ABCMdb

PMID: 7534226

Sheppard DN, Ostedgaard LS, Winter MC, Welsh MJ
Mechanism of dysfunction of two nucleotide binding domain mutations in cystic fibrosis transmembrane conductance regulator that are associated with pancreatic sufficiency.
EMBO J. 1995 Mar 1;14(5):876-83., [PubMed]

Sentences

No. Mutations Sentence Comment

1 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu To understand better how mutations in CFTR disrupt Cl- channel function and to learn about the relationship between genotype and phenotype, we studied two CF mutants, A455E and P574H, that are associated with pancreatic sufficiency. Login to comment 2 ABCC7 p.Pro574His ABCC7 p.Ala455Glu A455E and P574H are located close to conserved ATP binding motifs in CFTR. Login to comment 5 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu These mutants had normal or increased Cl- channel activity: A455E had an open-state probability (PO) similar to wild-type, and P574H had an increased PO because bursts of activity were prolonged. Login to comment 22 ABCC7 p.Arg334Trp ABCC7 p.Arg347Pro In contrast, three mutations (RI 17H, R334W and R347P) in the first membrane-spanning domain (MSD1) are associated with the PS phenotype (Kristidis et al., 1992; The Cystic Fibrosis Genotype-Phenotype Consortium, 1993). Login to comment 27 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Two such mutations are located within NBD1: A455E and P574H (Kerem et al., 1990; Kristidis et al., 1992). Login to comment 28 ABCC7 p.Ala455Glu Although these mutations are uncommon, A455E accounts for -10% of CF chromosomes in some areas of The Netherlands and in the Saguenay-Lac St Jean region of Quebec, Canada (Rozen et al., 1992; Gan et al., 1994). Login to comment 29 ABCC7 p.Pro574His ABCC7 p.Ala455Glu A455E and P574H affect residues in NBD1 that are located close to the Walker A (residues 458-464) and B (residues 568-572) motifs, respectively. Login to comment 31 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Therefore, we tested the hypothesis that A455E and P574H might alter the function of CFTR Cl- channels. Login to comment 33 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu 86 Oxford University Press Results Expression of A455E and P574H generates cAMP-activated CL currents To learn whether A455E and P574H could form regulated Cl- channels in epithelia, we expressed these mutants in cAMP~ N E ._0. -1 P574H cAMPI A455E 'cAMP 5 pA/cm2L 5 min b Fig. 1. cAMP agonists activate apical membrane CF- currents in FRT epithelia expressing A455E and P574H. Login to comment 34 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu -1 P574H cAMPI A455E 'cAMP 5 pA/cm2L 5 min b Fig. 1. cAMP agonists activate apical membrane CF- currents in FRT epithelia expressing A455E and P574H. Login to comment 44 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 1 shows that A455E and P574H generated cAMP-stimulated apical Cl- currents, but AF508 did not. Login to comment 45 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu However, the magnitude of current generated by A455E and P574H was <20% that of wild-type CFTR in the rank order: wild-type CFTR>> P574H > A455E > AF508. Login to comment 46 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu When we expressed A455E and P574H in HeLa cells and studied them with the whole-cell patch-clamp technique, we found properties qualitatively similar to those observed with wild-type CFTR (Welsh et al., 1992; Riordan, 1993). Login to comment 47 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Figure 2 shows data from studies of P574H; similar results were obtained with A455E (data not shown). Login to comment 48 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 2 shows data from studies of P574H; similar results were obtained with A455E (data not shown). Login to comment 51 ABCC7 p.Pro574His In contrast, when P574H was expressed using the same conditions, cAMP agonists stimulated current in only five out of 12 cells, IP Cl [CI]E 7 ; - 25~O20 h / -200 Fig. 2. Login to comment 52 ABCC7 p.Pro574His ABCC7 p.Pro574His Whole-cell properties of P574H. Login to comment 53 ABCC7 p.Pro574His Whole-cell properties of P574H. Login to comment 66 ABCC7 p.Pro574His ABCC7 p.Ala455Glu CFTR 0 * I1 -50 * CFTR A A455E * P574H pA Fig. 3. Login to comment 67 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Single-channel properties of A455E and P574H. Login to comment 68 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu (A) Representative single-channel recordings are from excised inside-out membrane patches from HeLa cells transiently expressing wild-type CFTR, A455E or P574H. Login to comment 69 ABCC7 p.Pro574His ABCC7 p.Ala455Glu (A) Representative single-channel recordings are from excised inside-out membrane patches from HeLa cells transiently expressing wild-type CFTR, A455E or P574H. Login to comment 72 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The voltages were -45 (CFTR), -46 (A455E) or -47 mV (P574H). Login to comment 73 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu (B) Single-channel I-V relationships of CFTR (U), A455E (A) and P574H (A). Login to comment 74 ABCC7 p.Pro574His ABCC7 p.Ala455Glu (B) Single-channel I-V relationships of CFTR (U), A455E (A) and P574H (A). Login to comment 76 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Conductance was CFT'R 7.76 ± 0.14, A455E 7.40 ± 0.25 and P574H 7.84 ± 0.26 pS; n = 6. Login to comment 77 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Conductance was CFT'R 7.76 &#b1; 0.14, A455E 7.40 &#b1; 0.25 and P574H 7.84 &#b1; 0.26 pS; n = 6. Login to comment 81 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Single-channel properties of A455E and P574H To determine why A455E and P574H generated less macroscopic Cl- current, we examined their single-channel properties in excised, inside-out patches of membrane. Login to comment 82 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Single-channel properties of A455E and P574H To determine why A455E and P574H generated less macroscopic Cl-current, we examined their single-channel properties in excised, inside-out patches of membrane. Login to comment 83 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The tracings show that the single-channel current amplitudes of A455E and P574H were similar to that of wild-type CFTR. Login to comment 84 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The tracings show that the single-channel current amplitudes of A455E and P574H were similar to that of wild-type CFTR. Login to comment 86 ABCC7 p.Ala455Glu Figure 3A shows that A455E had a pattern of gating 878 that closely resembled that of wild-type CFTR. Login to comment 87 ABCC7 p.Ala455Glu Figure 3A shows that A455E had a pattern of gating 878 that closely resembled that of wild-type CFTR. Login to comment 88 ABCC7 p.Pro574His However, P574H showed a different pattern of activity. Login to comment 89 ABCC7 p.Pro574His However, P574H showed a different pattern of activity. Login to comment 92 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 3C shows that the PO of A455E was similar to that of wild-type CFTR, whereas the PO of P574H was increased significantly. Login to comment 93 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 3C shows that the PO of A455E was similar to that of wild-type CFTR, whereas the PO of P574H was increased significantly. Login to comment 95 ABCC7 p.Pro574His We focused primarily on P574H to determine why the P0 of this mutant was greater than that of wild-type CFTR. Login to comment 96 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Maximum likelihood A 0.5pA 2s 50OC mV 0.6 PO 0.4 0.2- -0.1 - -1.2 0.0 J- - -0.4 A B pi (/S) al (f/S) pi 02I -- C2 _ ° a1 a2 12 CFTR A455E P574H CFTR A455E P574H Fig. 4. Login to comment 97 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Effect of A455E and P574H on single-channel kinetics. Login to comment 98 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Effect of A455E and P574H on single-channel kinetics. Login to comment 100 ABCC7 p.Pro574His ABCC7 p.Ala455Glu (B) Effect of A455E and P574H on rate constants determined by the maximum likelihood fit to the model in (A). Login to comment 101 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Data are from four CFTR, two A455E and six P574H single-channel patches. Login to comment 102 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Data are from four CFTR, two A455E and six P574H single-channel patches. Login to comment 103 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The voltages were -50 ± 2 (CFTR), -48 ± I (P574H) and -47 ± 1 mV (A455E). Login to comment 104 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The voltages were -50 &#b1; 2 (CFTR), -48 &#b1; I (P574H) and -47 &#b1; 1 mV (A455E). Login to comment 110 ABCC7 p.Pro574His The effect of P574H on the rate constants is summarized in Figure 4B. Login to comment 111 ABCC7 p.Pro574His ABCC7 p.Pro574His The major effect of the P574H mutation was to decrease PI and al to -40% of wild-type values. Login to comment 112 ABCC7 p.Pro574His The major effect of the P574H mutation was to decrease PI and al to -40% of wild-type values. Login to comment 118 ABCC7 p.Pro574His All of the changes in rate constants produced by P574H tend to increase Tb. Login to comment 119 ABCC7 p.Pro574His ABCC7 p.Pro574His The increase in burst duration is responsible for the increase in PO (Figure 3C), but it is partially offset by the decreased frequency with which P574H channels move into the bursting state (i.e. the decreased I31). Login to comment 120 ABCC7 p.Pro574His ABCC7 p.Ala455Glu In two experiments we examined the gating kinetics of single phosphorylated A455E Cl- channels. Login to comment 121 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu A455E did not alter PI or al, but tended to increase P2 and a2 in both experiments (Figure 4B). Login to comment 122 ABCC7 p.Ala455Glu A455E did not alter PI or al, but tended to increase P2 and a2 in both experiments (Figure 4B). Login to comment 124 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Regulation of A455E and P574H by intracellular nucleotides Intracellular MgATP regulates CFTR through interactions with the NBDs. Login to comment 125 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Regulation of A455E and P574H by intracellular nucleotides Intracellular MgATP regulates CFTR through interactions with the NBDs. Login to comment 126 ABCC7 p.Gly551Ser ABCC7 p.Gly1244Glu ABCC7 p.Gly1349Asp ABCC7 p.Ser1255Pro ABCC7 p.Ser1255Pro For example, G551S, G1244E, S1255P and G1349D had a markedly reduced PO at all concentrations of MgATP tested, and S1255P was less potently stimulated by MgATP (Anderson and Welsh, 1992; Smit et al., 1993). Login to comment 127 ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Gly551Ser ABCC7 p.Gly1244Glu ABCC7 p.Gly1349Asp ABCC7 p.Ser1255Pro ABCC7 p.Ser1255Pro Therefore, we tested the hypothesis that ATP-dependent regulation of A455E and P574H was altered. Login to comment 128 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Therefore, we tested the hypothesis that ATP-dependent regulation of A455E and P574H was altered. Login to comment 129 ABCC7 p.Pro574His ABCC7 p.Ala455Glu At each concentration of MgATP tested, A455E had PO values similar to those of wild-type CFTR, whereas P574H had higher values of PO. Login to comment 130 ABCC7 p.Pro574His ABCC7 p.Ala455Glu At each concentration of MgATP tested, A455E had PO values similar to those of wild-type CFTR, whereas P574H had higher values of PO. Login to comment 131 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 5B shows that ADP (1 mM) produced an equivalent inhibition of wild-type CFTR, A455E and P574H. Login to comment 132 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Figure 5B shows that ADP (1 mM) produced an equivalent inhibition of wild-type CFTR, A455E and P574H. Login to comment 133 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Analysis of the processing of A455E and P574H The data indicate that the reduced apical membrane Cl- current produced by A455E and P574H cannot be attributed to the reduced function of single mutant channels. Login to comment 134 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Analysis of the processing of A455E and P574H The data indicate that the reduced apical membrane Cl-current produced by A455E and P574H cannot be attributed to the reduced function of single mutant channels. Login to comment 139 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu However, band C production was reduced in A455E and P574H and was not apparent until 12-24 h after infection, and only then as a faint A B CFTR A455E P574H MgATP (mM) Fig. 5. Login to comment 140 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Effects of MgATP concentration and ADP on the activity of phosphorylated A455E and P574H channels. Login to comment 141 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Effects of MgATP concentration and ADP on the activity of phosphorylated A455E and P574H channels. Login to comment 142 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu and intracellular MgATP concentration for A455E, P574H and wild-type CFTR Cl- channels. Login to comment 143 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Data points are the mean ± SEM of n = 4-6 for CFTR (-), n = 5 for A455E (A) and n = 3-5 for P574H (0) at each concentration. Login to comment 144 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu The voltages were -86 ± 1 (CFTR), -49 ± 2 (A455E) and -49 ± 1 mV (P574H). Login to comment 147 ABCC7 p.Pro574His ABCC7 p.Ala455Glu (B) Effect of intracellular ADP on the activities of CFTR, A455E and P574H Cl- channels. Login to comment 149 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Values are the mean ± SEM of n = 4 for CFTR, A455E and P574H; the voltages were -49 ± 1 (CFTR), -48 ± 2 (A455E) and -52 ± 1 mV (P574H). Login to comment 150 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Percent inhibition of NXP0 by 1 mM ADP was CFTR 77 ± 3, A455E 70 ± 3 and P574H 75 ± 6%. Login to comment 156 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Although the production of mature band C protein was greatly reduced in A455E and P574H, it was greater than that observed with AF508. Login to comment 157 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Discussion Relationship between apical membrane CL currents and the properties of mutant CFTR Because the A455E and P574H mutations cause CF, we hypothesized that Cl- transport would be reduced in cells expressing these mutants. Login to comment 161 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu If we set each of these variables to 100% for wild-type CFTR, we can then compare the apical Cl- current generated by wild-type and AF508 CFTR with that produced by A455E and P574H. Table I presents values of each variable, the predicted value of fCI(apical) determined by calculating N x i X P0 and the observed value of fCI(apical). Login to comment 164 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Thus, these data provide a molecular explanation for the quantitative decrease in cAMP-stimulated apical membrane Cl- current generated by A455E and P574H. Table I. Login to comment 165 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Comparison of predicted apical membrane Cl- current, N X i X PO, and measured cAMP-stimulated apical membrane Cl- current, f'(apical), for wild-type and mutant CFTR N i PO N X i X PO fl'(apical) (%) (%) (%) (%) (%) Wild-type 100 100 100 100 100 AF508 4 100 38 1.6 0 A455E 11 100 93 10.5 8 P574H 15 100 139 21.1 17 N, the number of Cl- channels in the apical membrane; i, single-channel current; PO- open-state probability. Login to comment 171 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Mechanism of altered gating by A455E and P574H A455 and P574 lie near the conserved Walker A and B motifs of NBD1. Login to comment 174 ABCC7 p.Pro574His P574H decreased 01, a rate constant that is also controlled by ATP and which describes the transition to the bursting state. Login to comment 175 ABCC7 p.Pro574His Because of the proximity of P574 to D572 (the Walker B aspartate of NBD1), we speculate that P574H alters the interaction of NBD1 with MgATP, and/ or the consequences thereof, and hence slows the rate of channel opening. Login to comment 176 ABCC7 p.Pro574His P574H also altered al. Login to comment 177 ABCC7 p.Pro574His ABCC7 p.Ala455Glu ABCC7 p.Lys464Ala It is interesting to compare the effect that different mutations in NBD1 had on al: P574H decreased a1, A455E did not change al, and K464A (a mutant of the Walker A lysine of NBDl; Carson and Welsh, 1995) appeared to increase the rate of channel closing. Login to comment 181 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Expression of wild-type CFTR, AF508, A455E and P574H. Login to comment 187 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The radioactivity in gels of immunoprecipitated and phosphorylated wild-type CFTR, AF508, A455E and P574H was quantitated as described in Materials and methods. Login to comment 188 ABCC7 p.Pro574His ABCC7 p.Ala455Glu The values are the mean + SEM of n = 3 for CFTR, AF508, A455E and P574H. Login to comment 189 ABCC7 p.Pro574His understand how CFTR is regulated by the NBDs and to learn how P574H alters this function. Login to comment 190 ABCC7 p.Ala455Glu It is interesting that A455E appeared to increase IP2 and a2, thereby increasing the frequency of transitions within a burst of activity. Login to comment 192 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Our data from A455E, plus the finding that P574H altered P2, suggest that the NBDs may also be involved in regulating gating within a burst. Login to comment 193 ABCC7 p.Ala455Glu One possible interpretation of the effect of A455E is that the mutation reduced the height of an energy barrier between the short-lived closed state (C2) and the open state. Login to comment 194 ABCC7 p.Pro574His Cellular processing ofA455E and P574H An important mechanism of dysfunction of CFTR containing CF-associated mutations is misprocessing, so that the mutant protein fails to leave the endoplasmic reticulum and traffic to the Golgi complex and then on to the cell surface (Cheng et al., 1990). Login to comment 198 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Our data indicate that processing mutations are not necessarily an all-or-none defect; in contrast to AF508, some of the mutant A455E and P574H protein had a glycosylation pattern consistent with processing in the Golgi complex and generated cAMP-stimulated apical Cl- currents. Login to comment 199 ABCC7 p.Pro574His ABCC7 p.Ala455Glu We evaluated the processing of A455E and P574H in HeLa and FRT cells incubated at 37°C. Login to comment 203 ABCC7 p.Ala455Glu For example, A455E formed a channel that was misprocessed yet had conductive and regulatory properties very similar to those of wild-type CFTR. Login to comment 204 ABCC7 p.Pro574His More strikingly, P574H, which was also misprocessed, formed a channel that was even more active than wild-type. Login to comment 206 ABCC7 p.Gly551Asp Conversely, previous data have shown that some mutants, such as G551D, have little Cl- channel function, yet are processed correctly (Cheng et al., 1990; Kartner et al., 1992). Login to comment 209 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Implications for cystic fibrosis These data suggest that the mutations A455E and P574H cause CF because they disrupt the processing of CFTR and its delivery to the cell surface. Login to comment 210 ABCC7 p.Pro574His ABCC7 p.Pro574His ABCC7 p.Ala455Glu However, they also suggest that these mutations are associated with a milder (PS) clinical phenotype because a small amount of the 881 CFTR A kDa 210 - 170- 116 - 98 - AF508 N. A455E I I P574H 4- Band C 4- Band B B band C bands A+B P574H aL mutant protein is processed correctly, retains normal or greater than normal Cl- channel function and thus generates residual cAMP-stimulated apical membrane C1- cur- rents. Login to comment 212 ABCC7 p.Ala455Glu (1994) who demonstrated that rectal biopsies from patients bearing the A455E mutation retain residual Cl- secretion. Login to comment 215 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu (1994) found that patients with the A455E mutation were PS, but it is not clear whether or not the A455E mutation confers milder lung disease. Login to comment 216 ABCC7 p.Ala455Glu Uncertainty about the severity of lung disease in these patients highlights the difficulty in assessing the clinical phenotype associated with the A455E and P574H1mutations in relatively small numbers of patients. Login to comment 218 ABCC7 p.Arg117His ABCC7 p.Arg334Trp ABCC7 p.Arg347Pro It is interesting to compare our present results with those from our study of mild CF mutations that occur in MSD1 (R117H, R334W and R347P). Login to comment 220 ABCC7 p.Pro574His ABCC7 p.Ala455Glu These values are in the same range as those found for A455E (8%) and P574H (17%). Login to comment 223 ABCC7 p.Pro574His ABCC7 p.Ala455Glu A455E or P574H could prove to be of value in the development of therapies designed to augment the delivery to and the retention of mutant protein at the plasma membrane (Cheng et al., 1990; Lukacs et al., 1993). Login to comment 224 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Although AF508 could be used to assess the processing defect, A455E or P574H might prove to be more tractable models for evaluating strategies to correct mislocalization because the processing defect is only partial and the conductance and regulation of the Cl- channels is more nearly normal. Login to comment 226 ABCC7 p.Pro574His ABCC7 p.Ala455Glu Such strategies might also be applied to patients bearing the A455E and P574H mutations because they appear to have at least some functional protein present in the plasma membrane. Login to comment