Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity

Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity

Bibliographic Details
Main Author: Shetty, Shruthi Ratnakar
Language:English
Published: The Ohio State University / OhioLINK 2020
Subjects:
Pharmacology
Biochemistry
Cellular Biology
Cyclin Dependent Kinases
Inflammation
Endothelial Cells
iNOS
COX-2
Online Access:http://rave.ohiolink.edu/etdc/view?acc_num=osu1591802767017206
id ndltd-OhioLink-oai-etd.ohiolink.edu-osu1591802767017206
record_format oai_dc
collection NDLTD
language English
sources NDLTD
topic Pharmacology
Biochemistry
Cellular Biology
Cyclin Dependent Kinases
Inflammation
Endothelial Cells
iNOS
COX-2
spellingShingle Pharmacology
Biochemistry
Cellular Biology
Cyclin Dependent Kinases
Inflammation
Endothelial Cells
iNOS
COX-2
Shetty, Shruthi Ratnakar
Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
author Shetty, Shruthi Ratnakar
author_facet Shetty, Shruthi Ratnakar
author_sort Shetty, Shruthi Ratnakar
title Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
title_short Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
title_full Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
title_fullStr Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
title_full_unstemmed Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity
title_sort cyclin-dependent kinases and their role in inflammation, endothelial cell migration and autocrine activity
publisher The Ohio State University / OhioLINK
publishDate 2020
url http://rave.ohiolink.edu/etdc/view?acc_num=osu1591802767017206
work_keys_str_mv AT shettyshruthiratnakar cyclindependentkinasesandtheirroleininflammationendothelialcellmigrationandautocrineactivity
_version_ 1719457542144262144
spelling ndltd-OhioLink-oai-etd.ohiolink.edu-osu15918027670172062021-08-03T07:15:11Z Cyclin-Dependent Kinases and their role in Inflammation, Endothelial Cell Migration and Autocrine Activity Shetty, Shruthi Ratnakar Pharmacology Biochemistry Cellular Biology Cyclin Dependent Kinases Inflammation Endothelial Cells iNOS COX-2 Inflammation is the body’s response to infection or injury. Endothelial cells are among the different players involved in an inflammatory cascade. In response to an inflammatory stimuli such as bacterial lipopolysaccharide (LPS), endothelial cells get activated which is characterized by the production of important mediators, such as inducible nitric oxide synthase (iNOS) which, catalyzes the production of nitric oxide (NO) and reactive nitrogen species and cyclooxygenase-2 (COX-2) that catalyzes the production of prostaglandins. Though the production of these mediators is required for an inflammatory response, it is important that their levels are regulated. Continued production of iNOS results in increased accumulation of reactive nitrogen species (RNS) that might lead to cytotoxicity, whereas lack of/suppression results in endothelial and vascular dysfunction. On the other hand, severe cardiovascular, intestinal and renal side effects are observed with significant suppression of COX-2. Thus, studying factors that could regulate the levels of iNOS and COX-2 could provide useful insights for developing novel therapeutic targets. Regulation of protein levels involves control of protein induction or turnover. Since protein induction requires transcription, in this dissertation we studied the role of a promoter of transcription “Cyclin-dependent kinase 7 (CDK7)” in iNOS and COX-2 protein induction. We first depleted CDK7 in mouse aortic endothelial cells (MAEC) using siRNA, followed by stimulus with lipopolysaccharide/interferon gamma (LPS/IFN) to induce iNOS and COX-2 protein. Interestingly, CDK7 depletion resulted in excessive induction of both iNOS and COX-2 protein levels compared to control. This observation was recapitulated using a covalent inhibitor of CDK7, THZ1. Further investigation showed the protein induction due to lack of CDK7 was not due to increased mRNA levels. CDK7 knockdown however decreased the turnover of iNOS, but not COX-2 protein. We identified a novel mechanism by which CDK7 could regulate the stability of iNOS protein. We demonstrated how in the absence of CDK7, iNOS was resistant to degradation by protease, calpain, due to increased association of iNOS with its binding partner calmodulin. CDK7 depletion was also found to exert a post-transcriptional effect on COX-2, by activating upstream signaling cascades from mTOR and MAPK pathways that might promote translation. Activation of the components of the translation pre-initiation complex was also observed in the absence of CDK7, which contributed to increased COX-2 protein levels. This dissertation thus identifies CDK7 as a repressor of both iNOS and COX-2 proteins in endothelial cells by affecting their turnover and translation, respectively. The research provides novel insights, for the therapeutic manipulation of iNOS and COX-2 expression in endothelial cells via the less explored post-transcriptional effects of CDK7.We also investigated the roles of different transcriptional and atypical CDKs in angiogenesis and autocrine activity in endothelial cells. CDK5 and CDK9 were found to be potential regulators of endothelial cell migration whereas CDK7 and CDK9 were found to regulate autocrine activity in endothelial cells. Although these studies are highly preliminary, they pave the way to understand and further explore these CDKs as potential angiogenic or autocrine targets in various therapeutic conditions. 2020 English text The Ohio State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=osu1591802767017206 http://rave.ohiolink.edu/etdc/view?acc_num=osu1591802767017206 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.

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