Ultraviolet C Irradiation Induces Different Expression of Cyclooxygenase 2 in NIH 3T3 Cells and A431 Cells: The Roles of COX-2 Are Different in Various Cell Lines

Ultraviolet C (UVC) is a DNA damage inducer, and 20 J/m(2) of UVC irradiation caused cell growth inhibition and induced cell death after exposure for 24-36 h. The growth of NIH 3T3 cells was significantly suppressed at 24 h after UVC irradiation whereas the proliferation of A431 cells was inhibited until 36 h after UVC irradiation. UVC irradiation increased COX-2 expression and such up-regulation reached a maximum during 3-6 h in NIH 3T3 cells. In contrast, UVC-induced COX-2 reached a maximum after 24-36 h in A431 cells. Measuring prostaglandin E2 (PGE2) level showed a biphasic profile that PGE2 release was rapidly elevated in 1-12 h after UVC irradiation and increased again at 24 h in both cell lines. Treatment with the selective COX-2 inhibitor, SC-791, during maximum expression of COX-2 induction, attenuated the UVC induced-growth inhibition in NIH 3T3 cells. In contrast, SC-791 treatment after UVC irradiation enhanced death of A431 cells. These data showed that the patterns of UVC-induced PGE2 secretion from NIH 3T3 cells and A431 cells were similar despite the differential profile in UVC-induced COX-2 up-regulation. Besides, COX-2 might play different roles in cellular response to UVC irradiation in various cell lines.     

Role of Conserved Residues and F322 in the Extracellular Vestibule of the Rat P2X7 Receptor in Its Expression,Function and Dye Uptake Ability

Activation of the P2X7 receptor results in the opening of a large pore that plays a role in immune responses, apoptosis, and many other physiological and pathological processes. Here, we investigated the role of conserved and unique residues in the extracellular vestibule connecting the agonist-binding domain with the transmembrane domain of rat P2X7 receptor. We found that all residues that are conserved among the P2X receptor subtypes respond to alanine mutagenesis with an inhibition (Y51, Q52, and G323) or a significant decrease (K49, G326, K327, and F328) of 2′,3′-O-(benzoyl-4-benzoyl)-ATP (BzATP)-induced current and permeability to ethidium bromide, while the nonconserved residue (F322), which is also present in P2X4 receptor, responds with a 10-fold higher sensitivity to BzATP, much slower deactivation kinetics, and a higher propensity to form the large dye-permeable pore. We examined the membrane expression of conserved mutants and found that Y51, Q52, G323, and F328 play a role in the trafficking of the receptor to the plasma membrane, while K49 controls receptor responsiveness to agonists. Finally, we studied the importance of the physicochemical properties of these residues and observed that the K49R, F322Y, F322W, and F322L mutants significantly reversed the receptor function, indicating that positively charged and large hydrophobic residues are important at positions 49 and 322, respectively. These results show that clusters of conserved residues above the transmembrane domain 1 (K49–Y51–Q52) and transmembrane domain 2 (G326–K327–F328) are important for receptor structure, membrane expression, and channel gating and that the nonconserved residue (F322) at the top of the extracellular vestibule is involved in hydrophobic inter-subunit interaction which stabilizes the closed state of the P2X7 receptor channel.