Huayang Lake Complex is a typical wetland system in the middle and lower reaches of the Yangtze River, with core ecological functions including waterbird protection, hydrological connectivity and water quality purification, as well as social production guarantee functions such as water supply and flood control. Current water level regulation focuses on single factors and lacks an annual dynamic process curve, which is difficult to fully support the stable exertion of its core ecological functions. Based on nearly 50 years of hydrological, topographic, water quality and habitat data, this study establishes a multi-factor collaborative calculation framework of “hydrology–morphology–water quality–habitat” by using the IHA-RVA method, ArcGIS spatial analysis, hydrodynamic-water quality model and habitat suitability area method, and proposes an annual optimal ecological water level hydrograph to satisfy the hydrological stability and ecological health of the lake complex. The results show that the optimal ecological water level ranges from 11.86 m (April) to 13.53 m (September), with a threshold range of 11.56 m (March) to 14.87 m (August). It presents a temporal distribution characteristic of "convergence in winter and spring, expansion in summer and autumn", and the seasonal water levels adapt to the demands of biological habitats, hydrological rhythms and water quality improvement. This scheme can guarantee the overwintering of key waterbirds, inhabitation of aquatic organisms and water quality purification, and provide a scientific reference for ecological water level regulation of similar gate-controlled lakes or lake groups in the middle and lower reaches of the Yangtze River.