Endothelial senescence is induced by phosphorylation and nuclear export of telomeric repeat binding factor 2–interacting protein
Sivareddy Kotla, Hang Thi Vu, Kyung Ae Ko, Yin Wang, Masaki Imanishi, Kyung-Sun Heo, Yuka Fujii, Tamlyn N. Thomas, Young Jin Gi, Hira Mazhar, Jesus Paez-Mayorga, Ji-Hyun Shin, Yunting Tao, Carolyn J. Giancursio, Jan L.M. Medina, Jack Taunton, Aldos J. Lusis, John P. Cooke, Keigi Fujiwara, Nhat-Tu Le, Jun-ichi Abe
Sivareddy Kotla, Hang Thi Vu, Kyung Ae Ko, Yin Wang, Masaki Imanishi, Kyung-Sun Heo, Yuka Fujii, Tamlyn N. Thomas, Young Jin Gi, Hira Mazhar, Jesus Paez-Mayorga, Ji-Hyun Shin, Yunting Tao, Carolyn J. Giancursio, Jan L.M. Medina, Jack Taunton, Aldos J. Lusis, John P. Cooke, Keigi Fujiwara, Nhat-Tu Le, Jun-ichi Abe
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Research Article Cardiology Vascular biology

Endothelial senescence is induced by phosphorylation and nuclear export of telomeric repeat binding factor 2–interacting protein

Abstract

The interplay among signaling events for endothelial cell (EC) senescence, apoptosis, and activation and how these pathological conditions promote atherosclerosis in the area exposed to disturbed flow (d-flow) in concert remain unclear. The aim of this study was to determine whether telomeric repeat-binding factor 2–interacting protein (TERF2IP), a member of the shelterin complex at the telomere, can regulate EC senescence, apoptosis, and activation simultaneously, and if so, by what molecular mechanisms. We found that d-flow induced p90RSK and TERF2IP interaction in a p90RSK kinase activity–dependent manner. An in vitro kinase assay revealed that p90RSK directly phosphorylated TERF2IP at the serine 205 (S205) residue, and d-flow increased TERF2IP S205 phosphorylation as well as EC senescence, apoptosis, and activation by activating p90RSK. TERF2IP phosphorylation was crucial for nuclear export of the TERF2IP-TRF2 complex, which led to EC activation by cytosolic TERF2IP-mediated NF-κB activation and also to senescence and apoptosis of ECs by depleting TRF2 from the nucleus. Lastly, using EC-specific TERF2IP-knockout (TERF2IP-KO) mice, we found that the depletion of TERF2IP inhibited d-flow–induced EC senescence, apoptosis, and activation, as well as atherosclerotic plaque formation. These findings demonstrate that TERF2IP is an important molecular switch that simultaneously accelerates EC senescence, apoptosis, and activation by S205 phosphorylation.

Authors

Sivareddy Kotla, Hang Thi Vu, Kyung Ae Ko, Yin Wang, Masaki Imanishi, Kyung-Sun Heo, Yuka Fujii, Tamlyn N. Thomas, Young Jin Gi, Hira Mazhar, Jesus Paez-Mayorga, Ji-Hyun Shin, Yunting Tao, Carolyn J. Giancursio, Jan L.M. Medina, Jack Taunton, Aldos J. Lusis, John P. Cooke, Keigi Fujiwara, Nhat-Tu Le, Jun-ichi Abe

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Figure 5

TERF2IP S205 phosphorylation is crucial for d-flow–induced nuclear export of TERF2IP.

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TERF2IP S205 phosphorylation is crucial for d-flow–induced nuclear expor...
(A) HUVECs were exposed to d-flow for 4 hours and stained with anti-TERF2IP and DAPI. Single confocal optical sections roughly 5 μm above the substrate plane were captured using an Olympus confocal microscope (FV1200 MPE) with a 60× objective lens (NA 1.35na Oil). Faint but detectable anti-TERF2IP staining in the cytoplasm can be detected in cells exposed to d-flow. Scale bars: 20 μm. The percentages of cells with cytoplasmic staining were calculated and presented in B. More than 200 cells from 3 independent experiments were scored for each group. Data are presented as mean ± SD **P < 0.01 by unpaired 2-tailed t test. Scale bars: 20 μm. (C) HUVECs were transduced with adenovirus containing Flag-TERF2IP (Ad-Flag-TERF2IP WT) or Flag-TERF2IP S205A and stimulated with or without d-flow for 3 hours. After fixation, cells were then stained by anti-Flag and DAPI and imaged as in A. Note that a portion of exogenously expressed WT TERF2IP, which was localized in the nucleus in static cells, translocated into the cytoplasm only when cells were exposed to d-flow. This d-flow–dependent TERF2IP extranuclearization was not observed in cells expressing the S205A mutant. The percentages of cells with cytoplasmic staining were quantified by counting >200 cells/group from 3 independent experiments and are shown in D. Data are presented as mean ± SD **P < 0.01 by 1-way ANOVA followed by Bonferroni post hoc test.