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Plastic film mulching has been widely applied to improve crop productivity of full-season cotton (Gossypium hirsutum L.). However, in a previous study, plastic mulching did not increase seed cotton yield of short-season cotton compared with non-mulching. The aim of this study was to determine why plastic mulching fails to increase seed cotton yield of short-season cotton. A short-season cultivar Lumian 532 was planted with or without plastic mulching at an experimental station in Linqing in the Yellow River valley of China in 2020 and 2021. Yield, yield components, leaf senescence, photoassimilate accumulation and partitioning, photohormone contents, expression of senescence-related genes, and root traits were examined each year. Seed cotton yield produced with plastic mulching was not significantly different from that produced with non-mulching in short-season cotton. Although number and size of early- and mid-season bolls were significantly greater with plastic mulching than with non-mulching, plastic mulching decreased number and size of late-season bolls by 34.0% and 8.8%, respectively. At the initial boll opening stage, photosynthetic rate and chlorophyll content in main-stem functional leaves were 36.2% and 24.1% lower, respectively, with plastic mulching than with non-mulching, but malondialdehyde content was 34.2% higher, suggesting that plastic mulching accelerated late-season leaf senescence. Canopy photosynthesis and photoassimilate partitioning to reproductive organs at the initial boll-opening stage were 68.1% and 9.4% lower, respectively, with plastic mulching than with non-mulching. Compared with non-mulching, plastic mulching increased abscisic acid and methyl jasmonate contents but decreased those of cytokinins content because of differential expression of hormone metabolism-related genes in late-season main-stem leaves. Plastic mulching also increased root distribution in the shallow soil layer and lowered root vigor in the late season, compared with non-mulching. Overall, plastic mulching negatively affected root distribution and function and altered expression of senescence-related genes and hormone metabolism-related genes to affect hormone balance, which led to accelerated late-season leaf senescence. Because of late-season leaf senescence, canopy photosynthesis and photoassimilate partitioning to cotton bolls were suppressed with plastic mulching, which led to fewer and smaller late-season bolls than those with non-mulching. Thus, plastic mulching failed to improve the seed cotton yield. This study provides a clear explanation of how plastic mulching causes late-season leaf senescence and further demonstrates that plastic film mulching is not required for planting short-season cotton in the Yellow River valley of China or in other cotton growing areas with similar ecology. • Plastic mulching did not increase yield compared with non-mulching because of fewer and smaller late-season bolls in short-season cotton. • Reduced number and size of late-season cotton bolls was attributed to accelerated late-season leaf senescence in short-season cotton. • Accelerated leaf senescence was due to shallow root distribution and weak root vigor, and differential expression of senescence-related genes. [ABSTRACT FROM AUTHOR]