Export of newly synthesised hK_2P3.1 (TASK1) and hK_2P9.1 (TASK3) channels from the endoplasmic reticulum (ER) and delivery to the cell surface is subject to quality control mechanisms. ER retrieval motifs that bind βCOP have been demonstrated on both the N- and C- termini of both channels (O’Kelly et al., 2002; O’Kelly & Goldstein, 2008; Zuzarte et al., 2009) while a C-terminal motif (MKRRSSV.) recruits 14-3-3 which overcomes βCOP binding and enables forward transport of the channels to the membrane (O’Kelly et al., 2002; Rajan et al., 2002; O’Kelly & Goldstein, 2008). 14-3-3 binding to this motif is phosphorylation dependent. Previous studies have shown that while there are two potential phosphorylation sites within this motif, 14-3-3 binding is dependent on phosphorylation of the terminal serine (S393 for hK_2P3.1 or S373 for hK_2P9.1; O’Kelly et al., 2002; Rajan et al., 2002). To date the kinase responsible for this phosphorylation is unidentified. This study presents evidence that phosphorylation of the terminal serine by protein kinase A (PKA) is required for K_2P3.1 and K_2P9.1 functional expression. In vitro kinase assays demonstrate that alanine substitution of S393 (hK_2P3.1) resulted in a significant reduction in phosphorylation by PKA. When expressed in Xenopus oocytes, mutant channels in which the consensus PKA site was ablated (S393A for K_2P3.1 or S373A for K_2P9.1) showed a marked reduction in current (4.0 ± 0.37 µA and 0.59 ± 0.16 µA at 60mV for WT hK_2P3.1 and S373A respectively and 1.35 ± 0.31 µA and 0.54 ± 0.11 µA at 60mV WT K_2P9.1 and S373A respectively). Oocytes injected with the protein kinase A peptide inhibitor (PKI; 1 nM) 24 hours post WT hK_2P3.1 RNA injection and incubated for a further 16 hours failed to produce currents significantly different from that of water injected oocytes (0.23 ± 0.05 µA and 0.5 ± 0.21 µA for WT hK_2P3.1 plus PKI injected and water injected cells respectively at 60 mV). Cells injected with RNA but not PKI gave acid sensitive currents similar to those seen previously. Similarly hK_2P9.1 injected oocytes treated with PKI also resulted in loss of WT currents (0.3 ± 0.07 µA and 1.36 ± 0.26 µA for WT with and without PKI injection respectively at 60 mV). This data demonstrates that PKA phosphorylation of hK_2P3.1 or hK_2P9.1 on their terminal serine is required for functional expression of these channels.