Effects of MTS reagents on [3H]NBMPR binding and [3H]2-chloroadenosine uptake by cells transfected with wild-type hENT1 or the four cysteine mutants
Data shown are the means ± S.E.M. from at least five independent experiments conducted as described in Figs. 3, 4, and 9.
| Cell Line and MTS Reagent | [3H]NBMPR Binding (Bmax) | [3H]2-Chloroadenosine Uptake (Vmax) | ||||
|---|---|---|---|---|---|---|
| Control | Treated | Control | Treated | |||
| sites/cell × 105 | % change | pmol · μl−1 · s−1 | % change | |||
| WT | ||||||
| MMTS | 3.9 ± 0.4 | 5.6 ± 0.4 | 62 ± 11* | 10.8 ± 1.1 | 6.4 ± 1.0 | −36 ± 13* |
| MTSET | 3.6 ± 0.4 | 3.2 ± 0.3 | −13 ± 4* | 9.1 ± 1.6 | 11.3 ± 1.7 | 45 ± 24* |
| MTSES | 4.1 ± 0.5 | 4.0 ± 0.7 | −4 ± 8 | 8.1 ± 1.2 | 7.6 ± 1.1 | −7 ± 16 |
| C87S | ||||||
| MMTS | 1.3 ± 0.1 | 1.8 ± 0.1 | 49 ± 12* | 6.6 ± 0.8 | 5.2 ± 0.8 | −20 ± 14* |
| MTSET | 1.1 ± 0.1 | 1.1 ± 0.2 | 4 ± 11 | 7.3 ± 0.5 | 11.3 ± 2.5 | 51 ± 24* |
| MTSES | 1.4 ± 0.1 | 1.3 ± 0.1 | −7 ± 6 | 7.4 ± 0.6 | 8.3 ± 1.7 | 19 ± 27 |
| C193S | ||||||
| MMTS | 2.1 ± 0.5 | 4.6 ± 1.0 | 106 ± 28* | 5.8 ± 0.8 | 5.0 ± 1.9 | −11 ± 9 |
| MTSET | 2.5 ± 0.4 | 2.4 ± 0.5 | −6 ± 11 | 4.4 ± 0.6 | 4.0 ± 0.2 | −5 ± 10 |
| MTSES | 2.9 ± 0.3 | 3.0 ± 0.6 | 7 ± 21 | 5.7 ± 1.0 | 5.2 ± 0.7 | 18 ± 27 |
| C213S | ||||||
| MMTS | 2.8 ± 0.6 | 5.4 ± 1.5 | 56 ± 20* | 19 ± 4 | 12 ± 3 | −40 ± 7* |
| MTSET | 5.2 ± 1.5 | 3.8 ± 0.8 | −18 ± 12* | 25 ± 5 | 24 ± 5 | −3 ± 9 |
| MTSES | 11.4 ± 1.0 | 13.6 ± 1.5 | 21 ± 23 | 44 ± 9 | 47 ± 9 | 31 ± 30 |
| C222S | ||||||
| MMTS | 2.3 ± 0.3 | 2.2 ± 0.3 | 4 ± 8 | 10.8 ± 2.5 | 6.7 ± 2.1 | −53 ± 21* |
| MTSET | 1.9 ± 0.3 | 1.5 ± 0.3 | −18 ± 12* | 11.1 ± 1.7 | 7.1 ± 0.8 | −20 ± 22 |
| MTSES | 1.8 ± 0.1 | 1.8 ± 0.3 | −1 ± 14 | 10.1 ± 1.6 | 8.7 ± 2.0 | −12 ± 20 |
* P <0.05, significant change upon treatment with the MTS reagent (Student's t test for paired samples).