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<i>Populus alba</i> ‘Villafranca’ clone is well-known for its tolerance to cadmium (Cd). To determine the mechanisms of Cd tolerance of this species, wild-type (wt) plants were compared with transgenic plants over-expressing an aquaporin (<i>aqua1</i>, GenBank GQ918138). Plants were maintained in hydroponic conditions with Hoagland’s solution and treated with 10 µM of Cd, renewed every 5 d. The transcription levels of heavy metal transporter genes (<i>PaHMA2</i>, <i>PaNRAMP1.3</i>, <i>PaNRAMP2</i>, <i>PaNRAMP3.1</i>, <i>PaNRAMP3.2</i>, <i>PaABCC9</i>, and <i>PaABCC13</i>) were analyzed at 1, 7, and 60 d of treatment. Cd application did not induce visible toxicity symptoms in wt and <i>aqua1</i> plants even after 2 months of treatment confirming the high tolerance of this poplar species to Cd. Most of the analyzed genes showed in wt plants a quick response in transcription at 1 d of treatment and an adaptation at 60 d. On the contrary, a lower transcriptional response was observed in <i>aqua1</i> plants in concomitance with a higher Cd concentration in medial leaves. Moreover, <i>PaHMA2</i> showed at 1 d an opposite trend within organs since it was up-regulated in root and stem of wt plants and in leaves of <i>aqua1</i> plants. In summary, <i>aqua1</i> overexpression in poplar improved Cd translocation suggesting a lower Cd sensitivity of <i>aqua1</i> plants. This different response might be due to a different transcription of <i>PaNRAMP3</i> genes that were more transcribed in wt line because of the importance of this gene in Cd compartmentalization.