Phosphatidic acid signaling to mTOR: Signals for the survival of human cancer cells

https://doi.org/10.1016/j.bbalip.2009.02.009Get rights and content

Abstract

During the past decade elevated phospholipase D (PLD) activity has been reported in virtually all cancers where it has been examined. PLD catalyzes the hydrolysis of phosphatidylcholine to generate the lipid second messenger phosphatidic acid (PA). While many targets of PA signaling have been identified, the most critical target of PA in cancer cells is likely to be mTOR — the mammalian target of rapamycin. mTOR has been widely implicated in signals that suppress apoptotic programs in cancer cells — frequently referred to as survival signals. mTOR exists as two multi-component complexes known as mTORC1 and mTORC2. Recent data has revealed that PA is required for the stability of both mTORC1 and mTORC2 complexes — and therefore also required for the kinase activity of both mTORC1 and mTORC2. PA interacts with mTOR in a manner that is competitive with rapamycin, and as a consequence, elevated PLD activity confers rapamycin resistance — a point that has been largely overlooked in clinical trials involving rapamycin-based strategies. The earliest genetic changes occurring in an emerging tumor are generally ones that suppress default apoptotic programs that likely represent the first line of defense of cancer. Targeting survival signals in human cancers represents a rational anti-cancer therapeutic strategy. Therefore, understanding the signals that regulate PA levels and how PA impacts upon mTOR could be important for developing strategies to de-repress the survival signals that suppress apoptosis. This review summarizes the role of PA in regulating the mTOR-mediated signals that promote cancer cell survival.

Introduction

During the past fifteen years many studies have shown that in response to mitogenic signals and activated oncoproteins there is an increase in phospholipase D (PLD) activity. PLD activity is elevated in response to several growth factors including epidermal growth factor (EGF) [1], platelet-derived growth factor [2], fibroblast growth factor [3], [4], insulin [5], insulin-like growth factor 1 [6], and vascular endothelial growth factor [7]. In addition, PLD activity is elevated in response to the oncoproteins v-Src [8], v-Ras [9], [10], v-Fps [11], and v-Raf [12]. The elevated PLD activity in cells transformed by the Ras oncoprotein is required for cell transformation [13]. In addition, elevated expression of either c-Src or the EGF receptor, in combination with elevated expression of PLD, transforms rat fibroblasts [14], [15], [16]. PLD has also been reported to induce anchorage-independent growth and enhance cell cycle progression of mouse fibroblasts [17]. These studies reveal elevated PLD activity is correlated with mitogenic and oncogenic signals.

Elevated PLD expression has also been shown to prevent cell cycle arrest and apoptosis. High intensity Raf signaling induces cell senescence [18], [19] or, if cells are deprived of serum, apoptosis [20]. Elevated expression of PLD suppressed the apoptosis induced by high intensity Raf signals [20]. Similarly, rat fibroblasts overexpressing c-Src undergo apoptosis in response to growth factor deprivation, and elevated PLD expression suppressed this apoptosis [21]. These early studies in rodent fibroblasts indicate that in addition to enhancing cell proliferation, PLD is required for the “survival signals” that suppress default apoptotic programs.

The studies implicating PLD in mitogenic signaling and the suppression of apoptosis suggest that PLD activity might be a factor in human cancer where mitogenic signals are constitutively active and suppression of apoptotic programs is critical. Consistent with this hypothesis, elevated PLD activity and expression has been reported in a variety of human cancer tissues including breast, gastric, kidney, and colon [22], [23], [24], [25]. In addition, elevated PLD activity has been reported in several human cancer cell lines including those derived from breast, lung, bladder, pancreatic, and kidney cancers [26], [27], [28], [29], [30]. Importantly, the elevated PLD activity in these cells was shown to be critical for suppressing apoptosis in these cell lines. Thus, elevated PLD activity in human cancers is likely a critical aspect of tumorigenesis that promotes cell proliferation and suppresses the default apoptotic programs that prevent cancer.

There are two mammalian PLD isoforms – PLD1 and PLD2 – the distinct functions of which are poorly understood [31]. Both catalyze the hydrolysis of phosphatidylcholine to phosphatidic acid (PA) and choline. PA is a central node for lipid signaling and can be generated from lysophosphatidic acid (LPA) and diacylglycerol (DG) as well from phosphatidylcholine (Fig. 1A) — although it is likely that the most relevant source in cancer cells is via the PLD-mediated hydrolysis of phosphatidylcholine [31]. PA is also metabolically converted to the lipid second messengers LPA and DG (Fig. 1A). However, while DG and LPA have important second messenger function, there are several targets of PA have been identified (Fig. 1B) and it is believed that the most significant effects of elevated PLD activity are mediated by targets of PA. Significantly, these include Raf [32] and the mammalian target of rapamycin (mTOR) [33] — both of which are commonly dysregulated in human cancers. mTOR, like PLD, has been implicated in cancer cell survival signals [34], [35], [36]. While the role of PA in regulating mTOR has been controversial [37], recent reports have strongly implicated PLD and its metabolite PA in the regulation of both mTOR complexes — mTORC1 and mTORC2 [38], [39], [40]. This review summarizes the role of PLD in the regulation of mTOR and the potential of combining strategies for targeting PLD and mTOR signals in human cancer.

Section snippets

PA competes with rapamycin for binding to mTOR

The first report directly linking PLD activity with mTOR was the discovery that PA is required for mTOR activity [33]. PA was shown to bind the FKBP12-rapamycin binding (FRB) domain of mTOR in a manner that is competitive with rapamycin-FKBP12 [33]. Consistent with a competition between PA and rapamycin-FKBP12 for mTOR, it was subsequently demonstrated that elevated levels of PLD activity conferred rapamycin resistance in human breast cancer cell lines [42]. The recently reported NMR structure

PA is necessary, but not sufficient to activate mTOR

While recent studies clearly demonstrate a PA requirement for the activation of both mTORC1 and mTORC2, there is also evidence that PA is not sufficient to activate either mTORC1 or mTORC2. Chen and colleagues demonstrated that exogenously provided PA stimulated the activation of the mTORC1 as indicated by increased phosphorylation of the mTORC1 substrates S6 kinase eukaryotic initiation factor 4E binding protein-1 in HEK293 cells, however the effect was dependent on the presence of amino acids

Regulation of PLD1 and PLD2

There are two mammalian PLD isoforms – PLD1 and PLD2 – that can be distinguished by different mechanisms of regulation and sub-cellular distribution [31]. PLD1 has a predominantly peri-nuclear localization and is regulated by members of the Ras family of GTPases including ARF [62], Rho [63], and Ral [64]. Importantly, it was recently reported that the GTPase Rheb activates PLD1 [38]. PLD2 is largely restricted to lipid raft fractions on the plasma membrane and its mode of regulation is not well

Targeting PLD-mTOR survival signals

Targeting mTOR in anti-cancer therapies has attracted much attention in recent years largely due to a link between mTOR and survival signals in human cancer cells [34], [35], [36]. Much has been written about the potential of targeting mTOR because, in principle, the suppression of survival signals in cancer cells should result in apoptotic cell death and tumor regression. While the principle of targeting mTOR-mediated survival signals in human cancer offers an attractive therapeutic option,

Conclusions

Elevated PLD activity has been observed in a large number of human cancers and has been shown to suppress apoptosis [74]. PLD activity is commonly elevated in cancer cell lines in response to the stress of serum withdrawal [27]. It has now become apparent that a key target of PLD survival signals is mTOR. mTOR has been implicated as a key regulator of stress responses by shutting down under conditions of poor nutrition or hypoxia [36], [61]. In order for a cancer cell to survive and

Acknowledgements

Paige Yellen is acknowledged for thoughtful comments on the manuscript. The author of this work was supported by grants from the National Cancer Institute (CA46677) and a SCORE grant from the National Institutes of Health (GM60654). Research Centers in Minority Institutions (RCMI) award RR-03037 from the National Center for Research Resources of the National Institutes of Health, which supports infrastructure and instrumentation in the Biological Sciences Department at Hunter College, is also

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      PLD catalyzes the hydrolysis of membrane phospholipids, mainly phosphatidylcholine, which yields to the production of phosphatidic acid (PA) and choline. PA is a potent regulator of mTOR [25], actin cytoskeleton organization [26,27], and it also interacts with a vast array of proteins [28,29]. Above all, PA is negatively charged and has a “cone” like shape which is thought to be helping with vesicles formation and stability [30], facilitating exosome biogenesis.

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