A sodium-dependent sugar carrier expressed in oocytes

A laboratory clones a gene for a candidate sugar transporter, T1, and tests whether the protein moves glucose-like molecules across a membrane. Capped RNA encoding T1 is injected into frog oocytes from a single batch. Control oocytes from the same batch are injected with an equal volume of water. After incubation, groups of 10 oocytes are placed at room temperature in a bath containing a radiolabeled glucose analog that cells do not metabolize. Uptake is measured after a fixed 30-minute period that was shown earlier to lie within the linear range of accumulation. Values below are T1-specific uptake, defined as the mean for T1-injected oocytes minus the mean for water-injected oocytes. Water-injected background was small and rose linearly with analog concentration.

Experiment 1. The bath contained 100 mM NaCl. T1-specific uptake was measured at several analog concentrations.

Analog (mM)    Uptake (pmol per oocyte per hour)
0.5            12
1              20
2              30
5              43
10             50
20             55

Experiment 2. The assay was repeated with the 100 mM NaCl replaced by 100 mM choline chloride; T1 does not recognize the choline cation. At every analog concentration, T1-specific uptake was close to zero.

Experiment 3. Uptake was measured at two analog concentrations in the NaCl bath, with and without 10 mM of an unlabeled sugar, Q, that is structurally similar to the analog.

Analog (mM)    Without Q    With Q (pmol per oocyte per hour)
2              30           20
20             55           50

In a separate voltage-clamp experiment, adding the analog to the bath produced an inward current in T1-injected oocytes but not in water-injected oocytes.