June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Serine Phosphorylation of IQGAP1 Regulates Choroidal Endothelial Activation and Neovascularization
Author Affiliations & Notes
  • Haibo Wang
    John A Moran Eye Center, The University of Utah, Salt Lake City, Utah, United States
  • Eric Kunz
    John A Moran Eye Center, The University of Utah, Salt Lake City, Utah, United States
  • Aniket Ramshekar
    John A Moran Eye Center, The University of Utah, Salt Lake City, Utah, United States
  • Mary Elizabeth Hartnett
    John A Moran Eye Center, The University of Utah, Salt Lake City, Utah, United States
  • Footnotes
    Commercial Relationships   Haibo Wang, None; Eric Kunz, None; Aniket Ramshekar, None; Mary Elizabeth Hartnett, None
  • Footnotes
    Support  NIH Grants EY014800 (Core Funding); NIH R01EY015130 and R01EY017011 to M.E.H and an Unrestricted Grant from Research to Prevent Blindness, Inc., New York, NY, to the Department of Ophthalmology & Visual Sciences, University of Utah. The Retina Research Foundation of The Macula Society to MEH.; NIH Grant EY021721 to WWH, RPB, Inc. to WWH; P, C, AGTC (WWH).
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 641. doi:
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    • Get Citation

      Haibo Wang, Eric Kunz, Aniket Ramshekar, Mary Elizabeth Hartnett; Serine Phosphorylation of IQGAP1 Regulates Choroidal Endothelial Activation and Neovascularization. Invest. Ophthalmol. Vis. Sci. 2021;62(8):641.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Serine phosphorylation of IQGAP1 regulates signaling leading to neurite cell growth. We previously found that knockout of IQGAP1 inhibited VEGF-mediated choroidal endothelial cell (CEC) activation and migration, a necessary step in the development of choroidal neovascularization (CNV) in neovascular age-related macular degeneration (AMD). We, therefore, tested the hypothesis that IQGAP1 mediates the development of CNV through IQGAP1 serine phosphorylation.

Methods : Serine phosphorylation of IQGAP1 was measured by Co-immunoprecipitation (Co-IP) with anti-IQGAP1 and western blot with anti-phospho-serine antibodies in 1) primary human CECs transfected with plasmid DNA expressing wild type IQGAP1 (IQ-WT) or IQGAP1 with serine mutation into alanine at sites 1441/1443 (IQ-S/A) and treated with control PBS or VEGF (20 ng/ml) for 15 mins, and in 2) retinal pigment epithelium (RPE)/choroid tissues from wild type mice 7 days after laser treatment. Mice with IQGAP1 mutated at serine 1441 (Crispr/IQ) were created by CRISPR-Cas9 edited gene mutation. Seven days post laser, CNV lesion volume was measured in lectin-stained RPE/choroidal flatmounts of Crispr/IQ and littermate controls. IQGAP1 serine phosphorylation was determined in CECs isolated from three different donors and in RPE/choroid tissues from three lasered mice and three non-lasered controls. The volume of CNV lesions were measured in 7 Crispr-IQ mice and 16 littermate wild type controls.

Results : Compared to PBS, VEGF activated serine phosphorylation of IQGAP1 in IQ-WT transfected CECs. VEGF-induced IQGAP1 serine phosphorylation was abolished in CECs transfected with IQ-S/A. In RPE/choroid tissues and compared to non-lasered controls, laser treatment caused a 3-fold increase in IQGAP1 serine phosphorylation. Crispr-IQ mice had a trend toward reduced CNV lesions (p=0.07) compared to littermate wild type mice.

Conclusions : VEGF treatment and laser injury activate IQGAP1 by inducing serine phosphorylation. Serine phosphorylation of IQGAP1 mediates experimental CNV in vivo. These data may provide insight into mechanisms underlying neovascular AMD.

This is a 2021 ARVO Annual Meeting abstract.

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