Abstract: Background The study investigated the vertical distribution patterns and driving mechanisms of Soil microbial communities and their nutrient metabolic functions during forest restoration. Methods This study investigated a restoration sequence in a subtropical region, consisting of early-successional coniferous forest dominated by Pinus massoniana, conifer-broadleaf mixed forest, secondary evergreen broadleaf forest, and mature evergreen broadleaf forest. Soil samples were collected from the topsoil (0~10, 10~20 cm), subsurface soil (20~40, 40~60 cm), and deep soil (60~80, 80~100 cm) layers. By analyzing microbial phospholipid fatty acids (PLFAs), enzyme activities linked to carbon (C), nitrogen (N), and phosphorus (P) cycling, and microbial carbon use efficiency (CUE), to explored the microboal characteristics and their responses to environmental factors across soil layers. Results The results showed that: (1) forest restoration significantly increased total microbial PLFAs and the abundance of each microbial group across soil layers, though increases showed a delayed response in deep soil. Concurrently, restoration significantly lowered the ratio of fungal to bacterial biomass in both topsoil and subsurface soil. (2) Forest restoration significantly decreased β-glucosidase activity in all layers, while markedly increasing activities of other hydrolases and oxidases, alleviating microbial C limitation and improving microbial carbon use efficiency. (3) The drivers of microbial community assembly exhibited vertical stratification: the topsoil community was primarily governed by mineral N; the subsurface community by soluble organic N; and the deep soil community mainly by environmental filtering related to C availability and pH. (4) Enzyme activities were primarily regulated by soil C, N, and P contents and their stoichiometric ratios, with a shift in the primary regulation from the N:P ratio in topsoil C:P ratio and soil organic carbon in subsurface and deep soils. Conclusions The study demonstrates that regulating aboveground vegetation recovery processes and the improvements in soil nutrient availability and the optimization of C, N, and P stoichiometric ratios during subtropical forest restoration drive shifts in microbial metabolic strategies, thereby differentially regulating ecological functions across soil layers. Future subtropical forest restoration and management should emphasize whloe-profile soil carbon stabilization and microbe-mediated carbon cycling processes.