The Amazon is hypothesized to reach an irreversible ‘tipping point’ when deforestation slows the hydrological cycle sufficiently that tropical forest ecosystems cannot be sustained. However, inception of such a tipping point has not been supported by observations and the relevant links between deforestation and atmospheric moisture recycling are poorly understood. Here we show that reduction in evapotranspiration from 20 years of deforestation dried the atmosphere persistently and caused moisture decoupling between the lower and middle troposphere. Increased deforestation exacerbated lower troposphere drying and caused it to penetrate deeper into the middle troposphere in the dry and transition seasons over monsoon forests and savannas. Deforestation induced warming-enhanced buoyant updrafts, elevated hot and dry air, and thereby reduced downward mixing of water supplies from the tropical Atlantic that normally moisten the Amazon forests. The severe atmospheric desiccation in the southern and eastern Amazon cannot be compensated by enhanced water supplies from the Atlantic Ocean, demonstrating an irreversible transition in Amazon hydrological cycle exacerbated by rapid deforestation. The more recent drying through the seasons over rainforests and during the wet season over the transition zones from rainforests to monsoon forests and savannas, however, suggests a window of opportunity for preventing ecosystem collapse with forest conservation.