Thin-walled connection structures are commonly used in the hot-end components of aerospace vehicles. Large deflection nonlinear responses and fatigue failure occur due to their discontinuous mass distribution and prominent cross-sectional changes under the action of complex thermal, aerodynamic, and noise loads. A thermoacoustic fatigue test was carried out to obtain the acoustic and vibration responses and fatigue life changes of the connection structure under heat flow conditions in engineering applications. The high-temperature acoustic fatigue test system of aviation thin-walled structures was used, taking the high-temperature alloy thin-walled plate-load-bearing frame bolted connection structure as the research object. As a result, the vibration response and fatigue life under different thermoacoustic loads were obtained. The contact finite element method was used to simulate the connection pre-tightening force, and the coupled finite element/boundary element method was used to calculate the acoustic and vibration response of the heat flow conditions. The changing rules of the frequency response peak value at the critical point of the thin-walled connection structure under the effects of different temperature fields, fluid fields, and sound fields were obtained through the processing and analysis of the calculation results. Considering the structural vibration fatigue damage mechanism, this study employed an improved rainflow counting method to compute the rainflow circulation matrix (RFM) and rainflow damage matrix (RFD) of the vibration stress time history at critical points within the structure framework. Said method was combined with Miner’s linear cumulative damage theory to estimate the fatigue life under various thermal-fluid-acoustic coupled loads. A comprehensive analysis validates the accuracy of the established numerical simulation calculation model in identifying critical connection points within structures subjected to pre-tightening forces. This model effectively characterizes thermal, aerodynamic, and acoustic loads on high-temperature alloy thin-walled-load-bearing frame bolted connection structures. It delineates the relationship between vibration response and fatigue life while assessing the impact of three distinct load parameters.