bims-lymeca 2022-03-20 papers

bims-lymeca

Biomed News

on Lysosome metabolism in cancer

Issue of 2022–03–20
seven papers selected by
Harilaos Filippakis, Harvard University

Material properties of phase-separated TFEB condensates regulate the autophagy-lysosome pathway.
Regulation of basal autophagy by calmodulin availability.
Rheb regulates nuclear mTORC1 activity independent of farnesylation.
Autophagy flux inhibition mediated by lysosomal dysfunction participates in the cadmium exposure-induced cardiotoxicity in swine.
Lysosomal inhibition sensitizes TMEM16A-expressing cancer cells to chemotherapy.
Follicular lymphoma-associated mutations in the V-ATPase chaperone VMA21 activate autophagy creating a targetable dependency.
Lysosomal Ca2+-mediated TFEB activation modulates mitophagy and functional adaptation of pancreatic β-cells to metabolic stress.

J Cell Biol. 2022 May 02. pii: e202112024. [Epub ahead of print]221(5):

Material properties of phase-separated TFEB condensates regulate the autophagy-lysosome pathway.

Zheng Wang, Di Chen, Dongshi Guan, Xiaobo Liang, Jianfeng Xue, Hongyu Zhao, Guangtao Song, Jizhong Lou, Yan He, Hong Zhang.

Very little is known about how the material properties of protein condensates assembled via liquid-liquid phase separation (LLPS) are maintained and affect physiological functions. Here we show that liquid-like condensates of the transcription factor TFEB exhibit low fusion propensity in vitro and in living cells. We directly measured the attraction force between droplets, and we characterized the interfacial tension, viscosity, and elasticity of TFEB condensates. TFEB condensates contain rigid interfacial boundaries that govern their interaction behaviors. Several small molecules, including Ro-3306, modify the material properties of TFEB condensates, increasing their size and fusion propensity. These compounds promote lysosomal biogenesis and function in a TFEB-dependent manner without changing its cytoplasmic-nuclear translocation. Ro-3306 promotes autophagy activity, facilitating degradation of toxic protein aggregates. Our study helps explain how protein condensates are maintained as physically separate entities and reveals that the material properties of TFEB condensates can be harnessed to modulate TFEB activity.

DOI: https://doi.org/10.1083/jcb.202112024
FEBS J. 2022 Mar 14.

Regulation of basal autophagy by calmodulin availability.

Jennifer Giles, Vanessa Lopez, Elizabeth McConnaha, Matthew Hayden, Caleb Kragenbring, David Carli, Eric Wauson, Quang-Kim Tran.

  Macroautophagy (hereafter autophagy) is a process that degrades cellular components to maintain homeostasis. The Ca2+ sensor calmodulin (CaM) regulates numerous cell functions but is a limiting factor due to its insufficient availability for all target proteins. However, evidence that CaM availability regulates basal autophagy is lacking. Here, we have tested this hypothesis. CaM antagonists W-7, trifluoperazine and CGS9343b cause autophagosome accumulation and inhibit basal autophagic flux in the same manner as does chloroquine. These reagents promote the activity of AMP-activated protein kinase (AMPK) but not that of the mechanistic target of rapamycin (mTOR). Competitive binding assays using CaM sensors with different Ca2+ dependencies showed that chloroquine directly binds CaM in a Ca2+ -dependent fashion. The CaM antagonists have disparate effects on cytoplasmic Ca2+ , triggering from none to robust signals, indicating that their consistent inhibition of autophagy is due to inhibition of CaM and not Ca2+ . Chelating intracellular Ca2+ reduces the effect of the CaM antagonists to accumulate LC3-II, indicating that they do so by inhibiting CaM-dependent activities at basal Ca2+ level. The CaM antagonists cause lysosomal alkalinisation. Consistently, buffering CaM with a high-affinity CaM-binding protein that binds CaM at resting Ca2+ level increases lysosomal pH. Enhanced CaM buffering using a chimeric protein that contains two high-affinity CaM-binding sites that can collectively bind CaM at a large range of Ca2+ further increases lysosomal pH and increases LC3-II accumulation and AMPK activity, but not that of mTOR. These data demonstrate that CaM availability is required for basal autophagy.

Keywords:  autophagy; calmodulin; calmodulin antagonists; chloroquine; lysosomal acidification

DOI:  https://doi.org/10.1111/febs.16432
Cell Chem Biol. 2022 Mar 04. pii: S2451-9456(22)00087-3. [Epub ahead of print]

Rheb regulates nuclear mTORC1 activity independent of farnesylation.

Yanghao Zhong, Xin Zhou, Kun-Liang Guan, Jin Zhang.

  The small GTPase Ras homolog enriched in brain (Rheb) plays a critical role in activating the mechanistic target of rapamycin complex 1 (mTORC1), a signaling hub that regulates various cellular functions. We recently observed nuclear mTORC1 activity, raising an intriguing question as to how Rheb, which is known to be farnesylated and localized to intracellular membranes, regulates nuclear mTORC1. In this study, we found that active Rheb is present in the nucleus and required for nuclear mTORC1 activity. We showed that inhibition of farnesyltransferase reduced cytosolic, but not nuclear, mTORC1 activity. Furthermore, a farnesylation-deficient Rheb mutant, with preferential nuclear localization and specific lysosome tethering, enables nuclear and cytosolic mTORC1 activities, respectively. These data suggest that non-farnesylated Rheb is capable of interacting with and activating mTORC1, providing mechanistic insights into the molecular functioning of Rheb as well as regulation of the recently observed, active pool of nuclear mTORC1.

Keywords:  Compartmentation; PTM; TSC; biosensor; lipid modification; mTOR; small GTPase

DOI:  https://doi.org/10.1016/j.chembiol.2022.02.006
Biofactors. 2022 Mar 14.

Autophagy flux inhibition mediated by lysosomal dysfunction participates in the cadmium exposure-induced cardiotoxicity in swine.

Xia Zhao, Xu Shi, Yujie Yao, Xiaojing Li, Shiwen Xu.

  Cadmium (Cd), a common toxic heavy metal, is believed as a risk factor for the induction and progression of cardiovascular disease. Autophagy is a highly ordered intracellular lysosomal-mediated degradation pathway that is crucial for protein and organelle quality control. Autophagy dysfunction could develop exacerbated cardiac dysfunction. However, the role of autophagy in Cd exposure-induced cardiotoxicity remains largely unknown. In this study, the Cd-induced swine cardiotoxicity model was established by feeding with a CdCl2 suppled diet (20 mg Cd/kg diet). The results showed that Cd exposure increased the expression of endoplasmic reticulum stress-related genes (GRP78, GRP94, IRE1, XBP1, PERK, ATF4, and ATF6), increased the expression of Ca2+ release channels IP3R and RYR1 and decreased the expression of Ca2+ uptake pump SERCA1. Cd exposure upregulated the expression of autophagy-related genes (CAMKKII, AMPK, ATG5, ATG7, ATG12, Beclin1, LC3-II, and P62) and downregulated mTOR expression. Cd exposure inhibited the expression of V-ATPase and cathepsins (CTSB and CTSD), and increased the expression of cathepsins in cytoplasm. Cd exposure decreased the colocalization of autophagosome and lysosome. This study revealed that autophagy flux inhibition caused by lysosomal dysfunction participates in the cardiotoxicity induced by Cd exposure in swine.

Keywords:  autophagy flux; cadmium; endoplasmic reticulum stress; lysosome; myocardium

DOI:  https://doi.org/10.1002/biof.1834
Proc Natl Acad Sci U S A. 2022 Mar 22. 119(12): e2100670119

Lysosomal inhibition sensitizes TMEM16A-expressing cancer cells to chemotherapy.

Avani Vyas, Roberto Gomez-Casal, Silvia Cruz-Rangel, Hugo Villanueva, Andrew G Sikora, Pavithra Rajagopalan, Devraj Basu, Jonathan Pacheco, Gerald R V Hammond, Kirill Kiselyov, Umamaheswar Duvvuri.

  SignificanceCisplatin is the first line therapy for patients with head and neck cancer. However, resistance to cisplatin remains a major concern. High expression of the calcium-activated chloride channel TMEM16A in tumors portends poor survival in these patients, possibly because of drug resistance. Here, we show that TMEM16A drives the sequestration of cisplatin into lysosomes. Subsequently, cisplatin is expelled via the delivery of lysosomes to the cell surface. We show that TMEM16A enhances this process, thereby promoting cisplatin resistance. We also show that lysosomal inhibition synergizes with cisplatin to induce tumor cell death. Our data uncovers a new fundamental feature of both lysosomal physiology and cancer cell biology that can potentially impact the treatment of patients with head and neck cancer.

Keywords:  MITF; TMEM16A; cisplatin; hydroxychloroquine; lysosomal flux

DOI:  https://doi.org/10.1073/pnas.2100670119
Autophagy. 2022 Mar 15.

Follicular lymphoma-associated mutations in the V-ATPase chaperone VMA21 activate autophagy creating a targetable dependency.

Fangyang Wang, Ying Yang, Gabriel Boudagh, Eeva-Liisa Eskelinen, Daniel J Klionsky, Sami N Malek.

  The discovery of recurrent mutations in subunits and regulators of the vacuolar-type H+-translocating ATPase (V-ATPase) in follicular lymphoma (FL) highlights a role for macroautophagy/autophagy, amino-acid, and nutrient-sensing pathways in the pathogenesis of this disease. Here, we report on novel mutations in the ER-resident chaperone VMA21, which is involved in V-ATPase assembly in 12% of FL. Mutations in a novel VMA21 hotspot (p.93X) result in the removal of a C-terminal non-canonical ER retrieval signal thus causing VMA21 mislocalization to lysosomes. The resulting impairment in V-ATPase function prevents full lysosomal acidification and function, including impaired pH-dependent protein degradation as shown via lysosomal metabolomics and ultimately causes a degree of amino acid depletion in the cytoplasm. These deficiencies result in compensatory autophagy activation, as measured using multiple complementary assays in human and yeast cells. Of translational significance, the compensatory activation of autophagy creates a dependency for survival for VMA21-mutated primary human FL as shown using inhibitors to ULK1, the proximal autophagy-regulating kinase. Using high-throughput microscopy-based screening assays for autophagy-inhibiting compounds, we identify multiple clinical grade cyclin-dependent kinase inhibitors as promising drugs and thus provide new rationale for innovative clinical trials in FL harboring aberrant V-ATPase.

Keywords:  Follicular lymphoma; VMA21 mutations; autophagy; lysosomal dysfunction; survival

DOI:  https://doi.org/10.1080/15548627.2022.2050663
Nat Commun. 2022 Mar 14. 13(1): 1300

Lysosomal Ca2+-mediated TFEB activation modulates mitophagy and functional adaptation of pancreatic β-cells to metabolic stress.

Kihyoun Park, Hyejin Lim, Jinyoung Kim, Yeseong Hwang, Yu Seol Lee, Soo Han Bae, Hyeongseok Kim, Hail Kim, Shin-Wook Kang, Joo Young Kim, Myung-Shik Lee.

Although autophagy is critical for pancreatic β-cell function, the role and mechanism of mitophagy in β-cells are unclear. We studied the role of lysosomal Ca2+ in TFEB activation by mitochondrial or metabolic stress and that of TFEB-mediated mitophagy in β-cell function. Mitochondrial or metabolic stress induced mitophagy through lysosomal Ca2+ release, increased cytosolic Ca2+ and TFEB activation. Lysosomal Ca2+ replenishment by ER- > lysosome Ca2+ refilling was essential for mitophagy. β-cell-specific Tfeb knockout (TfebΔβ-cell) abrogated high-fat diet (HFD)-induced mitophagy, accompanied by increased ROS and reduced mitochondrial cytochrome c oxidase activity or O2 consumption. TfebΔβ-cell mice showed aggravation of HFD-induced glucose intolerance and impaired insulin release. Metabolic or mitochondrial stress induced TFEB-dependent expression of mitophagy receptors including Ndp52 and Optn, contributing to the increased mitophagy. These results suggest crucial roles of lysosomal Ca2+ release coupled with ER- > lysosome Ca2+ refilling and TFEB activation in mitophagy and maintenance of pancreatic β-cell function during metabolic stress.

DOI: https://doi.org/10.1038/s41467-022-28874-9