Abstract: To investigate the response differences of soybean varieties with varying shade tolerance to shading environments, the study used the strong shade-tolerant variety 'Changdou 33' (cd33) and the weak shade-tolerant variety 'Changdou 31' (cd31) as materials. The agronomic traits, leaf tissue structure, mesophyll ultrastructure, physiological and biochemical indicators, endogenous hormone content, and transcriptomics of soybean under natural light and shading conditions were analyzed. The results showed that: 1) Under shading stress, both cd31 and cd33 exhibited increased plant height, reduced stem diameter, and decreased above-ground dry matter accumulation; however, the changes in cd33 were less pronounced than those in cd31. 2) Soybean improved light utilization efficiency by reducing leaf thickness and palisade tissue thickness while increasing mesophyll cell area, perimeter, and chloroplast area. Cd33 exhibited a more intact cellular structure, which mitigated the impact of shading on photosynthesis. 3) Physiological and biochemical results indicated that under shading, SOD and POD activities increased, CAT activity significantly decreased, soluble sugar and proline content increased, soluble protein content decreased, MDA content declined, and the levels of auxin, gibberellin, and abscisic acid decreased. This suggests that soybean adapts to the environment by regulating the antioxidant system, osmotic adjustment substances, and endogenous hormones. Additionally, the abscisic acid and gibberellin content in cd33 leaves were lower than in cd31, which may inhibit plant height and contribute to stronger lodging resistance. 4) Transcriptomic analysis further revealed that the strong shade-tolerant variety had significantly more differentially expressed genes (5,783) under shading conditions compared to the weak shade-tolerant variety (2,319). These differentially expressed genes were mainly enriched in pathways such as hydrogen peroxide response, protein kinase activity, plant hormone signal transduction, and phenylpropanoid biosynthesis. In conclusion, the strong shade-tolerant variety demonstrated greater shade tolerance by activating more gene expression and metabolic pathways, maintaining cellular stability and light energy utilization efficiency.