Santosh Phuyal

• Phuyal, Santosh; Romani, Patrizia; Dupont, Sirio & Farhan, Hesso (2023). Mechanobiology of organelles: illuminating their roles in mechanosensing and mechanotransduction. Trends in Cell Biology. ISSN 0962-8924. 33(12), p. 1049–1061. doi: 10.1016/j.tcb.2023.05.001.
• Ghannoum, Salim; Fantini, Damiano; Zahoor, Muhammad; Reiterer, Veronika; Phuyal, Santosh & Leoncio Netto, Waldir [Show all 13 contributors for this article] (2023). A combined experimental-computational approach uncovers a role for the Golgi matrix protein Giantin in breast cancer progression. PLoS Computational Biology. ISSN 1553-734X. 19(4). doi: 10.1371/journal.pcbi.1010995. Full text in Research Archive
• Radulovic, Maja; Wenzel, Eva; Gilani, Sania; Holland, Lya KK; Lystad, Alf Håkon Lohsen & Phuyal, Santosh [Show all 12 contributors for this article] (2022). Cholesterol transfer via endoplasmic reticulum contacts mediates lysosome damage repair. EMBO Journal. ISSN 0261-4189. 41:e112677, p. 1–18. doi: 10.15252/embj.2022112677. Full text in Research Archive Show summary

  Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol-binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP–ORP1L interactions. The PtdIns 4-kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage-induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol–PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response.

• Phuyal, Santosh; Djaerff, Elena; Le Roux, Anabel-Lise; Baker, Martin J; Fankhauser, Daniela & Mahdizadeh, Sayyed Jalil [Show all 16 contributors for this article] (2022). Mechanical strain stimulates COPII-dependent secretory trafficking via Rac1. EMBO Journal. ISSN 0261-4189. 41(18). doi: 10.15252/embj.2022110596. Full text in Research Archive
• Besse, Lenka; Besse, Andrej; Stolze, Sara C.; Sobh, Amin; Zaal, Esther A & Ham, Alwin J.Van Der [Show all 25 contributors for this article] (2021). Treatment with HIV-protease inhibitor nelfinavir identifies membrane lipid composition and fluidity as a therapeutic target in advanced multiple myeloma. Cancer Research. ISSN 0008-5472. 81(17), p. 4581–4593. doi: 10.1158/0008-5472.CAN-20-3323. Full text in Research Archive
• Alswady-Hoff, Mayes; Erdem, Johanna Maria Samulin; Phuyal, Santosh; Knittelfelder, Oskar; Sharma, Animesh & Fonseca, Davi de Miranda [Show all 9 contributors for this article] (2021). Long-Term Exposure to Nanosized TiO2 Triggers Stress Responses and Cell Death Pathways in Pulmonary Epithelial Cells. International Journal of Molecular Sciences. ISSN 1661-6596. 22(10). doi: 10.3390/ijms22105349. Full text in Research Archive Show summary

  There is little in vitro data available on long-term effects of TiO2 exposure. Such data are important for improving the understanding of underlying mechanisms of adverse health effects of TiO2 . Here, we exposed pulmonary epithelial cells to two doses (0.96 and 1.92 µg/cm2 ) of TiO2 for 13 weeks and effects on cell cycle and cell death mechanisms, i.e., apoptosis and autophagy were determined after 4, 8 and 13 weeks of exposure. Changes in telomere length, cellular protein levels and lipid classes were also analyzed at 13 weeks of exposure. We observed that the TiO2 exposure increased the fraction of cells in G1-phase and reduced the fraction of cells in G2-phase, which was accompanied by an increase in the fraction of late apoptotic/necrotic cells. This corresponded with an induced expression of key apoptotic proteins i.e., BAD and BAX, and an accumulation of several lipid classes involved in cellular stress and apoptosis. These findings were further supported by quantitative proteome profiling data showing an increase in proteins involved in cell stress and genomic maintenance pathways following TiO2 exposure. Altogether, we suggest that cell stress response and cell death pathways may be important molecular events in long-term health effects of TiO2

• Eck, Franziska; Phuyal, Santosh; Smith, Matthew D.; Kaulich, Manuel; Wilkinson, Simon & Farhan, Hesso [Show all 7 contributors for this article] (2020). ACSL3 is a novel GABARAPL2 interactor that links ufmylation and lipid droplet biogenesis. Journal of Cell Science. ISSN 0021-9533. 133(18), p. 1–16. doi: 10.1242/jcs.243477. Full text in Research Archive
• Phuyal, Santosh & Baschieri, Francesco (2020). Endomembranes: Unsung Heroes of Mechanobiology? Frontiers in Bioengineering and Biotechnology. ISSN 2296-4185. 8. doi: 10.3389/fbioe.2020.597721. Full text in Research Archive
• Phuyal, Santosh & Farhan, Hesso (2019). Multifaceted Rho GTPase Signaling at the Endomembranes. Frontiers in Cell and Developmental Biology. ISSN 2296-634X. 7(127). doi: 10.3389/fcell.2019.00127. Full text in Research Archive
• Phuyal, Santosh; Kasem, Mayes; Knittelfelder, Oskar; Sharma, Animesh; Fonseca, Davi de Miranda & Vebraite, Vaineta [Show all 10 contributors for this article] (2018). Characterization of the proteome and lipidome profiles of human lung cells after low dose and chronic exposure to multiwalled carbon nanotubes. Nanotoxicology. ISSN 1743-5390. 12(2), p. 138–152. doi: 10.1080/17435390.2018.1425500. Show summary

  The effects of long-term chronic exposure of human lung cells to multi-walled carbon nanotubes (MWCNT) and their impact upon cellular proteins and lipids were investigated. Since the lung is the major target organ, an in vitro normal bronchial epithelial cell line model was used. Additionally, to better mimic exposure to manufactured nanomaterials at occupational settings, cells were continuously exposed to two non-toxic and low doses of a MWCNT for 13-weeks. MWCNT-treatment increased ROS levels in cells without increasing oxidative DNA damage and resulted in differential expression of multiple anti- and pro-apoptotic proteins. The proteomic analysis of the MWCNT-exposed cells showed that among more than 5000 identified proteins; more than 200 were differentially expressed in the treated cells. Functional analyses revealed association of these differentially regulated proteins to cellular processes such as cell death and survival, cellular assembly, and organization. Similarly, shotgun lipidomic profiling revealed accumulation of multiple lipid classes. Our results indicate that long-term MWCNT-exposure of human normal lung cells at occupationally relevant low-doses may alter both the proteome and the lipidome profiles of the target epithelial cells in the lung.

• Vestad, Beate; Llorente, Alicia; Neurauter, Axl; Phuyal, Santosh; Kierulf, Bente & Kierulf, Peter [Show all 10 contributors for this article] (2017). Size and concentration analyses of extracellular vesicles by nanoparticle tracking analysis: a variation study. Journal of Extracellular Vesicles. ISSN 2001-3078. 6(1), p. 1–11. doi: 10.1080/20013078.2017.1344087. Full text in Research Archive
• Phuyal, Santosh; Kasem, Mayes; Rubio, Laura; Karlsson, Hanna; Marcos, Ricard & Skaug, Vidar [Show all 7 contributors for this article] (2017). Effects on human bronchial epithelial cells following low-dose chronic exposure to nanomaterials: A 6-month transformation study. Toxicology in Vitro. ISSN 0887-2333. 44, p. 230–240. doi: 10.1016/j.tiv.2017.07.016. Show summary

  The most plausible exposure route to manufactured nanomaterials (MNM) remains pulmonary inhalation. Yet, few studies have attempted to assess carcinogenic properties in vitro following long-term exposure of human pulmonary cells to low and occupationally relevant doses. The most advanced in vitro tests for carcinogenicity, the cell transformation assay (CTA), rely mostly on rodent cells and short-term exposure. We hypothesized that long-term exposure of human bronchial epithelial cells with a normal phenotype could be a valuable assay for testing carcinogenicity of nanomaterials. Therefore, this study (performed within the framework of the FP7-NANoREG project) assessed carcinogenic potential of chronic exposure (up to 6 months) to low doses of multi-walled carbon nanotubes (MWCNT, NM-400 and NM-401) and TiO2 materials (NM62002 and KC7000). In order to harmonize and standardize the experiments, standard operating protocols of MNM dispersion (NANOGENOTOX) were used by three different NANoREG project partners. All nanomaterials showed low cytotoxicity in short-term tests for the tested doses (0.96 and 1.92 μg/cm2). During long-term exposure, however, NM-401 clearly affected cell proliferation. In contrast, no cell transformation was observed for NM-401 by any of the partners. NM-400 and NM62002 formed some colonies after 3 months. We conclude that agglomerated NM-401 in low doses affect cell proliferation but do not cause cell transformation in the CTA assay used.

• Moen, Aurora Lie; Jacobsen, Daniel ; Phuyal, Santosh; Legfeldt, Anna; Haugen, Fred & Røe, Cecilie [Show all 7 contributors for this article] (2017). MicroRNA-223 demonstrated experimentally in exosome-like vesicles is associated with decreased risk of persistent pain after lumbar disc herniation. Journal of Translational Medicine. ISSN 1479-5876. 15(89), p. 1–11. doi: 10.1186/s12967-017-1194-8. Full text in Research Archive Show summary

  Background: Previous findings have demonstrated that lumbar radicular pain after disc herniation may be associated with up-regulation of inflammatory mediators. In the present study we examined the possible role of extracellular microRNAs (miRs) in this process. Methods: Single unit recordings, isolation of exosome-like vesicles, electron microscopy, nanoparticle tracking analysis, western blot analysis and qPCR were used in rats to demonstrate the effect of nucleus pulposus (NP) applied onto the dorsal nerve roots. ELISA and qPCR were used to measure the level of circulating IL-6 and miRs in a 1-year observational study in patients after disc herniation. Results: In the rats, enhanced spinal cord nociceptive responses were displayed after NP applied onto the dorsal nerve roots. An increased release of small non-coding RNAs, including miR-223, miR-760 and miR-145, from NP in exosome-like vesicles was demonstrated. In particular, the NP expression of miR-223, which inhibited the nociceptive spinal signalling, was increased. In the patients, increased extracellular miR-223 was also verified in the acute phase after disc herniation. The increased miR-223 expression was, however, only observed in those who recovered (sex, age and smoking were included as covariates). Conclusions: Our findings suggest that miR-223, which can be released from the NP after disc herniation, attenuates the neuronal activity in the pain pathways. Dysregulation of miR-223 may predict chronic lumbar radicular pain.

• Phuyal, Santosh; Skotland, Tore; Hessvik, Nina Pettersen; Simolin, Helena; Øverbye, Anders & Brech, Andreas [Show all 10 contributors for this article] (2015). The ether lipid precursor hexadecylglycerol stimulates the release and changes the composition of exosomes derived from PC-3 cells. Journal of Biological Chemistry. ISSN 0021-9258. 290, p. 4225–4237. doi: 10.1074/jbc.M114.593962.
• Phuyal, Santosh; Hessvik, Nina Pettersen; Skotland, Tore; Sandvig, Kirsten & Llorente, Alicia (2014). Regulation of exosome release by glycosphingolipids and flotillins. The FEBS Journal. ISSN 1742-464X. 281(9), p. 2214–2227. doi: 10.1111/febs.12775.
• Hessvik, Nina Pettersen; Phuyal, Santosh; Brech, Andreas; Sandvig, Kirsten & Llorente, Alicia (2012). Profiling of microRNAs in exosomes released from PC-3 prostate cancer cells. Biochimica et Biophysica Acta. Gene Regulatory Mechanisms. ISSN 1874-9399. 1819(11-12), p. 1154–1163. doi: 10.1016/j.bbagrm.2012.08.016.

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• Gilmour, Brian Christopher; Chan, Hanna; Borowicz, Pawel; Phuyal, Santosh; Lossius, Andreas & Spurkland, Anne (2023). β2, or not β2? Connecting the Lck-adaptor TSAd to integrin β2 via a combined wet lab & bioinformatic approach.
• Phuyal, Santosh & Farhan, Hesso (2021). Want to leave the ER? We offer vesicles, tubules, and tunnels. Journal of Cell Biology. ISSN 0021-9525. 220(6). doi: 10.1083/jcb.202104062.
• Auer, Susann; Haelterman, Nele A.; Weissgerber, Tracey L.; Erlich, Jeffrey C.; Susilaradeya, Damar & Julkowska, Magdalena [Show all 38 contributors for this article] (2021). Science forum: A community-led initiative for training in reproducible research. eLIFE. ISSN 2050-084X. 10. doi: 10.7554/eLife.64719. Full text in Research Archive
• Zienolddiny, Shanbeh; Phuyal, Santosh; Kasem, Mayes; Knittelfelder, Oskar; Sharma, Animesh & Fonseca, Davi de Miranda [Show all 8 contributors for this article] (2017). Toxicoproteomic effects of carbon nanotubes on chronically exposed human lung cells. Toxicology Letters. ISSN 0378-4274. 280. doi: 10.1016/j.toxlet.2017.07.395.
• Phuyal, Santosh; Llorente, Alicia & Sandvig, Kirsten (2014). Composition and cellular release of exosomes from cancer cells. Akademika publishing.

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