathway. These observations demonstrate that Bnip3 mediates the inhibition of the mTOR pathway in response to hypoxia.Target of rapamycin (TOR), 2 is an evolutionarily conserved serine/threonine kinase that plays a central role in cell growth (1-3). TOR regulates many processes, including protein translation, ribosome biogenesis, autophagy, and metabolism. TOR functions to integrate a wide range of extracellular and intracellular signals to produce a concerted cellular response, such as to stimulate cell growth. For example, mammalian target of rapamycin (mTOR) is activated by growth factor and nutrient availability. In contrast, mTOR is inhibited by cellular energy starvation and various stress conditions, including osmotic stress and hypoxia. These observations established a fundamental importance of mTOR in signal integration in regulation of cell growth.Recent studies have demonstrated that TOR exists in two functionally distinct protein complexes, termed TOR complex 1 (TORC1) and TOR complex 2 (TORC2) (3, 4). The two TOR complexes were initially identified in yeast and subsequently were also characterized in Drosophila and mammalian cells. TORC1 contains mTOR, mLST8, PRAS40, and Raptor, whereas TORC2 contains mTOR, mLST8, Rictor, and Sin1 (5-12). TORC1 is responsible for phosphorylation of Thr 389 of S6K1 (ribosomal protein S6 kinase) and 4EBP1 (eukaryote initiation factor 4E-binding protein), two important regulators in protein synthesis (1). In contrast, TORC2 has different substrates and is responsible for phosphorylation of the hydrophobic sites in both AKT and PKC (9, 13). Interestingly, TORC1 but not TORC2 is inhibited by rapamycin (6,8,9). These observations clearly demonstrate that the two TOR complexes have different physiological functions in vivo.Much progress has been made regarding the mechanisms of TORC1 regulation. Tuberous sclerosis complex (TSC) is a genetic disease characterized by benign hamartomas in various tissues (14). Mutations in either the TSC1 or TSC2 tumor suppressor gene are responsible for TSC development. TORC1 is highly activated in TSC tumors or cells with mutation of either TSC1 or TSC2. Both genetic and biochemical data have demonstrated th...

BNIP3 protects HepG2 cells against etoposide-induced cell death under hypoxia by an autophagy-independent pathway

“…The involvement of BNIP3 in other steps of autophagy has also been described. It seems that BNIP3 can induce autophagy by interacting with RheB, resulting in mTOR inhibition [17]. Moreover, the involvement of BNIP3 in later stages of autophagy, such as autophagosome-lysosome fusion, has also been described [16].…”

“…However, the precise mechanisms involved are still poorly understood. It seems that BNIP3 could be involved in autophagosome-lysosome fusion or in earlier stages such as Beclin-1 activation and mTOR inhibition [10,16,17].…”

Understanding complexity in the HIF signaling pathway using systems biology and mathematical modeling

“…The TSC1/2 possesses GTPase activating function that renders the mTOR activator RHEB inactive [76]. BNIP3, another known HIF-1 target, has also been reported to facilitate the accumulation of the GDP-bound form of RHEB and the consequent down-regulation of mTOR in hypoxia [77]. In addition, the HIF-1-inducible miR-155 also targets elements of the mTOR pathway including RHEB, the mTORC2 member RICOR and the mTOR effector ribosomal protein S6KB2 [78].…”

The role of Bcl-2 family member BNIP3 in cell death and disease: NIPping at the heels of cell death

“…37 Another theory is that BNIP3 blocks autophagy by inhibiting mammalian target of rapamycin (mTOR) signaling through binding to Rheb (an activator of mTOR). 38 The molecular mechanism of the involvement of BNIP3 in the induction of autophagy remains to be defined.…”