ABCMdb

ABCC7 p.Ser737Ala

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Publications

PMID: 11053017 [PubMed] Baldursson O et al: "Contribution of R domain phosphoserines to the function of CFTR studied in Fischer rat thyroid epithelia."

No. Sentence Comment

14 Changing Ser737 to alanine increased current above wild-type levels, suggesting that phosphorylation of Ser737 may inhibit current in wild-type CFTR. Login to comment
56 In each of the serine to alanine mutants, S660A, S737A, S795A, and S813A, alanine replaced serine at the designated residue. Login to comment
108 The S795A channels generated more current, but it was still less than that produced by wild-type CFTR. Interestingly, current from the S737A variant tended to be greater than that of wild-type CFTR, although the difference was not significant. Login to comment
114 Current from the S795A variant was not different from that in wild-type CFTR. Interestingly, current generated by the S737A variant was more than twice that generated by wild-type CFTR. Login to comment
134 The S737A variant generated more current than the wild type, and the S-Quad-A mutant generated less. Login to comment
137 The data show that the S-Quad-A variant was less sensitive to forskolin than the wild type and that the S737A variant was more sensitive than wild-type CFTR. Login to comment
140 The results show that current increased more rapidly with S737A than with wild-type and S-Quad-A CFTR. Login to comment
152 The increase in current and the more rapid activation observed when Ser737 was mutated to alanine suggested that phosphorylation of Ser737 may inhibit current. Login to comment
157 Activation by increasing concentrations of forskolin of Cl- current in epithelia expressing wild-type CFTR, S737A, and SQuad-A. Login to comment
161 C: current in epithelia expressing wild-type CFTR, S737A, and S-Quad-A in response to forskolin. Login to comment
187 Time course of Cl- current activation in epithelia expressing wild-type CFTR, S737A, and S-Quad-A. Login to comment
225 A particularly striking example is the different function of S737A in cells and in excised membrane patches. Login to comment
226 In FRT cells and in Xenopus oocytes (28), Ser737 generated more current than wild-type CFTR, but, in patches, S737A showed the same open state probability as the wild type, and the sensitivity to ATP was reduced (29). Login to comment

PMID: 11244086 [PubMed] Ostedgaard LS et al: "Regulation of the cystic fibrosis transmembrane conductance regulator Cl- channel by its R domain."

No. Sentence Comment

35 In excised, cell-free patches, the S737A mutant reduced Po (20), but in Xenopus oocytes and epithelia, the S737A mutant increased cAMP-stimulated current (21, 23). Login to comment

PMID: 15155835 [PubMed] Ai T et al: "Capsaicin potentiates wild-type and mutant cystic fibrosis transmembrane conductance regulator chloride-channel currents."

No. Sentence Comment

142 We further explored the action of capsaicin using CFTR mutants whose eight major PKA consensus serines are substituted with alanine (S660A, S686A, S700A, S712A, S737A, S768A, S795A, and S813A), so called S-oct-A or 8SA. Login to comment

PMID: 15657296 [PubMed] Csanady L et al: "Preferential phosphorylation of R-domain Serine 768 dampens activation of CFTR channels by PKA."

No. Sentence Comment

208 In addition, to verify the inferred link between the major mobility shift of the R domain and phosphorylation of Ser 737 (Fig. 2), the other candidate inhibitory serine (Wilkinson et al., 1997), we studied phosphorylation of His-tagged R-domain proteins containing the single mutation S737A, or the double mutation S737A-S768A. Login to comment
209 The incremental mobility shifts already seen in Figs. 1 and 2 as phosphorylation of the R domain progressed were recapitulated in the His-tagged WT peptide (Fig. 8 A), and were not substantially altered by the S768A mutation in either the WT or S737A background (Fig. 8, B and D vs. A and C). Login to comment
210 As anticipated, however, the S737A mutation abolished the large mobility shift of both WT and S768A R-domain peptides (Fig. 8, C and D vs. A and B). Login to comment
217 WT (A) and mutant R-domain peptides, with Ser768 replaced by alanine (S768A; B), Ser737 replaced by alanine (S737A; C), or Ser737 and Ser768 both replaced by alanine (S737A-S768A; D), were phosphorylated with 10 nM PKA and 5 ␮M ␥32P-MgATP for 0.5-60 min as indicated. Login to comment
219 The major mobility shift (to band 3; Figs. 1 and 2) was seen in the WT and S768A peptides, but not in the S737A or S737A-S768A peptides. Login to comment
296 Mass spectrometry and site-directed mutagenesis revealed that the major mobility shift was linked to phosphorylation of Ser 737 both at low and high [MgATP] (Fig. 2), and this requirement was confirmed by the absence of the large mobility shift after mutation of Ser 737 to Ala in either WT or S768A R-domain peptide (Fig. 8, A-D; see also Borchardt et al., 1996; Kole et al., 1998). Login to comment
299 Though we did not examine its consequences for channel gating, the S737A mutation has been reported to leave burst duration unchanged from that of WT CFTR (Winter and Welsh, 1997), whereas we found the S768A mutation to roughly double burst duration (Fig. 6). Login to comment

PMID: 17700961 [PubMed] Quinton PM et al: "Too much salt, too little soda: cystic fibrosis."

No. Sentence Comment

78 Substitutionofalanine for phosphorylatable S737A or S768A enhanced the channel activity, suggesting that phosphorylation at either of these sites may inhibit phosphorylation of other stimulatory sites[72] . Login to comment

PMID: 19095655 [PubMed] Kongsuphol P et al: "Mechanistic insight into control of CFTR by AMPK."

No. Sentence Comment

43 EXPERIMENTAL PROCEDURES cRNAs for CFTR and Double Electrode Voltage Clamp-Oocytes were injected with cRNA (10 ng, 47 nl of double-distilled water) encoding wtCFTR, L1430A/L1431A, S573A, S1248A, F508del-CFTR, G551D-CFTR, S768A, S737A, S768D, S737D, E1474X, and AMPK␣1. Login to comment
93 Fig. 2B demonstrates that AMPK indeed phosphorylates the R domain in vitro and this AMPK phosphorylation is largely reduced in R domain mutants S737A and S768A (compare lanes 2 and 3 in Fig. 2B, lower panel). Login to comment
94 The data also suggest that serine 768 has a much greater reductive impact because S768A almost abolished all the phosphorylation, whereas some were preserved with S737A. Login to comment
96 We quantified data (supplemental Fig. S2) from three independent experiments and found that the S737A mutation leads to a small 23 Ϯ 8% reduction in counts, whereas S768A leads to a major 81 Ϯ 5% (mean Ϯ range) reduction similar to the double mutant (87% Ϯ 4%) when compared with the wild type (100%). Login to comment
125 It is clear that S768A (lower left panel) has a more profound inhibitory effect on R domain phosphorylation compared with S737A (upper right) given that all these experiments were run simultaneously with similar concentrations of R domain protein and kinase, and each imaged for identical lengths of time. Login to comment
126 Broadly, prolonged incubation that previously created two major spots found with AMPK alone was now supplemented by multiple small spots that almost disappeared after S768A mutation, but much less so after S737A mutation (Fig. 2D, compare lower two panels with upper). Login to comment
128 When expressed in oocytes, CFTR bearing the R domain mutants S737A and S768A as well as the double mutant S737A/ S768A (Fig. 3A) produced dramatically enhanced conductances (S768A Ͼ S737A; Fig. 3B) upon stimulation with forskolin (2 ␮M) and IBMX (1 mM). Login to comment
130 The large Cl-conductances generated by S737A/S768A were inhibited by 5 ␮M of the specific chloride channel blocker CFTRinh-172 (Fig. 3D), or alternatively, the PKA inhibitor KT5720 and Cl- replacement by gluconate (not shown). Login to comment
131 Once again the differential roles of these two serines were observed because the conductance observed with S737A was almost 100 ␮S lower than that found with S768A. Login to comment
135 B, phosphoryl- ationoftheRdomainwithPKAandAMPK.AMPKphosphorylationwaslargely reduced or abolished in S737A and S768A, respectively (lower panel). Login to comment
146 Overall the data suggest that S737A, S768A, and the double mutant S737A/ S768A were no longer sensitive to stimulation or inhibition of AMPK indicating that phosphorylation at both serines is required (Fig. 3C, middle panels). Login to comment
151 In contrast to wtCFTR the CFTR mutants S737A (not shown), S768A, and S737A/S768A produced a high Cl- con- ductance under basal conditions, i.e. in the absence of IBMX and forskolin (Fig. 4, A and B). Login to comment
155 A, whole cell currents activated by IBMX (1 mM) and forskolin (2 ␮M) in wtCFTR and S737A/ S768A-CFTRexpressingoocytes.B,summaryofwholecellconductancesgen- erated by wtCFTR and different CFTR mutants. Login to comment
156 C, summary of CFTR whole cell conductances generated by wtCFTR and different CFTR mutants, and effects of phenformin and compound C. D, summary of the whole cell conductance activated by S737A/S768A-CFTR and inhibition by the CFTR blocker CFTRinh-172. Login to comment
161 S737A/S768A-CFTR generates a baseline conductance. Login to comment
162 A, effect of extracellular Cl- replacement by gluconate (glcn) on whole cell currents generated by wtCFTR, S768A-CFTR, and S737A/S768A-CFTR in the absence of IBMX and forskolin. Login to comment
164 C, summary of the whole cell conductances generated by wtCFTR and S737A/S768A-CFTRintheabsenceofstimulationwithIBMXandforskolinand effectsofphenformin,compoundC,andCFTRinh-172.DandE,baselinewhole cell conductances generated by wild type CFTR (D) and S737A/S768A-CFTR (E) and effects of compound C and the PKA inhibitor KT520. Login to comment
170 Moreover, the PKA inhibitor KT5720 (50 ␮M) inhibited this enhanced baseline CFTR conductance generated by S737A/ S768A irrespective of the presence of compound C (Fig. 4E). Login to comment
172 We interpret this finding to suggest that the sensitivity of S737A/ S768A-CFTR toward PKA inhibition is retained, which implies that PKA is now active after the loss of AMPK sensitivity. Login to comment
178 The initial recovery from PKA stimulation was equal in S737A/ S768A (1.1 Ϯ 0.3 ␮S/min) and wtCFTR (1.1 Ϯ 0.1 ␮S/min) (Fig. 5C), but was enhanced in ooctyes coexpressing kinase-dead AMPK␣1-K45R (2.8 Ϯ 0.4 ␮S/min), whereas overexpression of wtAMPK␣1beta1␥1 literally eliminated CFTR currents (Fig. 5B). Login to comment
180 Although AMPK largely antagonizes activation of CFTR, the mutated serines S737A and S768A do not seem to influence the recovery time from forskolin stimulation. Login to comment
210 C, inactivation of conductances generated by wtCFTR and S737A/S768A-CFTR. Login to comment
225 Even maximal stimulation of wtCFTR with a mixture of 8-Br- - cAMP, IBMX, and forskolin does not produce the same high level of conductance as S737A-CFTR or S768A-CFTR. Login to comment

PMID: 20952391 [PubMed] Wang G et al: "State-dependent regulation of cystic fibrosis transmembrane conductance regulator (CFTR) gating by a high affinity Fe3+ bridge between the regulatory domain and cytoplasmic loop 3."

No. Sentence Comment

162 Although Fig. 4E indicates that both S768A and S737A mutants were much inhibited by Fe3ϩ , these sites may not be excluded as Fe3ϩ ligands. Login to comment
180 Unlike S768A, S737A exhibited a weak DTT effect (Fig. 6D), suggesting that phosphorylated Ser-737 may not participate in Fe3ϩ binding. Login to comment

PMID: 21059651 [PubMed] Wang G et al: "The inhibition mechanism of non-phosphorylated Ser768 in the regulatory domain of cystic fibrosis transmembrane conductance regulator."

No. Sentence Comment

143 In contrast, curcumin had no such effect on S737A and H954A mutants, suggesting that they may be weak inhibitory residues, although disulfide cross-linking of S737C to H954C strongly inhibited channel activity (Figs. 2E and 4E). Login to comment
144 Because curcumin increased initial channel activity of most mutants at the R-CL3 interface after ATP was present, it is reasonable that subsequent PKA dependence was greatly reduced except for S737A and H954A. Login to comment
234 Error bars, S.E. TABLE 1 Potential roles in hydrogen bonding at the CL3-R domain interface Note that mutants whose channel activity was increased by curcumin in the presence of ATP are highlighted in boldface type. Residues Role in H-bond Mutants Arg Strong donor H950R, S768R, H950R/S768R, H950R/S768D, H950D/S768R Asp Strong acceptor H950D, S768D, H950D/S768D, H950R/S768D, H950D/S768R Thr, Gln, Ser, His Donor/Acceptor H950Q, S768T, WT Ala Negative control K946A, H950A, K951A, H954A, S955A, Q958A, S737A, S768A, ⌬R Inhibition of CFTR by Ser768 JANUARY 21, 2011•VOLUME 286•NUMBER 3 JOURNAL OF BIOLOGICAL CHEMISTRY 2177 matter whether cAMP was present or not in the extracellular perfusate. Login to comment
311 In contrast, the S737A mutation failed to promote channel opening by ATP and curcumin, although it was also closed to CL3 (Figs. 2-4). Login to comment

PMID: 21455600 [PubMed] Siwiak M et al: "Structural models of CFTR-AMPK and CFTR-PKA interactions: R-domain flexibility is a key factor in CFTR regulation."

No. Sentence Comment

121 However, experiments [7] have shown an 80% decrease in AMPK phosphorylation for the CFTR double mutant S737A-S768A. Login to comment
156 This shows that, at the molecular level, it is possible that 20% of the currently unexplained phosphorylation signal observed in the CFTR double mutant S737A-S768A incubated with AMPK may come from "activator" serines such as S813. Login to comment
155 This shows that, at the molecular level, it is possible that 20% of the currently unexplained phosphorylation signal observed in the CFTR double mutant S737A-S768A incubated with AMPK may come from "activator" serines such as S813. Login to comment

PMID: 19328185 [PubMed] Hegedus T et al: "Role of individual R domain phosphorylation sites in CFTR regulation by protein kinase A."

No. Sentence Comment

132 On the other hand, the much lower rate of S813 phosphorylation appeared to be reduced somewhat further by either the S700A or S737A substitutions, but not by the S768A substitution. Login to comment
152 Membranes isolated from BHK cells expressing S700A (A), S737A (B), S768A (C), or S813A (D) were subjected to phosphorylation for 0, 2, 4, 8, 16, 32, and 64 min. Login to comment
242 The S737A and S768A constructs exhibit slightly higher basal open probability (Po =0.06) compared to wild type (Po ≪0.01) in good agreement with data of Csanady et al. [24]. Login to comment
245 The channel properties of the S737A mutant were previously reported to be very similar to those of the wild type by Wilkinson et al. [10]. Login to comment
131 On the other hand, the much lower rate of S813 phosphorylation appeared to be reduced somewhat further by either the S700A or S737A substitutions, but not by the S768A substitution. Login to comment
151 Membranes isolated from BHK cells expressing S700A (A), S737A (B), S768A (C), or S813A (D) were subjected to phosphorylation for 0, 2, 4, 8, 16, 32, and 64 min. Login to comment
241 The S737A and S768A constructs exhibit slightly higher basal open probability (Po =0.06) compared to wild type (Po âa;0.01) in good agreement with data of Csanady et al. [24]. Login to comment
244 The channel properties of the S737A mutant were previously reported to be very similar to those of the wild type by Wilkinson et al. [10]. Login to comment

PMID: 18423665 [PubMed] Hegedus T et al: "Computational studies reveal phosphorylation-dependent changes in the unstructured R domain of CFTR."

No. Sentence Comment

28 Replacement of serine with alanine at position 737 or 768 was reported to produce a channel with a higher level of activity in both the non-phosphorylated and phosphorylated state.16,19-21 These studies led to the indirect conclusion that these two sites are inhibitory in their phosphorylated state. Login to comment
27 Replacement of serine with alanine at position 737 or 768 was reported to produce a channel with a higher level of activity in both the non-phosphorylated and phosphorylated state.16,19-21 These studies led to the indirect conclusion that these two sites are inhibitory in their phosphorylated state. Login to comment

PMID: 9305845 [PubMed] Winter MC et al: "Stimulation of CFTR activity by its phosphorylated R domain."

No. Sentence Comment

162 Mutation of the individual serines reduced Po, but to different extents: the S795A mutation had the largest effect and S737A had little or no effect in the presence of 1 mM ATP (Fig. 3a, b). Login to comment
185 Number of experiments for b/c and d were: 17/6 for wild-type (WT), 8 for wild-type (-PKA), 8/7 for S660A, 5/5 for S737A, 8/6 for S795A, 9/7 for S813A, and 11/4 for S-Quad-A. Login to comment
188 0.0 0.1 0.2 0.3 0.4 0.5 0.6 Po 0 0.2 0.4 0.6 0.8 1 10 [ATP] (mM) S813A S795A S737A S660A WT (-PKA) WT Figure 4 Effect of ATP concentration on Po of CFTR containing phosphorylation site mutations. Login to comment

PMID: 9252549 [PubMed] Wilkinson DJ et al: "CFTR activation: additive effects of stimulatory and inhibitory phosphorylation sites in the R domain."

No. Sentence Comment

15 In contrast, alanine substitution at serine-737 or -768 actually decreased the KA for activation, suggesting that phosphorylation at either of these sites is inhibitory. Login to comment
87 S686A 0.75 ?I 0.07 9 N PKC site S700A 0.86 t 0.07* 12 S ++ +++ S712A 0.67 t 0.12 9 N - ++ S737A 0.35 5 0.05* 8 I +++ +++ S768A 0.09 IT 0.03* 8 I - ++++ S795A 1.24 + 0.22* 9 S +++ ++++ S813A 3.18-+0.36* 6 S ++++ ++ Values of half-maximal inhibition constant for activation (KA) are means + SE by 3-isobutyl-1-methylxanthine (IBMX) obtained for wild-type cystic fibrosis transmembrane conductance regulator (CFTR) and variants with single serine-to-alanine substitutions. Login to comment
107 Symbols show averaged IBMX dose-response data for wild-type CFTR and several single-site serine-to-alanine mutants (S660A, S737A, S768A, S795A, and S813A). Login to comment

PMID: 7690753 [PubMed] Rich DP et al: "Regulation of the cystic fibrosis transmembrane conductance regulator Cl- channel by negative charge in the R domain."

No. Sentence Comment

48 Mutants are named by the number of the amino acid residue preceded by the wild-type amino acid and followed bythe aminoacid to which the residue was changed using the single-letter aminoacid code; for example,inthemutant CFTR S66OA,S737A,S795A,S813A (also called CFTR S-Quad-A for convenience), alanines are substituted for serines at amino acid positions 660, 737, 795, and 813. Login to comment
66 In vivo phosphorylation ofwild-typeand mutant CFTR CFTR S-Quad-A (S66OA,S737A,S795A,S813A), or CFTR S-Oct-A (S660A,S686A,S700A,S712A,S737A,S768A,S795A,S813A) weretran- siently expressed in HeLa cells. Login to comment
89 Functional assay of CFTR S-Quad-Aby SPQ fluorescence.The changein SPQfluorescenceis shown for HeLa cellsexpress- ing wild-type CFTR (n = 53, where n = number of cells) or CFTR S-Quad-A tS660A,S737A,S795A,S813A)(n = 53) and for virus only- infectedcells(no plasmid) as a negative control (n = 47). Login to comment
94 Single-channelanalysis of gle-channel traces from an excised, in-cell expressing CFTR S-Quad-A side-out membrane patch from a HeLa (S660A,S737A,S795A,S813A).Each trace is about 14s long. Login to comment
109 RESULTS Serine-to-Alanine Substitutionsin the R Domain Do NotAbolish CFTR Cl- Channel Activity-& a firststep to address the possibility that additional PKA phosphorylation sites might regulate CFTR, we asked whether the S-Quad-A (S660A,S737A,S795A,S813A) mutant could still bephosphorylated in vivofollowing CAMPstimulation. Login to comment
121 The single-channel open-stateprobabil- ity for wild-type CFTR (n = 14), CFTR S-Quad-A (S600A,S737A, S712A,S737A,S768A,S795A,S813A) (n = 7), or CFTR S-Oct-D (S660D,S686D,S700D,S712D,S737D,S768D,S795D,S813D)(n = 7in ATP alone (-PKA);n = 10 inPKA and ATP (+PkX))C1-channels was determined as described under "Experimental Procedures." Login to comment
123 S795A,S813A) (n = 5). Login to comment
139 CFTR (A) or CFTR S-Quad-A (S660A,S737A,S795A,S813A)( B )were transiently expressed inCOS-7 cells. Login to comment
174 Fig. 1lA shows that simultaneous substitutionof the four in vivo PKA sites with aspartates (in the S-Quad-D mutant) did not generate constitutively active CFTR C1-channels as assessed by the SPQ fluorescence assay: the mutant channels opened only after stimulation by CAMP.We observed similar results withCFTR S-Quint-D which contained substitutionsof A MockS-Hept-A C S-OCt-A transfected (A) or were transfected with pMT-CFTR S-Hept-A (S660A,S686A,S700A,S712A,S737A,S768A,S795A) (B)or pMT-CFl`R S-Oct-A FIG.9. Login to comment
176 COS-7 cells were mock- (S660A,S686A,S700A,S712A,S737A,S768A,S795A,S813A)(C). Login to comment
47 Mutants are named by the number of the amino acid residue preceded by the wild-type amino acid and followed bythe aminoacid to which the residue was changed using the single-letter amino acid code; for example,inthemutant CFTR S66OA,S737A,S795A,S813A (also called CFTR S-Quad-A for convenience), alanines are substituted for serines at amino acid positions 660, 737, 795, and 813. Login to comment
65 In vivo phosphorylation ofwild-typeand mutant CFTR CFTR S-Quad-A (S66OA,S737A,S795A,S813A), or CFTR S-Oct-A (S660A,S686A,S700A,S712A,S737A,S768A,S795A,S813A) were transiently expressed in HeLa cells. Login to comment
95 Single-channelanalysis of gle-channel traces from an excised, in-cell expressing CFTR S-Quad-A side-out membrane patch from a HeLa (S660A,S737A,S795A,S813A).Each trace is about 14s long. Login to comment
111 RESULTS Serine-to-Alanine Substitutions in the R Domain Do NotAbolish CFTR Cl-Channel Activity-& a firststep to address the possibility that additional PKA phosphorylation sites might regulate CFTR, we asked whether the S-Quad-A (S660A,S737A,S795A,S813A) mutant could still be phosphorylated in vivofollowing CAMPstimulation. Login to comment
142 CFTR (A) or CFTR S-Quad-A (S660A,S737A,S795A,S813A) ( B )were transiently expressed inCOS-7 cells. Login to comment
178 Fig. 1lA shows that simultaneous substitution of the four in vivo PKA sites with aspartates (in the S-Quad-D mutant) did not generate constitutively active CFTR C1-channels as assessed by the SPQ fluorescence assay: the mutant channels opened only after stimulation by CAMP.We observed similar results withCFTR S-Quint-D which contained substitutionsof A Mock S-Hept-A C S-OCt-A transfected (A) or were transfected with pMT-CFTR S-Hept-A (S660A,S686A,S700A,S712A,S737A,S768A,S795A) (B) or pMT-CFl`R S-Oct-A FIG. 9. Login to comment
180 COS-7 cells were mock- (S660A,S686A,S700A,S712A,S737A,S768A,S795A,S813A) (C). Login to comment

PMID: 7684377 [PubMed] Chang XB et al: "Protein kinase A (PKA) still activates CFTR chloride channel after mutagenesis of all 10 PKA consensus phosphorylation sites."

No. Sentence Comment

7 A previous investigationconcluded thatactivationby PKA is crit- icallydependent on phosphorylation at four of the nine predicted PKA sites in the R domain (S660A, S737A, S795A,SS13A), becausea "Quad"mutant lacking these sites couldnotbeactivated. Login to comment
37 The following mutations were introduced into CFTR, S422A (TCT to GCT), S660A (TCA to GCA), S686A (TCT to GCT), S700A (TCT toGCT), S712A (TCC to GCC), S737A (TCC to GCC), S768A (TCT toGCT), T788A (ACAto GCA),S795A (TCA to GCA), S813A (TCA to GCA), S660E (TCA to GAA),S737E (TCC to GAG), S795E (TCA to GAA), and S813E (TCA to GAA). Login to comment
39 The counterparts of CFTR cDNA in pUCF2.5 were replaced by PCR- mutated versions by interchange of the following fragments, S660A, DraIIIIEcoRI fragment, S737A, EcoRIIHpaIfragment, S795A and S813A,StyIIStyI fragment(Fig. 1A). Login to comment
40 A mutant containing S660/737/ 795/813A (pUCF2.5/4SA) was assembled by replacing the counterparts of a plasmid containing S795/813A with the DraIII/EcoRI fragment (S660A) and EcoRI/HpaI fragment (S737A). Login to comment

PMID: 23760269 [PubMed] Billet A et al: "Role of tyrosine phosphorylation in the muscarinic activation of the cystic fibrosis transmembrane conductance regulator (CFTR)."

No. Sentence Comment

102 Carbachol Stimulates CFTR through PKA and Non-PKA Signaling Pathways-To explore PKA-independent regulation of CFTR without using inhibitors that might have confounding effects on other pathways, we studied the activation of 15SA-CFTR (S422A/S660A/S670A/S686A/T690A/S700A/S712A/ S737A/S753A/S768A/T787A/T788A/S790A/S795A/S813A). Login to comment

PMID: 24727471 [PubMed] Luz S et al: "LMTK2-mediated phosphorylation regulates CFTR endocytosis in human airway epithelial cells."

No. Sentence Comment

8 Compared with the wild-type CFTR, the phosphorylation-deficient mutant CFTR-S737A shows increased cell surface density and decreased endocytosis. Login to comment
203 To demonstrate specificity of the phosphosite antibody, parental CFBE41o-cells were transiently transfected with WT-CFTR or CFTR with the S737A substitution (CFTR-S737A) inactivating the phosphorylation site (31). Login to comment
204 Even with calyculin A treatment of the cells, the CFTR-S737A mutant was not detected by antibody Ab-737, unlike the WT-CFTR (Fig. 5B). Login to comment
233 B, parental CFBE41o-cells were transfected with the WT-CFTR (WT) or the mutant CFTR-S737A (S737A). Login to comment
234 Unlike the WT-CFTR, the CFTR-S737A was not detected by the antibody Ab-737 in WCL. Login to comment
287 The Phosphorylation-deficient CFTR-S737A Mutant is More Abundant at the Plasma Membrane Due to Decreased Endocytosis-If the LMTK2 mediated phosphorylation of CFTR-Ser737 induces CFTR endocytosis, eliminating the phosphorylation site by the S737A substitution should reduce CFTR endocytosis. Login to comment
288 Parental CFBE41o-cells were transfected with WT-CFTR or the CFTR-S737A mutant. Login to comment
308 The S737A mutation attenuated CFTR endocytosis and increased the plasma membrane abundance of CFTR (Fig. 8). Login to comment
331 Experiments demonstrating that compared with WT-CFTR the phosphorylation-deficient mutant CFTR S737A has increased plasma membrane abundance at steady-state and decreased endocytosis in CFBE41o-cells. Login to comment
332 Parental CFBE41o-cells were transfected with the WT-CFTR (WT) or the CFTR-S737A mutant (S737A), and cells were cultured on collagen-coated tissue culture plates for 48 h to form monolayers. Login to comment
334 Immunoblots (A) and summaryofdata(B)demonstratingthattheS737AmutationdecreasedCFTRendocytosis.TheamountofbiotinylatedWT-CFTRorCFTR-S737Aremainingafter the GSH treatment at 4 &#b0;C without warming to 37 &#b0;C was considered background and was subtracted from the amount of biotinylated WT-CFTR or CFTR-S737A remaining after warming to 37 &#b0;C. Login to comment
335 Endocytosis of WT-CFTR or CFTR-S737A was calculated after subtracting the background (see above) and was expressed as the percent of WT-CFTR or CFTR-S737A remaining biotinylated before and after warming to 37 &#b0;C. WT-CFTR and CFTR-S737A was detected with antibody CFF596. Login to comment
338 The whole cell lysate (WCL) expression of WT-CFTR or CFTR-S737A mutant detected with antibody CFF596 was used as a loading control to account for small differences in the expression levels of the transiently transfected proteins. Login to comment

PMID: 26079370 [PubMed] Malik FA et al: "Sphingosine-1-Phosphate Is a Novel Regulator of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Activity."

No. Sentence Comment

123 (F) S1P fails to inhibit iodide efflux in na&#ef;ve BHK cells transiently transfected with a plasmid encoding a mutated CFTR containing a serine-to-alanine substitution at amino acid 737 (CFTRS737A ). Login to comment