Intensified human activities and global climate change have led to the deterioration of the environment, degradation of ecosystem functioning, and loss of biodiversity in lakes in many parts of China over the past few decades. To date, lakes in Yunnan have been adversely affected by the synergic effects of multiple environmental stressors, such as eutrophication, climate change, and hydrological regulation. However, previous studies have mainly focused on the ecological impacts of a single environmental stressor. In this study, paleolimnological methods in combination with long-term monitoring data were used to understand the patterns of diatom community responses to eutrophication, climate change, and hydrological regulation in Xingyun Lake. First, the history of diatom community shift and limnological changes were reconstructed for the past two centuries. In addition, key environmental stressors and their strengths in driving diatom communities were identified by selecting minimum adequate models and using variance partitioning. Our sediment pigment results showed a clear trajectory of lake eutrophication during the last century, particularly since the 1980s. The diatom community changed significantly with an increase in lake productivity (i.e., sediment Chl-a concentrations). Principal component analysis and correlation analysis showed that lake eutrophication was the major driver that affected diatom community change (r = -0.63, P < 0.001). In addition, the minimum adequate models showed that lake production (18.8%) and hydrodynamics (2.9%) were the major drivers of diatom community changes over the last 200 years (i.e., core length of 38 cm). Since 1951, the major drivers were lake production (31.4%) and temperature (26.8%), and both factors showed a strong interaction while driving the community shift (16.2%), suggesting that increasing temperatures also contributed significantly to the increase in lake production during the past few decades. Our results suggested that nutrient enrichment was the most significant factor in driving the shift in the diatom community in Xingyun Lake during the last two centuries. Global warming and anthropogenic activities, which included hydrological fluctuation and the connectivity between Xingyun Lake and the neighboring Fuxian Lake, also played important roles, but to a lesser degree than lake eutrophication. Overall, the sediment-based patterns of lake ecological changes can help us identify multiple environmental stressors and quantify their ecological impacts. These can be further used to facilitate the ecological restoration of polluted lakes and conservation of lake ecosystems in Yunnan.