Marine heatwaves (MHWs) are becoming increasingly common, with devastating ecosystem impacts. However, MHW understanding has almost exclusively relied on sea surface temperature with limited knowledge about their subsurface characteristics. Here we estimate global MHWs from the surface to 2,000 m depth, covering the period 1993–2019, and explore biodiversity exposure to their effects. We find that MHWs are typically more intense in the subsurface at 50–200 m and their duration increases up to twofold with depth, although with large spatial variability linked to different oceanographic conditions. Cumulative intensity (a thermal stress proxy) was highest in the upper 250 m, exposing subsurface biodiversity to MHW effects. This can be particularly concerning for up to 22% of the ocean, where high cumulative intensity overlapped the warm range edge of species distributions, thus being more sensitive to thermal stress. Subsurface MHWs can hence drive biodiversity patterns, with consequent effects on ecological interactions and ecosystem processes.