Polycystic kidney disease (PKD) is characterized by cardiovascular irregularities, including hypertension. Dopamine, a circulating hormone, is implicated in essential hypertension in humans and animal models. Vascular endothelial primary cilia are known to function as mechano-sensory organelles. Though both primary cilia and dopamine receptors play important roles in vascular hypertension, their relationship has never been explored. To determine the roles of the dopaminergic system and mechanosensory cilia, we studied the effects of dopamine on ciliary length and function in wild-type (WT) and mechano-insensitive polycystic mutant cells (Pkd1−/− and Tg737orpk/orpk). We show for the first time that mouse vascular endothelia exhibit dopamine receptor-type 5 (DR5), which co-localizes to primary cilia in cultured cells and mouse arteries in vivo. DR5 activation increases cilia length in arteries and endothelial cells through cofilin and actin polymerization. DR5-activation also restores cilia function in the mutant cells. In addition, silencing DR5 completely abolishes mechano-ciliary function in WT cells. We find that DR5 plays very important roles in ciliary length and function. Furthermore, the chemosensory function of cilia can alter the mechanosensory function through changes in sensitivity to fluid-shear stress. We propose that activated ciliary DR5 has a functional mechanosensory role in endothelial cells.

The primary cilium is an important sensory organelle present in most mammalian cells. Our current studies aim at examining intracellular molecules that regulate cilia length and/or cilia function in vitro and ex vivo. For the first time, we show that intracellular cAMP and cAMP-dependent protein kinase (PKA) regulate both cilia length and function in vascular endothelial cells. Although calcium-dependent protein kinase modulates cilia length, it does not play a significant role in cilia function. Cilia length regulation also involves mitogen-activated protein kinase (MAPK), protein phosphatase-1 (PP-1), and cofilin. Furthermore, cofilin regulates cilia length through actin rearrangement. Overall, our study suggests that the molecular interactions between cilia function and length can be independent of one another. Although PKA regulates both cilia length and function, changes in cilia length by MAPK, PP-1, or cofilin do not have a direct correlation to changes in cilia function. We propose that cilia length and function are regulated by distinct, yet complex intertwined signaling pathways.

A disintegrin-like metalloproteinase with thrombospondin motifs-16 (Adamts16) is an important candidate gene for hypertension. The goal of the present study was to further assess the candidacy of Adamts16 by targeted disruption of this gene in a rat genetic model of hypertension. A rat model was generated by manipulating the genome of the Dahl Salt-sensitive (S) rat using zinc-finger nucleases, wherein the mutant rat had a 17 bp deletion in the first exon of Adamts16, introducing a stop codon in the transcript. Systolic blood pressure (BP) of the homozygous Adamts16 mutant rats was lower by 36 mmHg compared with the BP of the S rats. The Adamts16 mutant rats exhibited significantly lower aortic pulse wave velocity and vascular media thickness compared with S rats. Scanning electron and fluorescence microscopic studies indicated that the mechanosensory cilia of vascular endothelial cells from the Adamts16 mutant rats were longer than that of the S rats. Furthermore, Adamts16 mutant rats showed splitting and thickening of glomerular capillaries and had a longer survival rate, compared with the S rats. Taken together, these physiological observations functionally link Adamts16 to BP regulation and suggest the vasculature as the potential site of action of Adamts16 to lower BP.espite strong evidence that susceptibility or resistance to the development of hypertension is heritable, the identification of genetic variants that cause blood pressure (BP) to rise into a hypertensive state has remained difficult (1, 2). Classic genetic mapping and association studies in both humans and in rats point to several genetic elements as potential candidates causing hypertension (3, 4). Most of the prioritized candidate genes for hypertension await functional assessments.Linkage analysis in the Quebec Family Study identified a quantitative trait locus (QTL) for systolic BP on human chromosome 5p15 (5). The corresponding comparative segment of human chromosome 5p15 on rat chromosome 1 is also linked to a BP QTL in rats (6). Improved resolutions of this locus in rats were obtained through iterative substitution mapping using strains differentially susceptible to the development of hypertension (6-10). A disintegrin-like metalloproteinase with thrombospondin motifs-16 (Adamts16), which was the only known gene with exonic variants within the highly resolved congenic interval, was prioritized as a candidate BP quantitative trait gene (QTG) (8). More importantly, following the congenic mapping study in rats, human allelic variants of Adamts16 were confirmed as being associated with BP in two independent cohorts, one of which was the Quebec Family Study (8). Taken together, all these studies point to Adamts16 as a prominent candidate locus linked to BP control across two species. However, due to the limitations of recombination frequencies, both the linkage and substitution mapping studies in rats cannot validate Adamts16 as the BP QTG because of the presence of other candidate variants within the linked or introgressed flanking genomic ...

In this review, we will discuss several well-accepted signaling pathways toward calcium-mediated mechanisms of cystic expansion. The second messenger calcium ion has contributed to a vast diversity of signal transduction pathways. We will dissect calcium signaling as a possible mechanism that contributes to renal cyst formation. Because cytosolic calcium also regulates an array of signaling pathways, we will first discuss cilia-induced calcium fluxes, followed by Wnt signaling that has attributed to much-discussed planar cell polarity. We will then look at the relationship between cytosolic calcium and cAMP as one of the most important aspects of cyst progression. The signaling of cAMP on MAPK and mTOR will also be discussed. We infer that while cilia-induced calcium fluxes may be the initial signaling messenger for various cellular pathways, no single signaling mediator or pathway is implicated exclusively in the progression of the cystic expansion.

A Disintegrin And Metalloproteinase with ThromboSpondin motifs16 (ADAMTS-16) is a member of a family of metalloproteinases. Using a novel zinc-finger nuclease based gene-edited rat model harboring a targeted mutation of the Adamts16 locus, we previously reported this gene to be linked to blood pressure regulation. Here we document our observation with this model that Adamts16 is essential for normal development of the testis. Absence of Adamts16 in the homozygous Adamts16mutant males resulted in cryptorchidism and male sterility. Heterozygous Adamts16mutant males were normal, indicating that this is a recessive trait. Testes of homozygous Adamts16mutant males were significantly smaller with significant histological changes associated with the lack of sperm production. Temporal histological assessments of the testis demonstrated that the seminiferous tubules did not support active spermatogenesis, but progressively lost germ cells, accumulated vacuoles and did not have any sperm. These observations, taken together with our previous report of renal abnormalities observed with the same Adamts16mutant rats, suggest an important mechanistic link between Adamts16 and the functioning of the male genitourinary system.