Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation

Aurélie Philippe; Gunnar Kleinau; Jason Jannis Gruner; Sumin Wu; Daniel Postpieszala; David Speck; Harald Heidecke; Simon J. Dowell; Gabriela Riemekasten; Peter W. Hildebrand; Julian Kamhieh-Milz; Rusan Catar; Michal Szczepek; Duska Dragun; Patrick Scheerer

doi:10.3390/ijms23073984

Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation

Aurélie Philippe^*, Gunnar Kleinau, Jason Jannis Gruner, Sumin Wu, Daniel Postpieszala, David Speck, Harald Heidecke, Simon J. Dowell, Gabriela Riemekasten, Peter W. Hildebrand, Julian Kamhieh-Milz, Rusan Catar, Michal Szczepek, Duska Dragun, Patrick Scheerer^*

^*Corresponding author for this work

Clinic of Rheumatology

6 Citations (Scopus)

Abstract

The angiotensin II (Ang II) type 1 receptor (AT₁ R) is involved in the regulation of blood pressure (through vasoconstriction) and water and ion homeostasis (mediated by interaction with the endogenous agonist). AT₁ R can also be activated by auto-antibodies (AT₁ R-Abs), which are associated with manifold diseases, such as obliterative vasculopathy, preeclampsia and systemic sclerosis. Knowledge of the molecular mechanisms related to AT₁ R-Abs binding and associated signaling cascade (dys-)regulation remains fragmentary. The goal of this study was, therefore, to investigate details of the effects of AT₁ R-Abs on G-protein signaling and subsequent cell proliferation, as well as the putative contribution of the three extracellular receptor loops (ELs) to Abs-AT₁ R signaling. AT₁ R-Abs induced nuclear factor of activated T-cells (NFAT) signaling, which reflects G_q/11 and G_i activation. The impact on cell proliferation was tested in different cell systems, as well as activation-triggered receptor internalization. Blockwise alanine substitutions were designed to potentially investigate the role of ELs in AT₁ R-Abs-mediated effects. First, we demonstrate that Ang II-mediated internalization of AT₁ R is impeded by binding of AT₁ R-Abs. Secondly, exclusive AT₁ R-Abs-induced G_q/11 activation is most significant for NFAT stimulation and mediates cell proliferation. Interestingly, our studies also reveal that ligand-independent, baseline AT₁ R activation of G_i signaling has, in turn, a negative effect on cell proliferation. Indeed, inhibition of G_i basal activity potentiates proliferation triggered by AT₁ R-Abs. Finally, although AT₁ R containing EL1 and EL3 blockwise alanine mutations were not expressed on the human embryonic kidney293T (HEK293T) cell surface, we at least confirmed that parts of EL2 are involved in interactions between AT₁ R and Abs. This current study thus provides extended insights into the molecular action of AT₁ R-Abs and associated mechanisms of interrelated pathogenesis.

Original language	English
Article number	3984
Journal	International Journal of Molecular Sciences
Volume	23
Issue number	7
ISSN	1661-6596
DOIs	https://doi.org/10.3390/ijms23073984
Publication status	Published - 01.04.2022

Funding

Acknowledgments: The authors would like to dedicate this manuscript to Duska Dragun who was the first to understand the potential of this study. The authors are grateful to Heike Biebermann (Charité—Universitätsmedizin Berlin) for providing the pGL4.34 plasmids used in the luciferase reporter assays. This research was supported by the German Research Foundation (DFG, Deutsche Forschungsgemeinschaft) project-ID 394046635—SFB1365 (subproject A03 to D.D. and P.S.), project-ID 421152132—SFB1423 (subprojects A01/A05/Z03 to P.S. and A03/C01/Z04 to P.W.H.), and project-ID 25065445—SFB740 (subproject B6 to P.W.H. and P.S.). This research was also funded by the European Union’s Horizon 2020 MSCA Program under grant agreement 956314 (ALLODD) (G.K. and P.S.). The work was further supported through Stiftung Charité and the Einstein Center Digital Future (to P.W.H.). This research was funded by the German Research Foundation (DFG, Deutsche Forschungsgemeinschaft) project-ID 394046635?SFB1365 (subproject A03 to D.D. and P.S.), project-ID 421152132? SFB1423 (subprojects A01/A05/Z03 to P.S. and A03/C01/Z04 to P.W.H.), and project-ID 25065445?SFB740 (subproject B6 to P.W.H. and P.S.). This research was also funded by the European Union?s Horizon 2020 MSCA Program under grant agreement 956314 (ALLODD) (G.K. and P.S.). Funding: This research was funded by the German Research Foundation (DFG, Deutsche Forschungs-gemeinschaft) project-ID 394046635—SFB1365 (subproject A03 to D.D. and P.S.), project-ID 421152132— SFB1423 (subprojects A01/A05/Z03 to P.S. and A03/C01/Z04 to P.W.H.), and project-ID 25065445—SFB740 (subproject B6 to P.W.H. and P.S.). This research was also funded by the European Union’s Horizon 2020 MSCA Program under grant agreement 956314 (ALLODD) (G.K. and P.S.).

Research Areas and Centers

Academic Focus: Center for Infection and Inflammation Research (ZIEL)

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.3390/ijms23073984

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Philippe, A., Kleinau, G., Gruner, J. J., Wu, S., Postpieszala, D., Speck, D., Heidecke, H., Dowell, S. J., Riemekasten, G., Hildebrand, P. W., Kamhieh-Milz, J., Catar, R., Szczepek, M., Dragun, D., & Scheerer, P. (2022). Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation. International Journal of Molecular Sciences, 23(7), Article 3984. https://doi.org/10.3390/ijms23073984

@article{c632095523514411bd526fd9cbaa2b13,

title = "Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation",

abstract = "The angiotensin II (Ang II) type 1 receptor (AT1 R) is involved in the regulation of blood pressure (through vasoconstriction) and water and ion homeostasis (mediated by interaction with the endogenous agonist). AT1 R can also be activated by auto-antibodies (AT1 R-Abs), which are associated with manifold diseases, such as obliterative vasculopathy, preeclampsia and systemic sclerosis. Knowledge of the molecular mechanisms related to AT1 R-Abs binding and associated signaling cascade (dys-)regulation remains fragmentary. The goal of this study was, therefore, to investigate details of the effects of AT1 R-Abs on G-protein signaling and subsequent cell proliferation, as well as the putative contribution of the three extracellular receptor loops (ELs) to Abs-AT1 R signaling. AT1 R-Abs induced nuclear factor of activated T-cells (NFAT) signaling, which reflects Gq/11 and Gi activation. The impact on cell proliferation was tested in different cell systems, as well as activation-triggered receptor internalization. Blockwise alanine substitutions were designed to potentially investigate the role of ELs in AT1 R-Abs-mediated effects. First, we demonstrate that Ang II-mediated internalization of AT1 R is impeded by binding of AT1 R-Abs. Secondly, exclusive AT1 R-Abs-induced Gq/11 activation is most significant for NFAT stimulation and mediates cell proliferation. Interestingly, our studies also reveal that ligand-independent, baseline AT1 R activation of Gi signaling has, in turn, a negative effect on cell proliferation. Indeed, inhibition of Gi basal activity potentiates proliferation triggered by AT1 R-Abs. Finally, although AT1 R containing EL1 and EL3 blockwise alanine mutations were not expressed on the human embryonic kidney293T (HEK293T) cell surface, we at least confirmed that parts of EL2 are involved in interactions between AT1 R and Abs. This current study thus provides extended insights into the molecular action of AT1 R-Abs and associated mechanisms of interrelated pathogenesis.",

author = "Aur{\'e}lie Philippe and Gunnar Kleinau and Gruner, \{Jason Jannis\} and Sumin Wu and Daniel Postpieszala and David Speck and Harald Heidecke and Dowell, \{Simon J.\} and Gabriela Riemekasten and Hildebrand, \{Peter W.\} and Julian Kamhieh-Milz and Rusan Catar and Michal Szczepek and Duska Dragun and Patrick Scheerer",

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Philippe, A, Kleinau, G, Gruner, JJ, Wu, S, Postpieszala, D, Speck, D, Heidecke, H, Dowell, SJ, Riemekasten, G, Hildebrand, PW, Kamhieh-Milz, J, Catar, R, Szczepek, M, Dragun, D & Scheerer, P 2022, 'Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation', International Journal of Molecular Sciences, vol. 23, no. 7, 3984. https://doi.org/10.3390/ijms23073984

TY - JOUR

T1 - Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation

AU - Philippe, Aurélie

AU - Kleinau, Gunnar

AU - Gruner, Jason Jannis

AU - Wu, Sumin

AU - Postpieszala, Daniel

AU - Speck, David

AU - Heidecke, Harald

AU - Dowell, Simon J.

AU - Riemekasten, Gabriela

AU - Hildebrand, Peter W.

AU - Kamhieh-Milz, Julian

AU - Catar, Rusan

AU - Szczepek, Michal

AU - Dragun, Duska

AU - Scheerer, Patrick

PY - 2022/4/1

Y1 - 2022/4/1

N2 - The angiotensin II (Ang II) type 1 receptor (AT1 R) is involved in the regulation of blood pressure (through vasoconstriction) and water and ion homeostasis (mediated by interaction with the endogenous agonist). AT1 R can also be activated by auto-antibodies (AT1 R-Abs), which are associated with manifold diseases, such as obliterative vasculopathy, preeclampsia and systemic sclerosis. Knowledge of the molecular mechanisms related to AT1 R-Abs binding and associated signaling cascade (dys-)regulation remains fragmentary. The goal of this study was, therefore, to investigate details of the effects of AT1 R-Abs on G-protein signaling and subsequent cell proliferation, as well as the putative contribution of the three extracellular receptor loops (ELs) to Abs-AT1 R signaling. AT1 R-Abs induced nuclear factor of activated T-cells (NFAT) signaling, which reflects Gq/11 and Gi activation. The impact on cell proliferation was tested in different cell systems, as well as activation-triggered receptor internalization. Blockwise alanine substitutions were designed to potentially investigate the role of ELs in AT1 R-Abs-mediated effects. First, we demonstrate that Ang II-mediated internalization of AT1 R is impeded by binding of AT1 R-Abs. Secondly, exclusive AT1 R-Abs-induced Gq/11 activation is most significant for NFAT stimulation and mediates cell proliferation. Interestingly, our studies also reveal that ligand-independent, baseline AT1 R activation of Gi signaling has, in turn, a negative effect on cell proliferation. Indeed, inhibition of Gi basal activity potentiates proliferation triggered by AT1 R-Abs. Finally, although AT1 R containing EL1 and EL3 blockwise alanine mutations were not expressed on the human embryonic kidney293T (HEK293T) cell surface, we at least confirmed that parts of EL2 are involved in interactions between AT1 R and Abs. This current study thus provides extended insights into the molecular action of AT1 R-Abs and associated mechanisms of interrelated pathogenesis.

AB - The angiotensin II (Ang II) type 1 receptor (AT1 R) is involved in the regulation of blood pressure (through vasoconstriction) and water and ion homeostasis (mediated by interaction with the endogenous agonist). AT1 R can also be activated by auto-antibodies (AT1 R-Abs), which are associated with manifold diseases, such as obliterative vasculopathy, preeclampsia and systemic sclerosis. Knowledge of the molecular mechanisms related to AT1 R-Abs binding and associated signaling cascade (dys-)regulation remains fragmentary. The goal of this study was, therefore, to investigate details of the effects of AT1 R-Abs on G-protein signaling and subsequent cell proliferation, as well as the putative contribution of the three extracellular receptor loops (ELs) to Abs-AT1 R signaling. AT1 R-Abs induced nuclear factor of activated T-cells (NFAT) signaling, which reflects Gq/11 and Gi activation. The impact on cell proliferation was tested in different cell systems, as well as activation-triggered receptor internalization. Blockwise alanine substitutions were designed to potentially investigate the role of ELs in AT1 R-Abs-mediated effects. First, we demonstrate that Ang II-mediated internalization of AT1 R is impeded by binding of AT1 R-Abs. Secondly, exclusive AT1 R-Abs-induced Gq/11 activation is most significant for NFAT stimulation and mediates cell proliferation. Interestingly, our studies also reveal that ligand-independent, baseline AT1 R activation of Gi signaling has, in turn, a negative effect on cell proliferation. Indeed, inhibition of Gi basal activity potentiates proliferation triggered by AT1 R-Abs. Finally, although AT1 R containing EL1 and EL3 blockwise alanine mutations were not expressed on the human embryonic kidney293T (HEK293T) cell surface, we at least confirmed that parts of EL2 are involved in interactions between AT1 R and Abs. This current study thus provides extended insights into the molecular action of AT1 R-Abs and associated mechanisms of interrelated pathogenesis.

UR - https://www.scopus.com/pages/publications/85127396007

U2 - 10.3390/ijms23073984

DO - 10.3390/ijms23073984

M3 - Journal articles

C2 - 35409344

AN - SCOPUS:85127396007

SN - 1661-6596

VL - 23

JO - International Journal of Molecular Sciences

JF - International Journal of Molecular Sciences

IS - 7

M1 - 3984

ER -

Molecular Effects of Auto-Antibodies on Angiotensin II Type 1 Receptor Signaling and Cell Proliferation

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