화학공학소재연구정보센터
Biochemical and Biophysical Research Communications, Vol.476, No.4, 379-385, 2016
G-protein-dependency of orexin/hypocretin receptor signalling in recombinant Chinese hamster ovary cells
Multiple signalling pathways for orexin receptors have been discovered, and most thoroughly mapped in Chinese hamster ovary K1 (CHO-K1) cells. It is also known that orexin receptors can couple to the G-protein families G(i), G(s) and G(q). However, the connection between the G-proteins and the downstream signals is only vaguely established, and we now set out to resolve this for human orexin receptors expressed in CHO-Kl cells. Adenylyl cyclase (AC), phospholipase A(2), C and D, and diacylglycerol lipase activities were assessed by precursor radiolabelling and chromatographic separation, and calcium by fluorescent methods. Pertussis toxin, cholera toxin and the cyclic depsipeptide, UBO-QIC a.k.a. FR900359, were used to assess the involvement of G(i)-, G(s)- and G(q)-family G-proteins, respectively. Calcium elevations as well as activation of the phospholipases and diacylglycerol lipase were dependent on G(q), as they were fully blocked by UBO-QIC. The low-potency AC activation fully depended on G(s). Surprisingly, the assumed G(i)-dependent inhibition of AC was (fully or partially) inhibited by UBO-QIC, in opposition to the previous findings of no sensitivity of G(i) proteins to UBO-QIC. Orexin receptor signalling is indeed mostly G(q)-driven in CHO-Kl cells, even with respect to the less clearly mapped cascades such as phospholipase A(2) and C and calcium influx, underlining the importance of G(q) even under physiological conditions. AC regulation warrants more studies. (C) 2016 Elsevier Inc. All rights reserved.