The urban heat island effect is a prevalent urban concern, and studies on its influencing factors are essential to mitigate its negative effects and promote sustainable urban development. Although many studies have revealed the impact of land cover change on land surface temperature (LST) during urbanization, most have focused only on built-up areas without considering the relationship between urban internal functional structure and LST. To clarify the influence of changes in urban functional zones (UFZs) on LST, focusing on the downtown area of Nanjing City, China, this study proposed an effective framework using multi-source geospatial data for qualitative and quantitative analysis. Specifically, very high spatial resolution images and point of interest data were jointly used to generate UFZ maps using an advanced deep learning approach, and LST distribution maps were retrieved from Landsat 8 images using the radiative transfer equation. The UFZ and LST maps were then overlaid and analyzed to reveal the impact of UFZ change on LST. The results showed that in 2013, 2018, and 2022, the residential region constituted the largest proportion of area, comprising 23.52%, 24.52%, and 27.28%, respectively. The areas of unused region and transport region experienced the most rapid decrease and increase of 6.08% and 4.05% from 2013 to 2022, respectively. Furthermore, the area of sub-high temperature zone decreased the most, by 3.47%, whereas that of the high-temperature zone increased the most, by 6.05%. In the study area, water and green space maintained relatively low LSTs, whereas residential region and unused region exhibited higher LSTs. From 2013 to 2022, UFZ changes contributed to a 0.71°C increase in LST. Transfers from green space, water, commercial region, transport region, and industrial region mostly resulted in a higher LST, which was associated with a decrease in fractional vegetation cover (FVC). Transfers from unused region, residential region, and public service region mostly contributed to reducing LST, which was caused by the increase in FVC. The findings and the proposed framework can provide reliable references for urban managers to optimize urban spatial layout and promote sustainable urban development.