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Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins

Matausz Pospiech, Sian-eleri Owens Orcid Logo, David J. Miller, Karl Austin-Muttitt, Jonathan Mullins Orcid Logo, James Cronin Orcid Logo, Rudolf K. Allemann, Martin Sheldon Orcid Logo

The FASEB Journal, Volume: 35, Issue: 6, Pages: 1 - 19

Swansea University Authors: Matausz Pospiech, Sian-eleri Owens Orcid Logo, Karl Austin-Muttitt, Jonathan Mullins Orcid Logo, James Cronin Orcid Logo, Martin Sheldon Orcid Logo

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DOI (Published version): 10.1096/fj.202100164r

Abstract

Certain species of pathogenic bacteria damage tissues by secreting cholesterol‐dependent cytolysins, which form pores in the plasma membranes of animal cells. However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed b...

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Published in: The FASEB Journal
ISSN: 0892-6638 1530-6860
Published: Wiley 2021
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URI: https://cronfa.swan.ac.uk/Record/cronfa56884
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However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed by squalene synthase, we explored whether inhibiting this enzyme protected cells against cholesterol&#x2010;dependent cytolysins. We first synthesized 22 different nitrogen&#x2010;containing bisphosphonate molecules that were designed to inhibit squalene synthase. Squalene synthase inhibition was quantified using a cell&#x2010;free enzyme assay, and validated by computer modeling of bisphosphonate molecules binding to squalene synthase. The bisphosphonates were then screened for their ability to protect HeLa cells against the damage caused by the cholesterol&#x2010;dependent cytolysin, pyolysin. The most effective bisphosphonate reduced pyolysin&#x2010;induced leakage of lactate dehydrogenase into cell supernatants by &gt;80%, and reduced pyolysin&#x2010;induced cytolysis from &gt;75% to &lt;25%. In addition, this bisphosphonate reduced pyolysin&#x2010;induced leakage of potassium from cells, limited changes in the cytoskeleton, prevented mitogen&#x2010;activated protein kinases cell stress responses, and reduced cellular cholesterol. The bisphosphonate also protected cells against another cholesterol&#x2010;dependent cytolysin, streptolysin O, and protected lung epithelial cells and primary dermal fibroblasts against cytolysis. 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spelling 2022-05-10T13:18:23.6630813 v2 56884 2021-05-17 Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins 9e76dd1d6563d6c22720373f0ad861e0 Matausz Pospiech Matausz Pospiech true false 721deb4604d122019244cfdf08820cbe 0000-0003-1806-5235 Sian-eleri Owens Sian-eleri Owens true false fafc0917b48af4eaec154646854867f8 Karl Austin-Muttitt Karl Austin-Muttitt true false 4cf2dddedbe1dacb506ec925fdbd5b40 0000-0003-0144-2962 Jonathan Mullins Jonathan Mullins true false 9cfd17551c0d1f7438895121e4fbb6e8 0000-0002-0590-9462 James Cronin James Cronin true false ab0f74b794e59cc270c69e63ee1d9748 0000-0001-7902-5558 Martin Sheldon Martin Sheldon true false 2021-05-17 BMS Certain species of pathogenic bacteria damage tissues by secreting cholesterol‐dependent cytolysins, which form pores in the plasma membranes of animal cells. However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed by squalene synthase, we explored whether inhibiting this enzyme protected cells against cholesterol‐dependent cytolysins. We first synthesized 22 different nitrogen‐containing bisphosphonate molecules that were designed to inhibit squalene synthase. Squalene synthase inhibition was quantified using a cell‐free enzyme assay, and validated by computer modeling of bisphosphonate molecules binding to squalene synthase. The bisphosphonates were then screened for their ability to protect HeLa cells against the damage caused by the cholesterol‐dependent cytolysin, pyolysin. The most effective bisphosphonate reduced pyolysin‐induced leakage of lactate dehydrogenase into cell supernatants by >80%, and reduced pyolysin‐induced cytolysis from >75% to <25%. In addition, this bisphosphonate reduced pyolysin‐induced leakage of potassium from cells, limited changes in the cytoskeleton, prevented mitogen‐activated protein kinases cell stress responses, and reduced cellular cholesterol. The bisphosphonate also protected cells against another cholesterol‐dependent cytolysin, streptolysin O, and protected lung epithelial cells and primary dermal fibroblasts against cytolysis. Our findings imply that treatment with bisphosphonates that inhibit squalene synthase might help protect tissues against pathogenic bacteria that secrete cholesterol‐dependent cytolysins. Journal Article The FASEB Journal 35 6 1 19 Wiley 0892-6638 1530-6860 bisphosphonate; cholesterol cholesterol‐dependent cytolysins; cytoprotection; pore‐forming toxins; squalene synthase 1 6 2021 2021-06-01 10.1096/fj.202100164r COLLEGE NANME Biomedical Sciences COLLEGE CODE BMS Swansea University SU Library paid the OA fee (TA Institutional Deal) Sêr Cymru National Research Network of the Welsh Government NRNS3APR009 2022-05-10T13:18:23.6630813 2021-05-17T14:02:15.3014973 Swansea University Medical School Swansea University Medical School Matausz Pospiech 1 Sian-eleri Owens 0000-0003-1806-5235 2 David J. Miller 3 Karl Austin-Muttitt 4 Jonathan Mullins 0000-0003-0144-2962 5 James Cronin 0000-0002-0590-9462 6 Rudolf K. Allemann 7 Martin Sheldon 0000-0001-7902-5558 8 56884__19915__79ea9e12887549c98aa7eba1cea4d3fd.pdf fj.202100164R.pdf 2021-05-17T14:06:40.8276853 Output 1943195 application/pdf Version of Record true © 2021 The Authors. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License true eng https://creativecommons.org/licenses/by-nc/4.0/
title Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
spellingShingle Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
Matausz Pospiech
Sian-eleri Owens
Karl Austin-Muttitt
Jonathan Mullins
James Cronin
Martin Sheldon
title_short Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
title_full Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
title_fullStr Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
title_full_unstemmed Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
title_sort Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins
author_id_str_mv 9e76dd1d6563d6c22720373f0ad861e0
721deb4604d122019244cfdf08820cbe
fafc0917b48af4eaec154646854867f8
4cf2dddedbe1dacb506ec925fdbd5b40
9cfd17551c0d1f7438895121e4fbb6e8
ab0f74b794e59cc270c69e63ee1d9748
author_id_fullname_str_mv 9e76dd1d6563d6c22720373f0ad861e0_***_Matausz Pospiech
721deb4604d122019244cfdf08820cbe_***_Sian-eleri Owens
fafc0917b48af4eaec154646854867f8_***_Karl Austin-Muttitt
4cf2dddedbe1dacb506ec925fdbd5b40_***_Jonathan Mullins
9cfd17551c0d1f7438895121e4fbb6e8_***_James Cronin
ab0f74b794e59cc270c69e63ee1d9748_***_Martin Sheldon
author Matausz Pospiech
Sian-eleri Owens
Karl Austin-Muttitt
Jonathan Mullins
James Cronin
Martin Sheldon
author2 Matausz Pospiech
Sian-eleri Owens
David J. Miller
Karl Austin-Muttitt
Jonathan Mullins
James Cronin
Rudolf K. Allemann
Martin Sheldon
format Journal article
container_title The FASEB Journal
container_volume 35
container_issue 6
container_start_page 1
publishDate 2021
institution Swansea University
issn 0892-6638
1530-6860
doi_str_mv 10.1096/fj.202100164r
publisher Wiley
college_str Swansea University Medical School
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hierarchy_top_id swanseauniversitymedicalschool
hierarchy_top_title Swansea University Medical School
hierarchy_parent_id swanseauniversitymedicalschool
hierarchy_parent_title Swansea University Medical School
department_str Swansea University Medical School{{{_:::_}}}Swansea University Medical School{{{_:::_}}}Swansea University Medical School
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description Certain species of pathogenic bacteria damage tissues by secreting cholesterol‐dependent cytolysins, which form pores in the plasma membranes of animal cells. However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed by squalene synthase, we explored whether inhibiting this enzyme protected cells against cholesterol‐dependent cytolysins. We first synthesized 22 different nitrogen‐containing bisphosphonate molecules that were designed to inhibit squalene synthase. Squalene synthase inhibition was quantified using a cell‐free enzyme assay, and validated by computer modeling of bisphosphonate molecules binding to squalene synthase. The bisphosphonates were then screened for their ability to protect HeLa cells against the damage caused by the cholesterol‐dependent cytolysin, pyolysin. The most effective bisphosphonate reduced pyolysin‐induced leakage of lactate dehydrogenase into cell supernatants by >80%, and reduced pyolysin‐induced cytolysis from >75% to <25%. In addition, this bisphosphonate reduced pyolysin‐induced leakage of potassium from cells, limited changes in the cytoskeleton, prevented mitogen‐activated protein kinases cell stress responses, and reduced cellular cholesterol. The bisphosphonate also protected cells against another cholesterol‐dependent cytolysin, streptolysin O, and protected lung epithelial cells and primary dermal fibroblasts against cytolysis. Our findings imply that treatment with bisphosphonates that inhibit squalene synthase might help protect tissues against pathogenic bacteria that secrete cholesterol‐dependent cytolysins.
published_date 2021-06-01T04:12:42Z
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