Ecosystem services are the beneficial goods and services that ecosystems provide to humans. Urbanization is an important feature of human social development. While promoting economic and social development, it also brings about land degradation, resource depletion, environmental pollution and other problems, intensifying the transformation of natural ecosystems into semi-natural and artificial ecosystems, ultimately leading to the loss of ecosystem service functions and declining value. The study of the impact of urbanization on the value of ecosystem services is of critical importance for the conservation of ecosystems and sustainable development. This study examined the spatio-temporal patterns of urbanization’s impacts on ecosystem service value in the Yellow River Basin from the perspective of spatio-temporal heterogeneity. Findings: (1) Both the ecosystem service value (ESV) and urbanization level (UL) in the Yellow River Basin were on the rise on the whole, but they were significantly spatially negatively correlated and mainly characterized by the high–low spatial clustering of “low ESV–high UL” and “high ESV–low UL”. This negative correlation was gradually weakened with the transformation of the urbanization development mode and ecological restoration projects in the Yellow River Basin. (2) The impacts of the five urbanization subsystems on the value of ecosystem services were diverse. Landscape urbanization had a negative impact on the value of ecosystem services in all regions; economic urbanization and innovation urbanization changed from having a negative to a positive impact; and demographic urbanization and social urbanization had both a positive and a negative impact. (3) To promote the coordinated development of ecological environmental protection and urbanization in the YRB, this paper proposes to change the urbanization development model, implement ecological restoration by zoning, and formulate classified development plans. This study compensates for the shortcomings of current studies that ignore the different impacts of urbanization subsystems on ecosystem service value and lack sufficient consideration of the spatio-temporal heterogeneity characteristics of urbanization and ESVs, enriches the theoretical understanding of the interrelationships between natural and human systems in basin areas, and provides a scientific basis for the rational formulation of urban planning and ecological protection policies in the region, which is of great theoretical and practical significance.