Physical interaction and functional coupling between ACDP4 and the intracellular ion chaperone COX11, an implication of the role of ACDP4 in essential metal ion transport and homeostasis

Divalent metal ions such as copper, manganese, and cobalt are essential for cell development, differentiation, function and survival. These essential metal ions are delivered into intracellular domains as cofactors for enzymes involved in neuropeptide and neurotransmitter synthesis, superoxide metabolism, and other biological functions in a target specific fashion. Altering the homeostasis of these essential metal ions is known to connect to a number of human diseases including Alzheimer disease, amyotrophic lateral sclerosis, and pain. It remains unclear how these essential metal ions are delivered to intracellular targets in mammalian cells. Here we report that rat spinal cord dorsal horn neurons express ACDP4, a member of Ancient Conserved Domain Protein family. By screening a pretransformed human fetal brain cDNA library in a yeast two-hybrid system, we have identified that ACDP4 specifically interacts with COX11, an intracellular metal ion chaperone. Ectopic expression of ACDP4 in HEK293 cells resulted in enhanced toxicity to metal ions including copper, manganese, and cobalt. The metal ion toxicity became more pronounced when ACDP4 and COX11 were co-expressed ectopically in HEK293 cells, suggesting a functional coupling between them. Our results indicate a role of ACDP4 in metal ion homeostasis and toxicity. This is the first report revealing a functional aspect of this ancient conserved domain protein family. We propose that ACDP is a family of transporter protein or chaperone proteins for delivering essential metal ions in different mammalian tissues. The expression of ACDP4 on spinal cord dorsal horn neurons may have implications in sensory neuron functions under physiological and pathological conditions

Contribution of cytochrome P450 4A isoforms to renal functional response to inhibition of nitric oxide production in the rat

20-Hydroxyeicosatetraenoic acid (20-HETE), a major renal eicosanoid, regulates renal function and contributes to renal responses following withdrawal of nitric oxide (NO). However, the role of 20-HETE-synthesizing isoforms in renal function resulting from NO inhibition is unknown. The present study evaluated the role of cytochrome (CYP)4A1, −4A2 and −4A3 isoforms on renal function in the presence and absence of NO. Antisense oligonucleotides (ASODN) to CYP4A1, −4A2 and −4A3 reduced 20-HETE synthesis and downregulated the expression of CYP4A isoforms in renal microsomes. Nω-L-nitromethyl arginine ester (L-NAME, 25 mg kg−1), an inhibitor of NO production, increased mean arterial blood pressure (MABP, Δ=+18 to 26 mmHg), reduced renal blood flow (RBF, Δ= −1.8 to 2.9 ml min−1), increased renal vascular resistance (RVR, Δ=+47 to 54 mmHg ml−1 min−1), reduced glomerular filtration rate (GFR), but increased sodium excretion (UNaV). ASODN to CYP4A1 and −4A2 but not −4A3 reduced basal MABP and RVR and increased basal GFR, while ASODN to CYP4A2 significantly reduced basal UNaV suggesting a differential role for CYP4A isoforms in the regulation of renal function. ASODN to CYP4A2 but not −4A1 or −4A3 blunted the increase in MABP by L-NAME (38 ± 9 %, P < 0.05). ASODN to CYP4A1, −4A2 and −4A3 attenuated the reduction in RBF and the consequent increase in RVR by L-NAME with a potency order of CYP4A2 = CYP4A1 > CYP4A3. ASODN to CYP4A1 and −4A2 but not −4A3 attenuated L-NAME-induced reduction in GFR, but ASODN to all three CYP4A isoforms blunted the L-NAME-induced increase in UNaV (CYP4A3 > CYP4A1 >> CYP4A2). We conclude from these data that CYP4A isoforms contribute to different extents to basal renal function. Moreover, CYP4A2 contributes greatest to haemodynamic responses while CYP4A3 contributes greatest to tubular responses following NO inhibition. We therefore propose that NO differentially regulates the function of CYP4A1, −4A2, and −4A3 isoforms in the renal vasculature and the nephron

mPGES-1 deletion impairs diuretic response to acute water loading

PGE2 has an established role in renal water handling. The present study was undertaken to examine the role of microsomal prostaglandin E synthase-1 (mPGES-1) in the diuretic response to acute and chronic water loading. Compared with wild-type (+/+) controls, mPGES-1 −/− mice exhibited impaired ability to excrete an acute, but not chronic water load. In response to acute water loading, urinary PGE2 excretion in the +/+ mice increased at 2 h, in parallel with increased urine flow. In contrast, the −/− mice exhibited a delayed increase in urinary PGE2 excretion, coinciding with the stimulation of renal medullary mRNA expression of cytosolic prostaglandin E synthase but not mPGES-2. At baseline, renal aquaporin-2 (AQP2) expression in mPGES-1 −/− mice was enhanced compared with the +/+ control. In response to acute water loading, renal AQP2 expression in the +/+ mice was significantly reduced, and this reduction was blunted in the −/− mice. Despite striking changes in AQP2 protein expression, renal AQP2 mRNA in both genotypes largely remained unchanged. Overall, these data support an important role of mPGES-1 in provoking the diuretic response to acute water loading