RESUMO
N-(3-Oxododecanoyl)-l-homoserine lactone (C12) is produced by Pseudomonas aeruginosa to function as a quorum-sensing molecule for bacteria-bacteria communication. C12 is also known to influence many aspects of human host cell physiology, including induction of cell death. However, the signalling pathway(s) leading to C12-triggered cell death is (are) still not completely known. To clarify cell death signalling induced by C12, we examined mouse embryonic fibroblasts deficient in "initiator" caspases or "effector" caspases. Our data indicate that C12 selectively induces the mitochondria-dependent intrinsic apoptotic pathway by quickly triggering mitochondrial outer membrane permeabilisation. Importantly, the activities of C12 to permeabilise mitochondria are independent of activation of both "initiator" and "effector" caspases. Furthermore, C12 directly induces mitochondrial outer membrane permeabilisation in vitro. Overall, our study suggests a mitochondrial apoptotic signalling pathway triggered by C12, in which C12 or its metabolite(s) acts on mitochondria to permeabilise mitochondria, leading to activation of apoptosis.
Assuntos
4-Butirolactona/análogos & derivados , Apoptose/fisiologia , Homosserina/análogos & derivados , Membranas Mitocondriais/metabolismo , Percepção de Quorum/fisiologia , 4-Butirolactona/metabolismo , Animais , Caspase 3/genética , Caspase 7/genética , Caspase 9/genética , Caspase 9/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Fibroblastos/metabolismo , Células HCT116 , Homosserina/metabolismo , Humanos , Camundongos , Camundongos Knockout , Interações Microbianas/fisiologia , Mitocôndrias/metabolismo , Pseudomonas aeruginosa/metabolismoRESUMO
Pseudomonas aeruginosa use quorum-sensing molecules, including N-(3-oxododecanoyl)-homoserine lactone (C12), for intercellular communication. C12 activated apoptosis in mouse embryo fibroblasts (MEF) from both wild type (WT) and Bax/Bak double knock-out mice (WT MEF and DKO MEF that were responsive to C12, DKOR MEF): nuclei fragmented; mitochondrial membrane potential (Δψmito) depolarized; Ca(2+) was released from the endoplasmic reticulum (ER), increasing cytosolic [Ca(2+)] (Cacyto); and caspase 3/7 was activated. DKOR MEF had been isolated from a nonclonal pool of DKO MEF that were non-responsive to C12 (DKONR MEF). RNAseq analysis, quantitative PCR, and Western blots showed that WT and DKOR MEF both expressed genes associated with cancer, including paraoxonase 2 (PON2), whereas DKONR MEF expressed little PON2. Adenovirus-mediated expression of human PON2 in DKONR MEF rendered them responsive to C12: Δψmito depolarized, Cacyto increased, and caspase 3/7 activated. Human embryonic kidney 293T (HEK293T) cells expressed low levels of endogenous PON2, and these cells were also less responsive to C12. Overexpression of PON2, but not PON2-H114Q (no lactonase activity) in HEK293T cells caused them to become sensitive to C12. Because [C12] may reach high levels in biofilms in lungs of cystic fibrosis (CF) patients, PON2 lactonase activity may control Δψmito, Ca(2+) release from the ER, and apoptosis in CF airway epithelia. Coupled with previous data, these results also indicate that PON2 uses its lactonase activity to prevent Bax- and Bak-dependent apoptosis in response to common proapoptotic drugs like doxorubicin and staurosporine, but activates Bax- and Bak-independent apoptosis in response to C12.
Assuntos
4-Butirolactona/análogos & derivados , Apoptose , Arildialquilfosfatase/metabolismo , Homosserina/análogos & derivados , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/fisiologia , Percepção de Quorum , 4-Butirolactona/metabolismo , Animais , Células Cultivadas , Células HEK293 , Homosserina/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Camundongos , Infecções por Pseudomonas/microbiologia , Infecções por Pseudomonas/patologiaRESUMO
Paraoxonase 2 (PON2) is a multifunctional intracellular enzyme that has received growing attention for its ability to modulate various aspects of normal and malignant cellular physiology. Recent research has revealed that PON2 is upregulated in tissues from patients with various types of solid tumors and hematologic cancers, likely due to its ability to suppress oxidative stress and evade apoptosis. However, the effects of PON2 on pulmonary oncogenesis are unknown. Here, we conducted studies to investigate how PON2 influences lung cancer cell proliferation in vitro and lung tumorigenesis in vivo using a variety of cellular and animal models. It was found that PON2 expression deficiency hampered the proliferation of cultured lung cancer cells with concomitant cell cycle arrest at the G1 phase. In addition, the loss of endogenous PON2 expression impaired key aspects of oxidative metabolism in lung adenocarcinoma cells. Moreover, we investigated how the interplay between PON2 expression in lung tumors and host mice influences lung tumor initiation and progression. PON2 status in both transplanted tumor cells and mice failed to influence the development of subcutaneously grafted Lewis lung carcinoma (LLC) tumors, orthotopically implanted LLC tumors, and oncogenic Kras-driven primary lung adenocarcinoma tumors. Importantly, the frequencies of tumor-infiltrating myeloid subsets that include myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages were not impacted by PON2 expression in LLC tumor-bearing mice. Overall, our studies indicate that PON2 plays a limited role in murine lung tumorigenesis.
Assuntos
Adenocarcinoma de Pulmão , Arildialquilfosfatase , Neoplasias Pulmonares , Animais , Camundongos , Adenocarcinoma de Pulmão/genética , Arildialquilfosfatase/genética , Arildialquilfosfatase/metabolismo , Carcinogênese/genética , Transformação Celular Neoplásica/genética , Pulmão/metabolismo , Neoplasias Pulmonares/genéticaRESUMO
Lung cancer is the leading cause of cancer death in the United States and worldwide, and a major source of cancer health disparities. Lung cancer cell lines provide key in vitro models for molecular studies of lung cancer development and progression, and for pre-clinical drug testing. To ensure health equity, it is imperative that cell lines representing different lung cancer histological types, carrying different cancer driver genes, and representing different genders, races, and ethnicities should be available. This is particularly relevant for cell lines from Black men, who experience the highest lung cancer mortality in the United States. Here, we undertook a review of the available lung cancer cell lines and their racial and ethnic origin. We noted a marked imbalance in the availability of cell lines from different races and ethnicities. Cell lines from Black patients were strongly underrepresented, and we identified no cell lines from Hispanic/Latin(x) (H/L), American Indian/American Native (AI/AN), or Native Hawaiian or other Pacific Islander (NHOPI) patients. The majority of cell lines were derived from White and Asian patients. Also missing are cell lines representing the cells-of-origin of the major lung cancer histological types, which can be used to model lung cancer development and to study the effects of environmental exposures on lung tissues. To our knowledge, the few available immortalized alveolar epithelial cell lines are all derived from White subjects, and the race and ethnicity of a handful of cell lines derived from bronchial epithelial cells are unknown. The lack of an appropriately diverse collection of lung cancer cell lines and lung cancer cell-of-origin lines severely limits racially and ethnically inclusive lung cancer research. It impedes the ability to develop inclusive models, screen comprehensively for effective compounds, pre-clinically test new drugs, and optimize precision medicine. It thereby hinders the development of therapies that can increase the survival of minority and underserved patients. The noted lack of cell lines from underrepresented groups should constitute a call to action to establish additional cell lines and ensure adequate representation of all population groups in this critical pre-clinical research resource.
RESUMO
Pseudomonas aeruginosa produces N-(3-oxododecanoyl)-homoserine lactone (C12) as a quorum-sensing molecule for bacterial communication. C12 has also been reported to induce apoptosis in various types of tumor cells. However, the detailed molecular mechanism of C12-triggerred tumor cell apoptosis is still unclear. In addition, it is completely unknown whether C12 possesses any potential therapeutic effects in vivo. Our data indicate that, unlike most apoptotic inducers, C12 evokes a novel form of apoptosis in tumor cells through inducing mitochondrial membrane permeabilization independent of both pro- and anti-apoptotic Bcl-2 proteins. Importantly, C12 inhibits tumor growth in animals regardless of either pro- or anti-apoptotic Bcl-2 proteins. Furthermore, opposite to conventional chemotherapeutics, C12 requires paraoxonase 2 (PON2) to exert its cytotoxicity on tumor cells in vitro and its inhibitory effects on tumor growth in vivo. Overall, our results demonstrate that C12 inhibits tumor growth independent of both pro- and anti-apoptotic Bcl-2 proteins, and through inducing unique apoptotic signaling mediated by PON2 in tumor cells.