bims-lymeca 2022-03-27 papers

Issue of 2022–03–27
eight papers selected by
Harilaos Filippakis, Harvard University

Autophagy. 2022 Mar 22. 1-21

TFEB- and TFE3-dependent autophagy activation supports cancer proliferation in the absence of centrosomes.

Chien-Han Kao, Ting-Yu Su, Wei-Syun Huang, Xin-Ying Lu, Wann-Neng Jane, Chien-Yung Huang, Hung-Hsiang Huang, Won-Jing Wang.

Centrosome amplification is a phenomenon frequently observed in human cancers, so centrosome depletion has been proposed as a therapeutic strategy. However, despite being afflicted with a lack of centrosomes, many cancer cells can still proliferate, implying there are impediments to adopting centrosome depletion as a treatment strategy. Here, we show that TFEB- and TFE3-dependent autophagy activation contributes to acentrosomal cancer proliferation. Our biochemical analyses uncover that both TFEB and TFE3 are novel PLK4 (polo like kinase 4) substrates. Centrosome depletion inactivates PLK4, resulting in TFEB and TFE3 dephosphorylation and subsequent promotion of TFEB and TFE3 nuclear translocation and transcriptional activation of autophagy- and lysosome-related genes. A combination of centrosome depletion and inhibition of the TFEB-TFE3 autophagy-lysosome pathway induced strongly anti-proliferative effects in cancer cells. Thus, our findings point to a new strategy for combating cancer.

Keywords: Anti-cancer therapy; PLK4; autophagy; centrosome; lysosomal biogenesis; transcription factor E3; transcription factor EB

DOI: https://doi.org/10.1080/15548627.2022.2051880

Sci Rep. 2022 Mar 23. 12(1): 5052

Activation of lysosomal mediated cell death in the course of autophagy by mTORC1 inhibitor.

Sameer Ullah Khan, Anup Singh Pathania, Abubakar Wani, Kaneez Fatima, Mubashir Javed Mintoo, Baseerat Hamza, Masroor Ahmad Paddar, Wadhwa Bhumika, Loveleena Kour Anand, Mir Shahid Maqbool, Sameer Ahmad Mir, Jaspreet Kour, Vunnam Venkateswarlu, Dilip Manikrao Mondhe, Sanghapal D Sawant, Fayaz Malik.

Lysosomal biogenesis plays a vital role in cell fate. Under certain conditions, excessive lysosomal biogenesis leads to susceptibility for lysosomal membrane permeabilization resulting in various pathological conditions including cell death. In cancer cells apoptosis machinery becomes dysregulated during the course of treatment, thus allows cancer cells to escape apoptosis. So it is therefore imperative to identify cytotoxic agents that exploit non-apoptotic mechanisms of cell death. Our study showed that pancreatic cancer cells treated with SDS-203 triggered an incomplete autophagic response and a nuclear translocation of transcriptional factor TFEB. This resulted in abundant biosynthesis and accumulation of autophagosomes and lysosomes into the cells leading to their death. It was observed that the silencing of autophagy genes didn't alter the cell fate, whereas siRNA-mediated silencing of TFEB subdued SDS-203 mediated lysosomal biogenesis and associated cell death. Further mouse tumors treated with SDS-203 showed a significant reduction in tumor burden and increased expression of lysosomal markers. Taken together this study demonstrates that SDS-203 treatment triggers non-apoptotic cell death in pancreatic cancer cells through a mechanism of lysosome over accumulation.

DOI: https://doi.org/10.1038/s41598-022-07955-1

Nat Commun. 2022 Mar 21. 13(1): 1540

RUFY3 links Arl8b and JIP4-Dynein complex to regulate lysosome size and positioning.

Gaurav Kumar, Prateek Chawla, Neha Dhiman, Sanya Chadha, Sheetal Sharma, Kanupriya Sethi, Mahak Sharma, Amit Tuli.

The bidirectional movement of lysosomes on microtubule tracks regulates their whole-cell spatial arrangement. Arl8b, a small GTP-binding (G) protein, promotes lysosome anterograde trafficking mediated by kinesin-1. Herein, we report an Arl8b effector, RUFY3, which regulates the retrograde transport of lysosomes. We show that RUFY3 interacts with the JIP4-dynein-dynactin complex and facilitates Arl8b association with the retrograde motor complex. Accordingly, RUFY3 knockdown disrupts the positioning of Arl8b-positive endosomes and reduces Arl8b colocalization with Rab7-marked late endosomal compartments. Moreover, we find that RUFY3 regulates nutrient-dependent lysosome distribution, although autophagosome-lysosome fusion and autophagic cargo degradation are not impaired upon RUFY3 depletion. Interestingly, lysosome size is significantly reduced in RUFY3 depleted cells, which could be rescued by inhibition of the lysosome reformation regulatory factor PIKFYVE. These findings suggest a model in which the perinuclear cloud arrangement of lysosomes regulates both the positioning and size of these proteolytic compartments.

DOI: https://doi.org/10.1038/s41467-022-29077-y

STAR Protoc. 2022 Jun 17. 3(2): 101262

Live cell microscopy of mitochondria-lysosome contact site formation and tethering dynamics.

Tayler B Belton, Eric D Leisten, Jasmine Cisneros, Yvette C Wong.

Mitochondria-lysosome contact sites are critical for maintaining cellular homeostasis by regulating mitochondrial and lysosomal network dynamics and mediating metabolite exchange. Here, we present a protocol to quantitatively analyze the formation and tethering duration of mitochondria-lysosome contact sites by using time-lapse live confocal microscopy of LAMP1 and TOMM20. Although this protocol focuses on mammalian HeLa cells, it can be applied to other cell types for further studies on mitochondria-lysosome contact regulation and function, and elucidation of their role in human disorders. For complete details on the use and execution of this protocol, please refer to Wong et al. (2018) and Wong et al. (2019b).

Keywords: Cell Biology; Cell culture; Microscopy

DOI: https://doi.org/10.1016/j.xpro.2022.101262

Dev Cell. 2022 Mar 15. pii: S1534-5807(22)00126-5. [Epub ahead of print]

The central moTOR of metabolism.

Judith Simcox, Dudley W Lamming.

The protein kinase mechanistic target of rapamycin (mTOR) functions as a central regulator of metabolism, integrating diverse nutritional and hormonal cues to control anabolic processes, organismal physiology, and even aging. This review discusses the current state of knowledge regarding the regulation of mTOR signaling and the metabolic regulation of the four macromolecular building blocks of the cell: carbohydrate, nucleic acid, lipid, and protein by mTOR. We review the role of mTOR in the control of organismal physiology and aging through its action in key tissues and discuss the potential for clinical translation of mTOR inhibition for the treatment and prevention of diseases of aging.

Keywords: amino acids; lipids; mTOR; mTORC1; mTORC2; metabolism; protein; rapamycin

DOI: https://doi.org/10.1016/j.devcel.2022.02.024

Cells. 2022 Mar 08. pii: 921. [Epub ahead of print]11(6):

Current Methods to Unravel the Functional Properties of Lysosomal Ion Channels and Transporters.

Margherita Festa, Velia Minicozzi, Anna Boccaccio, Laura Lagostena, Antonella Gradogna, Tianwen Qi, Alex Costa, Nina Larisch, Shin Hamamoto, Emanuela Pedrazzini, Stefan Milenkovic, Joachim Scholz-Starke, Matteo Ceccarelli, Alessandro Vitale, Petra Dietrich, Nobuyuki Uozumi, Franco Gambale, Armando Carpaneto.

A distinct set of channels and transporters regulates the ion fluxes across the lysosomal membrane. Malfunctioning of these transport proteins and the resulting ionic imbalance is involved in various human diseases, such as lysosomal storage disorders, cancer, as well as metabolic and neurodegenerative diseases. As a consequence, these proteins have stimulated strong interest for their suitability as possible drug targets. A detailed functional characterization of many lysosomal channels and transporters is lacking, mainly due to technical difficulties in applying the standard patch-clamp technique to these small intracellular compartments. In this review, we focus on current methods used to unravel the functional properties of lysosomal ion channels and transporters, stressing their advantages and disadvantages and evaluating their fields of applicability.

Keywords: ion channels; lysosomes; patch-clamp; plant vacuole; transporters

DOI: https://doi.org/10.3390/cells11060921

Sci Rep. 2022 Mar 23. 12(1): 4998

Effects of melatonin and metformin in preventing lysosome-induced autophagy and oxidative stress in rat models of carcinogenesis and the impact of high-fat diet.

Natalia Kurhaluk, Halyna Tkachenko.

Imbalanced glucose tolerance and insulin resistance remain important as high cancer risk factors. Metformin administration to diabetic patients may be associated with a reduced risk of malignancy. The combined effects of the hormone melatonin and metformin in oncology practice have shown positive results. The relevance of our study is to find out the role of specific biomarkers of lysosome destruction and oxidative stress data in carcinogenesis models. The present study was designed to investigate the comparative synergic effect of peroral antidiabetic metformin (MF) and pineal hormone melatonin (MEL) administered alone and in combination in two different rat's models of mammary tumour proliferation in vivo (N-methyl-N-nitrosourea, NMU or 7,12-dimethylbenz[a]anthracene, DMBA). We have studied the processes of lysosomal destruction (alanyl aminopeptidase AAP, leucyl aminopeptidase LAP, acid phosphatase AcP, β-N-acetylglucosaminidase NAG, β-galactosidase β-GD and β-glucuronidase β-GR) caused by evaluated oxidative stress in three types of tissues (liver, heart, and spleen) in female Sprague-Dawley rats fed a high-fat diet (10% of total fat: 2.5% from lard and 7.5% from palm olein). Our results revealed an increase in the activity of the studied lysosomal enzymes and their expression in a tissue-specific manner depending on the type of chemical agent (NMU or DMBA). MANOVA tests in our study confirmed the influence of the three main factors, type of tissue, chemical impact, and chemopreventive agents, and the combinations of these factors on the lysosomal activity induced during the process of cancerogenesis. The development and induction of the carcinogenesis process in the different rat models with the high-fat diet impact were also accompanied by initiation of free-radical oxidation processes, which we studied at the initial (estimated by the level of diene conjugates) and final (TBARS products) stages of this process. The combined effects of MEL and MF for the two models of carcinogenesis at high-fat diet impact for AAP, LAP, and AcP showed a significant synergistic effect when they impact together when compared with the effects of one substance alone (either MEL or MF) in the breast cancer model experiments. Synergistic effects of limiting destructive processes of lysosomal functioning β-GD enzyme activity we obtained in experiments with MEL and MF chemoprevention for both models of carcinogenesis for three tissues. The statistical SS test allowed us to draw the following conclusions on the role of each lysosomal parameter analyzed as an integral model: NAG > AcP > β-GD > β-GR > AAP > LAP.

DOI: https://doi.org/10.1038/s41598-022-08778-w