Inactivation of cell death is a major step in tumor development, and p53, a tumor suppressor frequently mutated in cancer, is a critical mediator of cell death. While a role for p53 in apoptosis is ...well established, direct links to other pathways controlling cell death are unknown. Here we describe
DRAM (
damage-
regulated
autophagy
modulator), a p53 target gene encoding a lysosomal protein that induces macroautophagy, as an effector of p53-mediated death. We show that p53 induces autophagy in a DRAM-dependent manner and, while overexpression of DRAM alone causes minimal cell death, DRAM is essential for p53-mediated apoptosis. Moreover, analysis of
DRAM in primary tumors revealed frequent decreased expression often accompanied by retention of wild-type p53. Collectively therefore, these studies not only report a stress-induced regulator of autophagy but also highlight the relationship of DRAM and autophagy to p53 function and damage-induced programmed cell death.
Sensing nutrient availability is essential for appropriate cellular growth, and mTORC1 is a major regulator of this process. Mechanisms causing mTORC1 activation are, however, complex and diverse. We ...report here an additional important step in the activation of mTORC1, which regulates the efflux of amino acids from lysosomes into the cytoplasm. This process requires DRAM-1, which binds the membrane carrier protein SCAMP3 and the amino acid transporters SLC1A5 and LAT1, directing them to lysosomes and permitting efficient mTORC1 activation. Consequently, we show that loss of DRAM-1 also impacts pathways regulated by mTORC1, including insulin signaling, glycemic balance, and adipocyte differentiation. Interestingly, although DRAM-1 can promote autophagy, this effect on mTORC1 is autophagy independent, and autophagy only becomes important for mTORC1 activation when DRAM-1 is deleted. These findings provide important insights into mTORC1 activation and highlight the importance of DRAM-1 in growth control, metabolic homeostasis, and differentiation.
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•DRAM-1 is required for efficient activation of the nutrient-sensing complex mTORC1•DRAM-1 and SCAMP3 direct newly synthesized amino acid transporters to lysosomes•DRAM-1 drives lysosomal amino acid efflux to promote mTORC1 activation•Loss of DRAM-1 increases insulin sensitivity and enhances adipocyte differentiation
mTORC1 is a nutrient-sensing complex that affects many cellular processes. Beaumatin et al. show that DRAM-1 and SCAMP3 are required for efficient activation of mTORC1. Consequently, they show that loss of DRAM-1 impairs cell growth and amino-acid-induced autophagy repression while promoting insulin sensitivity, glycemic regulation, and adipocyte differentiation.
p53 is a central factor in tumor suppression as exemplified by its frequent loss in human cancer. p53 exerts its tumor suppressive effects in multiple ways, but the ability to invoke the eradication ...of damaged cells by programmed cell death is considered a key factor. The ways in which p53 promotes cell death can involve direct activation or engagement of the cell death machinery, or can be via indirect mechanisms, for example though regulation of ER stress and autophagy. We present here another level of control in p53-mediated tumor suppression by showing that p53 activates the glycosidase, FUCA1, a modulator of N-linked glycosylation. We show that p53 transcriptionally activates FUCA1 and that p53 modulates fucosidase activity via FUCA1 up-regulation. Importantly, we also report that chemotherapeutic drugs induce FUCA1 and fucosidase activity in a p53-dependent manner. In this context, while we found that over-expression of FUCA1 does not induce cell death, RNAi-mediated knockdown of endogenous FUCA1 significantly attenuates p53-dependent, chemotherapy-induced apoptotic death. In summary, these findings add an additional component to p53s tumor suppressive response and highlight another mechanism by which the tumor suppressor controls programmed cell death that could potentially be exploited for cancer therapy.
Glioblastoma (GBM) is the most prevalent malignant primary brain tumour in adults. GBM typically has a poor prognosis, mainly due to a lack of effective treatment options leading to tumour ...persistence or recurrence. We investigated the therapeutic potential of targeting anti-apoptotic BCL-2 proteins in GBM. Levels of anti-apoptotic BCL-xL and MCL-1 were consistently increased in GBM compared with non-malignant cells and tissue. Moreover, we found that relative to their differentiated counterparts, patient-derived GBM stem-like cells also displayed higher expression of anti-apoptotic BCL-2 family members. High anti-apoptotic BCL-xL and MCL-1 expression correlated with heightened susceptibility of GBM to BCL-2 family protein-targeting BH3-mimetics. This is indicative of increased apoptotic priming. Indeed, GBM displayed an obligate requirement for MCL-1 expression in both tumour development and maintenance. Investigating this apoptotic sensitivity, we found that sequential inhibition of BCL-xL and MCL-1 led to robust anti-tumour responses in vivo, in the absence of overt toxicity. These data demonstrate that BCL-xL and MCL-1 pro-survival function is a fundamental prerequisite for GBM survival that can be therapeutically exploited by BH3-mimetics.
Glycan degradation promotes macroautophagy Baudot, Alice D; Wang, Victoria M-Y; Leach, Josh D ...
Proceedings of the National Academy of Sciences - PNAS,
06/2022, Letnik:
119, Številka:
26
Journal Article
Recenzirano
Odprti dostop
Macroautophagy promotes cellular homeostasis by delivering cytoplasmic constituents to lysosomes for degradation Mizushima,
20, 521-527 (2018). However, while most studies have focused on the ...mechanisms of protein degradation during this process, we report here that macroautophagy also depends on glycan degradation via the glycosidase, α-l-fucosidase 1 (FUCA1), which removes fucose from glycans. We show that cells lacking FUCA1 accumulate lysosomal glycans, which is associated with impaired autophagic flux. Moreover, in a mouse model of fucosidosis-a disease characterized by inactivating mutations in
Stepien
,
11, E1383 (2020)-glycan and autophagosome/autolysosome accumulation accompanies tissue destruction. Mechanistically, using lectin capture and mass spectrometry, we identified several lysosomal enzymes with altered fucosylation in FUCA1-null cells. Moreover, we show that the activity of some of these enzymes in the absence of FUCA1 can no longer be induced upon autophagy stimulation, causing retardation of autophagic flux, which involves impaired autophagosome-lysosome fusion. These findings therefore show that dysregulated glycan degradation leads to defective autophagy, which is likely a contributing factor in the etiology of fucosidosis.
What's known on the subject? and What does the study add?
Androgen‐ablation therapy (AAT) and chemotherapy are commonly used to treat incurable prostate cancer. To improve outcome, there is major ...on‐going research to develop more effective treatments with less toxicity. Autophagy has been suggested from previous studies to play a potential role in cell survival and may be associated with resistance to chemotherapy.
Autophagy is known to be upregulated by nutrient starvation or AAT in prostate cancer. However, its functional impact is not fully known. The present study describes the potential synergism between the blockade of autophagy and AAT alone or AAT combined with taxane chemotherapy. Hence, future combined treatment options are warranted to further investigate the clinical impact of autophagy suppression as a treatment strategy.
Objective
To study the cellular effects of the anti‐androgen bicalutamide on autophagy and its potential impact on response to androgen‐ablation therapy (AAT) alone or combined with docetaxel chemotherapy in human prostate cancer LNCaP cells.
Materials and Methods
LNCaP cells were treated with bicalutamide ± docetaxel, and cellular effects were assayed: lipidated LC3 (a microtubule‐associated protein) for autophagy and its trafficking to fuse with lysosome; flow cytometry using propidium iodide or caspase 3 for cell death; and sulforhodamine B assay for cell growth.
Results
Bicalutamide treatment enhanced autophagy in LNCaP cells with increased level of autophagosome coupled with an altered cellular morphology reminiscent of neuroendocrine differentiation.
Consistent with the literature on the interaction between androgen receptor activation and taxane chemotherapy, bicalutamide diminished docetaxel mediated cytotoxicity.
Significantly, pharmacological inhibition of autophagy with 3‐methyladenine significantly enhanced the efficacy cell kill mediated by AAT ± docetaxel.
Conclusion
Autophagy associated with bicalutamide treatment in LNCaP cells may have a pro‐survival effect and strategy to modulate autophagy may have a potential therapeutic value.
Significance It is known that autophagy plays a major role in cellular homeostasis and impaired autophagy has been implicated in various forms of human disease. Here we investigate one way in which ...autophagy is critically connected to cellular integrity by showing that autophagy loss diminishes DNA repair by the error-free process, homologous recombination. As a result, this causes reliance on the error-prone process of nonhomologous end joining for repair of DNA double-strand breaks, providing one explanation of why autophagy-deficient cells accumulate genomic damage. These findings therefore have important implications for diseases in which autophagy is impaired and for therapeutic strategies designed to inhibit autophagy, particularly if autophagy inhibition is undertaken in combination with agents that cause genomic damage.
(Macro)autophagy delivers cellular constituents to lysosomes for degradation. Although a cytoplasmic process, autophagy-deficient cells accumulate genomic damage, but an explanation for this effect is currently unclear. We report here that inhibition of autophagy causes elevated proteasomal activity leading to enhanced degradation of checkpoint kinase 1 (Chk1), a pivotal factor for the error-free DNA repair process, homologous recombination (HR). We show that loss of autophagy critically impairs HR and that autophagy-deficient cells accrue micronuclei and sub-G1 DNA, indicators of diminished genomic integrity. Moreover, due to impaired HR, autophagy-deficient cells are hyperdependent on nonhomologous end joining (NHEJ) for repair of DNA double-strand breaks. Consequently, inhibition of NHEJ following DNA damage in the absence of autophagy results in persistence of genomic lesions and rapid cell death. Because autophagy deficiency occurs in several diseases, these findings constitute an important link between autophagy and DNA repair and highlight a synthetic lethal strategy to kill autophagy-deficient cells.
Autophagy is a membrane-trafficking process that directs degradation of cytoplasmic material in lysosomes. The process promotes cellular fidelity, and while the core machinery of autophagy is known, ...the mechanisms that promote and sustain autophagy are less well defined. Here we report that the epigenetic reader BRD4 and the methyltransferase G9a repress a TFEB/TFE3/MITF-independent transcriptional program that promotes autophagy and lysosome biogenesis. We show that BRD4 knockdown induces autophagy in vitro and in vivo in response to some, but not all, situations. In the case of starvation, a signaling cascade involving AMPK and histone deacetylase SIRT1 displaces chromatin-bound BRD4, instigating autophagy gene activation and cell survival. Importantly, this program is directed independently and also reciprocally to the growth-promoting properties of BRD4 and is potently repressed by BRD4-NUT, a driver of NUT midline carcinoma. These findings therefore identify a distinct and selective mechanism of autophagy regulation.
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•BRD4 represses a program of autophagy and lysosome genes independently of MiT/TFE•BRD4 de-repression promotes certain types of autophagy, but not others•Nutrient deprivation de-represses BRD4 via AMPK signaling to promote cell survival•Oncoprotein BRD4-NUT is a potent repressor of autophagy and lysosome function
Sakamaki et al. show that BRD4 represses autophagy and lysosome gene expression. This repression is alleviated during nutrient deprivation through AMPK-SIRT1 signaling, allowing autophagy activation. BRD4 inhibition enhances autophagic flux and lysosomal function and promotes the degradation of protein aggregates.
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