Microorganism plays a pivotal role in regulating sustainable development of agriculture. The excessive application of nitrogen fertilizer is considered to affect the microbial structure in many agricultural systems. The present study aimed to assess the impacts of nitrogen application rate on microbial diversity, community and functionality in rhizosphere of Tartary buckwheat in short-time. The nitrogen fertilizer was applied at rates of 90 kg (N90), 120 kg (N120) and 150 kg (N150) urea per hectare, respectively. The soil properties were measured chemical analysis and displayed no difference among treatments. Metagenome analysis results showed that the microbial diversity was not affected, but the microbial community and functionality were affected by the nitrogen application rate. According to the Linear discriminant analysis effect size (LEfSe) analysis, 15 taxa were significantly enriched in the N120 and N150 groups, no taxon was enriched in the N90 group. Kyoto Encyclopaedia of Genes and Genomes (KEGG) annotation results revealed that the genes related to butanoate and beta alanine metabolism were significantly enriched in the N90 group, the genes related to thiamine metabolism, lipopolysaccharide biosynthesis and biofilm formation were significantly enriched in the N120 group, and the genes related to neurodegenerative disease was significantly enriched in the N150 group. In conclusion, short-time nitrogen fertilizer application shifted the microbial community structure and functionality.