UNDERSTANDING HOW MAP KINASES INFLUENCE ENDOTHELIAL NITRIC-OXIDE SYNTHASE ACTIVITY

<p>Mitogen activated protein kinases (MAPK) p38 and ERK have both been reported to bind endothelial nitric oxide synthase (eNOS) with submicromolar affinity via proposed interactions with a pentabasic non-canonical MAPK binding sequence in the autoinhibitory insertion of eNOS. The neuronal isoform, which lacks the pentabasic motif, did not bind either MAPK significantly. In the present study, the pentabasic motif was validated using predictive modeling programming, and eNOS phosphorylation by MAPKs (P38, ERK and JNK) was examined using <em>in vitro</em> kinase assays and immunoblotting. JNK phosphorylation at Ser¹¹⁴ contrasts with ERK, which phosphorylated Ser⁶⁰⁰, and p38, which phosphorylated both sites. Literature reports showed that JNK1/2 activation does not alter eNOS activity and prior work in our lab showed that ERK phosphorylation negativity affects NO output. This thesis presents <em>in vitro </em>results comparing the direct impact of each kinase on eNOS activity using the oxyhemoglobin (Oxy-Hb) assay. In HMEC-1 cells and <em>in vitro </em>it is observe that phosphorylation at these MAPK sites happens concurrently with activating sites (Ser¹¹⁷⁷), creating dually phosphorylated eNOS. These results underscore the importance of MAPK interactions with eNOS showing that each MAPK creates a unique phosphorylation pattern and NO production outcome. These findings strengthen the emerging paradigm of eNOS as a junction of multiple signaling pathways.</p>

Name thesis:degree_name

Master of Science in Integrative Biology (MSIB)

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Biology

Year dc:date.available

2019

Author dc:creator

• Solone, Xzaviar

Contributors dc:contributor

• Dr. Susan Smith
• Jonathan McMurry
• Dr. Scott Nowak

• eNOS
• MAP Kinases
• nitric oxide synthase
• phosphorylation
• nitric-oxide
• Analytical Chemistry
• Biochemistry
• Biology
• Integrative Biology
• Molecular Biology
• Structural Biology

Repository record dc:identifier

https://digitalcommons.kennesaw.edu/integrbiol_etd/40

OAI identifier oai:identifier

oai:digitalcommons.kennesaw.edu:integrbiol_etd-1040