Despite significant advances in targeted therapies against the hyperactivated BRAFV600/MEK pathway for patients with unresectable metastatic melanoma, acquired resistance remains an unsolved clinical problem. In this study, we focused on melanoma cells resistant to trametinib, an agent broadly used in combination therapies. Molecular and cellular changes were assessed during alternating periods of trametinib withdrawal and rechallenge in trametinib-resistant cell lines displaying either a differentiation phenotype (MITFhigh/NGFRlow) or neural crest stem-like dedifferentiation phenotype (NGFRhigh/MITFlow). Neither drug withdrawal nor drug rechallenge induced cell death, and instead of loss of fitness, trametinib-resistant melanoma cells adapted to altered conditions by phenotype switching. In resistant cells displaying a differentiation phenotype, trametinib withdrawal markedly decreased MITF level and activity, which was associated with reduced cell proliferation capacity, and induced stemness assessed as NGFR-positive cells and senescence features, including IL-8 expression and secretion. All these changes could be reversed by trametinib re-exposure, which emphasizes melanoma cell plasticity. Trametinib-resistant cells displaying a dedifferentiation phenotype were less responsive presumably due to the already low level of MITF, a master regulator of the melanoma phenotype. Considering new directions of the development of anti-melanoma treatment, our study suggests that the phenotype of melanomas resistant to targeted therapy might be a crucial determinant of the selection of second-line therapy for melanoma patients.