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Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells

Cancer stem cells (CSCs) are crucial regulators of tumor recurrence/progression. The maintenance of CSCs is dependent on aberrant activation of various pathways, including Hedgehog. Prevalent sialylations contribute to aggressiveness in CSCs. Here, we have addressed the role of sialylation in regula...

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Autores principales: Nath, Shalini, Mondal, Susmita, Butti, Ramesh, Prasanna Gunasekaran, Vinoth, Chatterjee, Uttara, Halder, Aniket, Kundu, Gopal C., Mandal, Chitra
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349614/
https://www.ncbi.nlm.nih.gov/pubmed/32575925
http://dx.doi.org/10.3390/cells9061512
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author Nath, Shalini
Mondal, Susmita
Butti, Ramesh
Prasanna Gunasekaran, Vinoth
Chatterjee, Uttara
Halder, Aniket
Kundu, Gopal C.
Mandal, Chitra
author_facet Nath, Shalini
Mondal, Susmita
Butti, Ramesh
Prasanna Gunasekaran, Vinoth
Chatterjee, Uttara
Halder, Aniket
Kundu, Gopal C.
Mandal, Chitra
author_sort Nath, Shalini
collection PubMed
description Cancer stem cells (CSCs) are crucial regulators of tumor recurrence/progression. The maintenance of CSCs is dependent on aberrant activation of various pathways, including Hedgehog. Prevalent sialylations contribute to aggressiveness in CSCs. Here, we have addressed the role of sialylation in regulating stemness-like properties of pancreatic cancer sphere-forming cells (PCS) through modulation of the Hedgehog (Hh) pathway. The status of CD133/CD44/surface-sialylation was checked by flow cytometry and effects of Neu2 overexpression in PCS were compared using qPCR, immunoblotting, co-immunoprecipitation and also by colony-formation assays. The work was also validated in a xenograft model after Neu2 overexpression. Neu2 and Shh status in patient tissues were examined by immunohistochemistry. PCS showed higher Hh-pathway activity and sialylation with reduced cytosolic-sialidase (Neu2). Neu2 overexpression caused desialylation of Shh, thereby reducing Shh-Patched1 binding thus causing decreased Hh-pathway activity with lower expression of Snail/Slug/CyclinD1 leading to reduction of stemness-like properties. Neu2-overexpression also induced apoptosis in PCS. Additionally, Neu2-overexpressed PCS demonstrated lower mTORC2 formation and inhibitory-phosphorylation of Gsk3β, reflecting a close relationship with reduced Hh pathway. Moreover, both Neu2 and Rictor (a major component of mTORC2) co-transfection reduced stem cell markers and Hh-pathway activity in PCS. Neu2-overexpressed tumors showed reduction in tumor mass with downregulation of stem cell markers/Shh/mTOR and upregulation of Bax/Caspase8/Caspase3. Thus, we established that reduced sialylation by Neu2 overexpression leads to decreased stemness-like properties by desialylation of Shh, which impaired its association with Patched1 thereby inhibiting the Hh pathway. All these may be responsible for enhanced apoptosis in Neu2-overexpressed PCS.
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spelling pubmed-73496142020-07-14 Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells Nath, Shalini Mondal, Susmita Butti, Ramesh Prasanna Gunasekaran, Vinoth Chatterjee, Uttara Halder, Aniket Kundu, Gopal C. Mandal, Chitra Cells Article Cancer stem cells (CSCs) are crucial regulators of tumor recurrence/progression. The maintenance of CSCs is dependent on aberrant activation of various pathways, including Hedgehog. Prevalent sialylations contribute to aggressiveness in CSCs. Here, we have addressed the role of sialylation in regulating stemness-like properties of pancreatic cancer sphere-forming cells (PCS) through modulation of the Hedgehog (Hh) pathway. The status of CD133/CD44/surface-sialylation was checked by flow cytometry and effects of Neu2 overexpression in PCS were compared using qPCR, immunoblotting, co-immunoprecipitation and also by colony-formation assays. The work was also validated in a xenograft model after Neu2 overexpression. Neu2 and Shh status in patient tissues were examined by immunohistochemistry. PCS showed higher Hh-pathway activity and sialylation with reduced cytosolic-sialidase (Neu2). Neu2 overexpression caused desialylation of Shh, thereby reducing Shh-Patched1 binding thus causing decreased Hh-pathway activity with lower expression of Snail/Slug/CyclinD1 leading to reduction of stemness-like properties. Neu2-overexpression also induced apoptosis in PCS. Additionally, Neu2-overexpressed PCS demonstrated lower mTORC2 formation and inhibitory-phosphorylation of Gsk3β, reflecting a close relationship with reduced Hh pathway. Moreover, both Neu2 and Rictor (a major component of mTORC2) co-transfection reduced stem cell markers and Hh-pathway activity in PCS. Neu2-overexpressed tumors showed reduction in tumor mass with downregulation of stem cell markers/Shh/mTOR and upregulation of Bax/Caspase8/Caspase3. Thus, we established that reduced sialylation by Neu2 overexpression leads to decreased stemness-like properties by desialylation of Shh, which impaired its association with Patched1 thereby inhibiting the Hh pathway. All these may be responsible for enhanced apoptosis in Neu2-overexpressed PCS. MDPI 2020-06-21 /pmc/articles/PMC7349614/ /pubmed/32575925 http://dx.doi.org/10.3390/cells9061512 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Nath, Shalini
Mondal, Susmita
Butti, Ramesh
Prasanna Gunasekaran, Vinoth
Chatterjee, Uttara
Halder, Aniket
Kundu, Gopal C.
Mandal, Chitra
Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title_full Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title_fullStr Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title_full_unstemmed Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title_short Desialylation of Sonic-Hedgehog by Neu2 Inhibits Its Association with Patched1 Reducing Stemness-Like Properties in Pancreatic Cancer Sphere-forming Cells
title_sort desialylation of sonic-hedgehog by neu2 inhibits its association with patched1 reducing stemness-like properties in pancreatic cancer sphere-forming cells
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349614/
https://www.ncbi.nlm.nih.gov/pubmed/32575925
http://dx.doi.org/10.3390/cells9061512
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