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Grasslands have an important global role in biological methane (CH 4) oxidation and therefore it is crucial to understand the key factors determining the assembly of high-affinity methanotroph populations and explore mechanisms that regulate CH 4 oxidation activity in these ecosystems. This study explored the linkage between high-affinity methanotroph community assembly processes and CH 4 oxidation potential across three grassland types in Inner Mongolia with or without livestock grazing. We found the largest proportion of deterministic selection in temperate desert grasslands, but it was increasingly structured by stochastic dispersal in temperate meadow and temperate typical grasslands. The distance decay relationship (DDR) and variation partitioning analysis (VPA) showed the strongest relationship and highest proportion of environmental factors in desert grasslands, followed by meadow and typical grasslands. Mean annual precipitation (MAP), environmental factors (moisture content, soil organic matter), and microbial traits (abundance, diversity and normalized stochasticity ratio) were significantly related with CH 4 oxidation activity. Multiple statistical analyses consistently indicated the key role of MAP in shaping high-affinity methanotroph community diversity, composition, co-occurrence networks, assembly processes and CH 4 oxidation activity. We found an association of increasing stochastic assembly and high-affinity methanotroph diversity with decreasing CH 4 oxidation potential. This study revealed the mechanisms governing high-affinity methanotroph community assembly processes and integrates high-affinity methanotroph community assembly with CH 4 oxidation activity in grassland soils. [Display omitted] ● The proportion of determinism was highest in desert than other grasslands. ● MAP was key factor regulating methanotrophs community and their activity. ● Stochastic assembly processes are dominant in high-diversity communities. ● CH 4 oxidation potential decreased with the increase of stochastic assembly. [ABSTRACT FROM AUTHOR]