Mechanisms underlying the protective effects of cAMP in cardiac fibrosis

: actin cytoskeletal remodelling and role of MKL1/MKL2 and YAP/TAZ-TEAD dependent transcription

Abstract

Cardiac fibrosis is the main pathophysiological processes contributing to heart failure, affecting
900,000 people in the U.K. Excessive extracellular matrix (ECM) production and increased
proliferation of cardiac fibroblasts contributes towards cardiac fibrosis, which increases
myocardium stiffness, causing systolic/diastolic cardiac dysfunction and ultimately heart failure.
Current treatment options are only partially effective. Hence, a better understanding of the
underlying mechanisms is required to help develop novel therapies. In this thesis, the effects of
cAMP-signalling on cardiac fibroblast proliferation and migration were investigated.
cAMP signalling, via PKA and EPAC1, inhibited cardiac fibroblasts proliferation but not migration.
This was associated with a change in cell morphology, actin remodelling and reduced RhoA-ROCK
signalling. Cyclic-AMP also inhibited SRF and TEAD-dependent transcription. Inhibition of SRF
resulted from reduced nuclear localisation of MKL1, which involved PKA and EPAC1 signalling.
Pharmacological and siRNA-mediated inhibition of SRF and TEAD co-factors MKL1/2 and
YAP/TAZ, respectfully, inhibited cardiac fibroblasts proliferation.
cAMP signalling was associated with a reduction in EPAC1 gene expression, which was mediated
by both PKA and EPAC1 signalling and dependent on inhibition of actin polymerisation. Promoter
analysis identified an EPAC1 promoter TEAD binding-element that was essential for maximal
EPAC1 promoter activity. Inhibition of TEAD activity with siRNA gene silencing, dominant
negative mutants or pharmacological inhibitors repressed EPAC1 expression. Importantly, a
constitutively active YAP mutant rescued EPAC1 promoter activity but not endogenous EPAC1
mRNA levels after cAMP elevation, indicating involvement of additional mechanism. cAMP
reduced histone3-lysine27 acetylation at the EPAC1 proximal promoter, which contributed towards
EPAC1 repression.
Taken together, these data demonstrate an important role of PKA and EPAC induced actin
remodelling and inhibition of MKL-SRF and YAP/TAZ-TEAD in mediating the anti-mitogenic
effects of cAMP in cardiac fibroblasts. The data also describes the existence of a negative feedback
loop that regulates EPAC1 expression in response to cAMP-induced actin and chromatin
remodelling.

Date of Award	29 Sept 2020
Original language	English
Awarding Institution	University of Bristol
Supervisor	Mark Bond (Supervisor), Andrew C Newby (Supervisor) & Saadeh Suleiman (Supervisor)