bims-lypmec 2024-11-17 papers

Curr Biol. 2024 Nov 10. pii: S0960-9822(24)01457-X. [Epub ahead of print]

Lysosomal damage triggers a p38 MAPK-dependent phosphorylation cascade to promote lysophagy via the small heat shock protein HSP27.

Elizabeth R Gallagher, Peace T Oloko, Tessa C Fitch, Elizabeth M Brown, Lynn A Spruce, Erika L F Holzbaur.

Maintenance of lysosomal integrity is essential for cell viability. Upon injury, lysosomes may be targeted for degradation via a selective form of autophagy known as lysophagy. The engulfment of a damaged lysosome by an autophagosome is mediated by the recruitment of adaptor proteins, including SQSTM1/p62. p62 promotes lysophagy via the formation of phase-separated condensates in a mechanism that is regulated by the heat shock protein HSP27. Here, we demonstrate a direct interaction between HSP27 and p62. We used structural modeling to predict the binding interface between HSP27 and p62 and identify several disease-associated mutations that map to this interface. We used proteomics to identify post-translational modifications of HSP27 that regulate HSP27 recruitment to stressed lysosomes, finding robust phosphorylation at several serine residues. Next, we characterized the upstream signaling mechanism leading to HSP27 phosphorylation and found that p38 mitogen-activated protein kinase (MAPK) and its effector kinase MAP kinase-activated protein kinase 2 (MK2) are activated upon lysosomal damage by the kinase mTOR and the production of intracellular reactive oxygen species (ROS). Increased ROS activates p38 MAPK, which in turn allows MK2-dependent phosphorylation of HSP27. Depletion of HSP27 or the inhibition of HSP27 phosphorylation alters the dynamics of p62 condensates on stressed lysosomes, significantly inhibiting p62-dependent lysophagy. Thus, we define a novel lysosomal quality control mechanism in which lysosomal injury triggers a p38 MAPK/MK2 signaling cascade promoting p62-dependent lysophagy. Further, this signaling cascade is activated by many cellular stressors, including oxidative and heat stress, suggesting that other forms of selective autophagy may be regulated by p38 MAPK/MK2/HSP27.

Keywords: HSP27; SQSTM1/p62; lysophagy; oligomers; p38 MAPK; phase separation; phosphorylation

DOI: https://doi.org/10.1016/j.cub.2024.10.061

EMBO J. 2024 Nov 12.

Calcium signaling from damaged lysosomes induces cytoprotective stress granules.

Jacob Duran, Jay E Salinas, Rui Ping Wheaton, Suttinee Poolsup, Lee Allers, Monica Rosas-Lemus, Li Chen, Qiuying Cheng, Jing Pu, Michelle Salemi, Brett Phinney, Pavel Ivanov, Alf Håkon Lystad, Kiran Bhaskar, Jaya Rajaiya, Douglas J Perkins, Jingyue Jia.

Lysosomal damage induces stress granule (SG) formation. However, the importance of SGs in determining cell fate and the precise mechanisms that mediate SG formation in response to lysosomal damage remain unclear. Here, we describe a novel calcium-dependent pathway controlling SG formation, which promotes cell survival during lysosomal damage. Mechanistically, the calcium-activated protein ALIX transduces lysosomal damage signals to SG formation by controlling eIF2α phosphorylation after sensing calcium leakage. ALIX enhances eIF2α phosphorylation by promoting the association between PKR and its activator PACT, with galectin-3 inhibiting this interaction; these regulatory events occur on damaged lysosomes. We further find that SG formation plays a crucial role in promoting cell survival upon lysosomal damage caused by factors such as SARS-CoV-2ORF3a, adenovirus, malarial pigment, proteopathic tau, or environmental hazards. Collectively, these data provide insights into the mechanism of SG formation upon lysosomal damage and implicate it in diseases associated with damaged lysosomes and SGs.

Keywords: ALG2-ALIX; Calcium-dependent Pathway; Lysosomal Damage; PACT-PKR-eIF2α; Stress Granules

DOI: https://doi.org/10.1038/s44318-024-00292-1

Trends Cell Biol. 2024 Nov 14. pii: S0962-8924(24)00227-7. [Epub ahead of print]

TFEB links the cGAS-STING pathway to lysosome biogenesis.

Yaping Meng, Xinran Li, Haoxing Xu.

The cGAS-STING pathway senses the level of double-stranded (ds)DNA in the cytosol, and is required for innate immunity through its effector, TBK1. A recent study by Lv et al. reports that STING activation also simultaneously promotes lysosomal biogenesis by inducing nuclear translocation of the transcription factors TFEB/TFE3 independent of TBK1.

Keywords: STING; TFEB; cGAS; lysosome

DOI: https://doi.org/10.1016/j.tcb.2024.10.011

Cell. 2024 Nov 14. pii: S0092-8674(24)01215-7. [Epub ahead of print]187(23): 6518-6520

Bound by the love for cholesterol: A transporter meets a GPCR.

Divyanshu Tiwari, Nabarun Roy, Arun K Shukla.

In a recently published article in Nature, Bayly-Jones et al. report the cryo-EM structures of a lysosomal cholesterol sensor, LYCHOS, also known as GPR155, which reveals a unique fusion of a plant auxin-transporter-like domain with a seven-transmembrane GPCR-like domain and elucidates mechanistic insights into cellular regulation of mTORC1 activity.

DOI: https://doi.org/10.1016/j.cell.2024.10.033

Neurobiol Dis. 2024 Nov 07. pii: S0969-9961(24)00330-9. [Epub ahead of print]202 106728

LRRK2 and RAB8A regulate cell death after lysosomal damage in macrophages through cholesterol-related pathways.

Josefine Fussing Tengberg, Francesco Russo, Tau Benned-Jensen, Jacob Nielsen.

Activating mutations in Leucine Rich Repeat Kinase 2 (LRRK2) are among the most common genetic causes of Parkinson's disease (PD). The mechanistic path from LRRK2 mutations to PD is not established, but several lines of data suggest that LRRK2 modulation of lysosomal function is involved. It has previously been shown that LRRK2 is recruited to lysosomes upon lysosomal damage leading to increased phosphorylation of its RAB GTPase substrates in macrophage-derived RAW 264.7 cells. Here, we find that LRRK2 kinase inhibition reduces cell death induced by the lysosomotropic compound LLOMe in RAW 264.7 cells showing that lysosomal damage and LRRK2 functionally interacts in both directions: lysosomal damage can lead to activation of LRRK2 signaling and LRRK2 inhibition can attenuate LLOMe-induced cell death. The effect is lysosome specific, as only lysosomal stressors and not a variety of other cell death inducers could be modulated by LRRK2 kinase inhibition. We show with timing and Lysotracker experiments that LRRK2 inhibition does not affect the immediate lysosomal permeabilization induced by LLOMe, but rather modulates the subsequent cellular response to lysosomal damage. siRNA-mediated knockdown of LRRK2 and its main substrates, the RAB GTPases, showed that LRRK2 and RAB8A knockdown could attenuate LLOMe-induced cell death, but not other RAB GTPases tested. An RNA sequencing study was done to identify downstream pathways modulated by LLOMe and LRRK2 inhibition. The most striking finding was that almost all cholesterol biosynthesis genes were strongly downregulated by LLOMe and upregulated with LRRK2 inhibition in combination with LLOMe treatment. To explore the functional relevance of the transcriptional changes, we pretreated cells with the NPC1 inhibitor U18666A that can lead to accumulation of lysosomal cholesterol. U18666A-treated cells were less sensitive to LLOMe-induced cell death, but the attenuation of cell death by LRRK2 inhibition was strongly reduced suggesting that LRRK2 inhibition and lysosomal cholesterol reduces cell death by overlapping mechanisms. Thus, our data demonstrates a LRRK2- and RAB8A-mediated attenuation of RAW 264.7 cell death induced by lysosomal damage that is modulated by lysosomal cholesterol.

Keywords: Cholesterol; Endolysosomal damage; LRRK2; MLi-2; RNAseq; U18666A; mTOR

DOI: https://doi.org/10.1016/j.nbd.2024.106728

Nat Commun. 2024 Nov 16. 15(1): 9937

Functionally overlapping intra- and extralysosomal pathways promote bis(monoacylglycero)phosphate synthesis in mammalian cells.

Dominik Bulfon, Johannes Breithofer, Gernot F Grabner, Nermeen Fawzy, Anita Pirchheim, Heimo Wolinski, Dagmar Kolb, Lennart Hartig, Martin Tischitz, Clara Zitta, Greta Bramerdorfer, Achim Lass, Ulrike Taschler, Dagmar Kratky, Peter Greimel, Robert Zimmermann.

Bis(monoacylglycero)phosphate (BMP) is a major phospholipid constituent of intralumenal membranes in late endosomes/lysosomes, where it regulates the degradation and sorting of lipid cargo. Recent observations suggest that the Batten disease-associated protein CLN5 functions as lysosomal BMP synthase. Here, we show that transacylation reactions catalyzed by cytosolic and secreted enzymes enhance BMP synthesis independently of CLN5. The transacylases identified in this study are capable of acylating the precursor lipid phosphatidylglycerol (PG), generating acyl-PG, which is subsequently hydrolyzed to BMP. Extracellularly, acyl-PG and BMP are generated by endothelial lipase in cooperation with other serum enzymes of the pancreatic lipase family. The intracellular acylation of PG is catalyzed by several members of the cytosolic phospholipase A2 group IV (PLA2G4) family. Overexpression of secreted or cytosolic transacylases was sufficient to correct BMP deficiency in HEK293 cells lacking CLN5. Collectively, our observations suggest that functionally overlapping pathways promote BMP synthesis in mammalian cells.

DOI: https://doi.org/10.1038/s41467-024-54213-1

J Diabetes. 2024 Nov;16(11): e70033

Subcellular mass spectrometric detection unveils hyperglycemic memory in the diabetic heart.

Jiabing Zhan, Yufei Zhou, Yifan Chen, Kunying Jin, Zhaoyang Chen, Chen Chen, Huaping Li, Dao Wen Wang.

BACKGROUND: Intensive glycemic control is insufficient to reduce the risk of heart failure in patients with diabetes mellitus. While the hyperglycemic memory in the diabetic cardiomyopathy has been well documented, its underlying mechanisms are not fully understood. The present study tried to investigate whether the dysregulated proteins/biological pathways, which persistently altered in diabetic hearts during normoglycemia, participate in the hyperglycemic memory.
METHODS: Hearts of streptozotocin-induced diabetic mice, with or without intensive glycemic control using slow-release insulin implants, were collected. Proteins from total heart samples and subcellular fractions were assessed by mass spectrometry, Western blotting, and KEGG pathway enrichment analysis. mRNA sequencing was used to determine whether the persistently altered proteins were regulated at the transcriptional or post-transcriptional level.
RESULTS: Western blot validation of several proteins with high pathophysiological importance, including MYH7, HMGCS2, PDK4, and BDH1, indicated that mass spectrometry was able to qualitatively, but not quantitatively, reflect the fold changes of certain proteins in diabetes. Pathway analysis revealed that the peroxisome, PPAR pathway, and fatty acid metabolism could be efficiently rescued by glycemic control. However, dysregulation of oxidative phosphorylation and reactive oxygen species persisted even after normalization of hyperglycemia. Notably, mRNA sequencing revealed that dysregulated proteins in the oxidative phosphorylation pathway were not accompanied by coordinated changes in mRNA levels, indicating post-transcriptional regulation. Moreover, literature review and bioinformatics analysis suggested that hyperglycemia-induced persistent alterations of miRNAs targeted genes from the persistently dysregulated oxidative phosphorylation pathway, whereas, oxidative phosphorylation dysfunction-induced ROS regulated miRNA expression, which thereby might sustained the dysregulation of miRNAs.
CONCLUSIONS: Glycemic control cannot rescue hyperglycemia-induced alterations of subcellular proteins in the diabetic heart, and persistently altered proteins are involved in multiple functional pathways, including oxidative phosphorylation. These findings might provide novel insights into hyperglycemic memory in diabetic cardiomyopathy.

Keywords: diabetic cardiomyopathy; hyperglycemic memory; mass spectrometry; subcellular

DOI: https://doi.org/10.1111/1753-0407.70033