Few continental palaeoenvironmental sedimentary sequences from Southern Europe are long enough to span the last interglacial period (Marine Isotopic Stage-MIS 5), the last glacial cycle (MIS 4 to 2) and the Holocene. El Cañizar de Villarquemado (North-Eastern Iberian Peninsula) is an exceptional sedimentary lacustrine sequence spanning the last ca. 135,000 years of environmental change in an area of inland Iberia characterized by Mediterranean climate with strong continentality. We present a multiproxy study which combines palynological, sedimentological and geochemical analyses framed by an independent, robust chronology. Hydrological and climate evolutions were reconstructed by sedimentological and geochemical proxies. Development of wetlands and shallow carbonate lakes support relatively humid conditions during MIS 6, till the onset of MIS 4, and during the Holocene. Palaeohydrological conditions were drier during MIS 5 (dominance of peat environments) than during the Holocene (more frequent carbonate-producing lakes). Sedimentological evidence indicates extremely arid conditions during MIS 3 with greater activity of alluvial fans prograding into the basin. Sedimentary facies variability highlights a large environmental and hydrological variability during MIS 2 and a rapid humidity response to the onset of the Holocene.Compared to classic Mediterranean sites, we found novel pollen assemblages for the end of MIS 6 and MIS 5 indicating that the vegetation cover was essentially represented by sustained high proportions of continentality-adapted taxa dominated by Juniperus during the relatively humid conditions since MIS 6 till the onset of MIS 4. Higher evapotranspiration in inland Iberia would have increased during periods of higher seasonal insolation maxima, impeding soil development and the usual mesophyte expansion during interglacials observed in other Mediterranean areas. Four main periods of forest development occurred in Villarquemado during MIS 5e, MIS 5c, MIS 5a and the Holocene; secondary peaks occurred also during MIS 3. During colder but still relatively humid MIS 4, junipers and Mediterranean taxa disappear but some mesophytes and cold-tolerant species persisted and Pinus became the dominant tree up to modern times. Pollen assemblages and geochemical data variability suggest a dominant control of seasonality and the impact of North Atlantic dynamics both during MIS 5 (cold events C18-C24) and full glacial conditions (HE and D-O interstadials). At millennial scales, steppe herbaceous assemblages dominated during the extremely arid conditions of MIS 3 and pines and steppe taxa during glacial period MIS 2. Villarquemado sequence demonstrates that the resilient behaviour of conifers in continental areas of inland Southern European regions is key to understand the glacial–interglacial vegetation evolution and to evaluate scenarios for potential impacts of global change.