Nitish R Mahapatra

Catestatin is a 21-amino acid residue, cationic and hydrophobic peptide that is formed endogenously by proteolytic cleavage of its precursor chromogranin A, a major protein co-stored and co-released with catecholamines from the storage vesicles in adrenal chromaffin cells and adrenergic neurons. This peptide exhibits potent catecholamine release-inhibitory activity by acting on the neuronal nicotinic acetylcholine receptor. It also stimulates histamine release from mast cells via heterotrimeric G-proteins in a receptor-independent manner. Plasma levels of catestatin are diminished not only in hypertensive patients but also in their still-normotensive offspring, indicating its role in the pathogenesis of hypertension. Consistently, exogenous catestatin rescues hypertension in chromogranin A knockout mice and diminishes blood pressure responses to activation of sympathetic outflow in rats. These hypotensive actions of catestatin may be caused directly by autocrine inhibition of catecholamine release from the sympathoadrenal system and indirectly by paracrine stimulation of the potent vasodilator histamine release from mast cells. Recently, three human variants of catestatin displaying differential potencies for inhibition of catecholamine secretion have been identified. One of these variants (Gly364Ser) causes increased baroreceptor sensitivity, increased cardiac parasympathetic activity, and decreased cardiac sympathetic activity, and it seems to alter the risk for hypertension. These cardiovascular effects may have resulted by action of this peptide in the baroreceptor centre of the nucleus tractus solitarius. Thus, accumulating evidence documents the endogenous peptide catestatin as a novel regulator of cardiac function and blood pressure. © The Author 2008.

Secretion of proteins and peptides from eukaryotic cells takes place by both constitutive and regulated pathways. Regulated secretion may involve interplay of proteins that are currently unknown. Recent studies suggest an important role of chromogranin A (CHGA) in the regulated secretory pathway in neuroendocrine cells, but the mechanism by which CHGA enters the regulated pathway, or even triggers the formation of the pathway, remains unclear. In this study, we used a transcriptome/proteome-wide approach, to discover binding partners for CHGA, by employing a phage display cDNA library method. Several proteins within or adjacent to the secretory pathway were initially detected as binding partners of recombinant human CHGA. We then focused on the trans-Golgi protein SCLIP (STMN3) and its stathmin paralog SCG10 (STMN2) for functional study. Co-immunoprecipitation experiments confirmed the interaction of each of these two proteins with CHGA in vitro. SCLIP and SCG10 were colocalized to the Golgi apparatus of chromaffin cells in vivo and shared localization with CHGA as it transited the Golgi. Downregulation of either SCLIP or SCG10 by synthetic siRNAs virtually abolished chromaffin cell secretion of a transfected CHGA-EAP chimera (expressing CHGA fused to an enzymatic reporter, and trafficked to the regulated pathway). SCLIP siRNA also decreased the level of secretion of endogenous CHGA and SCG2, as well as transfected human growth hormone, while SCG10 siRNA decreased the level of regulated secretion of endogenous CHGB. Moreover, a dominant negative mutant of SCG10 (Cys22,Cys 24→Ala22,Ala24) significantly blocked secretion of the transfected CHGA-EAP chimera. A decrease in the buoyant density of chromaffin granules was observed after downregulation of SCG10 by siRNA, suggesting participation of these stathmins in granule formation or maturation. We conclude that SCLIP and SCG10 interact with CHGA, share partial colocalization in the Golgi apparatus, and may be necessary for typical transmitter storage and release from chromaffin cells. © 2008 American Chemical Society.

Renal organic anion transporters (OAT) are known to mediate the excretion of many drugs, but their function in normal physiology is not well understood. In this study, mice lacking organic anion transporter 3 (Oat3) had a 10 to 15% lower BP than wild-type mice, raising the possibility that Oat3 transports an endogenous regulator of BP. The aldosterone response to a low-salt diet was blunted in Oat3-null mice, but baseline aldosterone concentration was higher in these mice, suggesting that aldosterone dysregulation does not fully explain the lower BP in the basal state; therefore, both targeted and global metabolomic analyses of plasma and urine were performed, and several potential endogenous substrates of Oat3 were found to accumulate in the plasma of Oat3-null mice. One of these substrates, thymidine, was transported by Oat3 expressed in vitro. In vivo, thymidine, as well as two of the most potent Oat3 inhibitors that were characterized, reduced BP by 10 to 15%; therefore, Oat3 seems to regulate BP, and Oat3 inhibitors might be therapeutically useful antihypertensive agents. Moreover, polymorphisms in human OAT3 might contribute to the genetic variation in susceptibility to hypertension. Copyright © 2008 by the American Society of Nephrology.

Chromogranin A (CHGA) is stored and released from the same secretory vesicles that contain catecholamines in chromaffin cells and noradrenergic neurons. We had previously identified common genetic variants at the CHGA locus in several human populations. Here we focus on whether inter-individual variants in the promoter region are of physiological significance. A common haplotype, CGATA (Hap-B), blunted the blood pressure response to cold stress and the effect exhibited molecular heterosis with the greatest blood pressure change found in Hap-A/Hap-B heterozygotes. Homozygosity for three minor alleles with peak effects within the haplotype predicted lower stress-induced blood pressure changes. The G-462A variant predicted resting blood pressure in the population with higher pressures occurring in heterozygotes (heterosis). Using cells transfected with CHGA promoter-luciferase reporter constructs, the Hap-B haplotype had decreased luciferase expression compared to the TTGTC (Hap-A) haplotype under both basal conditions and after activation by pre-ganglionic stimuli. The G-462A variant altered a COUP-TF transcriptional control motif. The two alleles in transfected promoters differed in basal activity and in the responses to COUP-II-TF transactivation and to retinoic acid. In vitro findings of molecular heterosis were also noted with the transfected CHGA promoter wherein the diploid combination of the two G-462A alleles gave rise to higher luciferase expression than either allele in isolation. Our results suggest that common genetic variants in the CHGA promoter may regulate heritable changes in blood pressure. © 2008 International Society of Nephrology.