PKCδ Mediates Mineralocorticoid Receptor Activation by Angiotensin II to Modulate Smooth Muscle Cell Function

Abstract

Angiotensin II (AngII) and the mineralocorticoid receptor (MR) ligand aldosterone both contribute to cardiovascular disorders, including hypertension and adverse vascular remodeling. We previously demonstrated that AngII activates MR-mediated gene transcription in human vascular smooth muscle cells (SMCs), yet the mechanism and the impact on SMC function are unknown. Using an MR-responsive element-driven transcriptional reporter assay, we confirm that AngII induces MR transcriptional activity in vascular SMCs and endothelial cells, but not in Cos1 or human embryonic kidney-293 cells. AngII activation of MR was blocked by the MR antagonist spironolactone or eplerenone and the protein kinase C-δ (PKCδ) inhibitor rottlerin, implicating both in the mechanism. Similarly, small interfering RNA knockdown of PKCδ in SMCs prevented AngII-mediated MR activation, whereas knocking down of MR blocked both aldosterone- and AngII-induced MR function. Coimmunoprecipitation studies reveal that endogenous MR and PKCδ form a complex in SMCs that is enhanced by AngII treatment in association with increased serine phosphorylation of the MR N terminus. AngII increased mRNA expression of the SMC-MR target gene, FKBP51, via an MR-responsive element in intron 5 of the FKBP51 gene. The impact of AngII on FKBP51 reporter activity and gene expression in SMCs was inhibited by spironolactone and rottlerin. Finally, the AngII-induced increase in SMC number was also blocked by the MR antagonist spironolactone and the PKCδ inhibitor rottlerin. These data demonstrate that AngII activates MR transcriptional regulatory activity, target gene regulation, and SMC proliferation in a PKCδ-dependent manner. This new mechanism may contribute to synergy between MR and AngII in driving SMC dysfunction and to the cardiovascular benefits of MR and AngII receptor blockade in humans.

Wnt5a/Ror2 promotes vascular smooth muscle cells proliferation via activating PKC

“…Wnt5a is involved in the development of various diseases such as atherosclerosis, cancer, and inflammation [24][25][26] and plays a critical role in numerous cellular processes from embryonic morphogenesis to postnatal development [5]. Our previous study has illustrated that the combination of Wnt5a and Ror2 together suppresses the expression of adenosine triphosphate-binding cassette transporter A1 (ABCA1) promotes cholesterol accumulation and secretion of pro-inflammatory cytokines and nuclear translocation of NF-κB in VSMCs, and ultimately leads to atherosclerosis [16]. OxLDL increased Wnt5a expression, induced foam cell formation, and affected the migration and phenotype of VSMCs [27].…”

“…The Protein kinase C (PKC) family of serine/threonine kinases is evolutionarily conserved and expressed in a wide variety of species [15]. Emerging evidence has shown that PKC is involved in the proliferation of VSMCs [16]. Interestingly, Ror2 activation by a noncanonical Wnt ligand can activate PKC and c-Jun N-terminal kinases (JNK) and promote N-methyl-d-aspartic acid (NMDA) receptor synaptic responses [17].…”

Mineralocorticoid Receptor in Myeloid Cells Mediates Angiotensin II-Induced Vascular Dysfunction in Female Mice

“…In addition, it is also important to highlight that macrophages do not express 11β-hydroxysteroid dehydrogenase type 2, the enzyme that inactivates glucocorticoids and provides specificity to mineralocorticoid actions. Therefore, MR activation in macrophages is likely to be mediated instead by glucocorticoids ( Rickard et al, 2009 ; Usher et al, 2010 ), although direct MR activation by Ang II has been described ( Lu et al, 2019 ). Despite these limitations of the MyMRKO model, our findings support a critical role for myeloid cells, in particular macrophages, as potential mediators of vascular injury resulting from inappropriate MR activation.…”

Posttranslational Modifications of the Mineralocorticoid Receptor and Cardiovascular Aging

“…For PKCδ, angiotensin II led to MR-PKCδ complex formation in association with increased serine phosphorylation of the MR-NTD in vascular cells. AngII also activated MR transcriptional activity, target gene expression, and SMC proliferation in a PKCδ-dependent manner that may involve phosphorylation of MR-NTD (Lu et al, 2019). Although MR phosphorylation by PKC isoforms has been demonstrated by several groups, it is not always clear if the effects described evolve from a direct MR phosphorylation or indirectly through phosphorylation of co-regulators or a combination of both.…”