Chicken gastrointestinal microbiota plays important roles in health, productivity, and disease. However, knowledge of the relationship between heat stress and the gut microbial ecosystem of poultry, especially laying hens, is still limited. Here, we aimed to provide important knowledge for heat stress intervention in the egg industry. We performed high-throughput sequencing metagenomics on fecal contents to unravel the microbial taxa and functional capacity of the gut microbiome of caged laying hens under heat stress. Results showed that the fecal microbial communities of laying hens were dominated by Firmicutes, Bacteroidetes, and Proteobacteria phyla. The Firmicutes were significantly decreased, and Bacteroidetes were increased in the fecal microbiota under heat stress. Functional prediction of these changes in microbiota revealed that metabolism-related pathways, including cysteine and methionine metabolism and benzoate degradation, were more abundant. Conversely, retinol metabolism and phenylpropanoid biosynthesis were decreased by heat stress, suggesting differences in metabolism between layers in different temperature environments. Clear contributions were identified between active taxa (genus level) and metabolic pathways, which were associated with the liver and intestinal dysfunction in layers. These data revealed that heat stress induced a significant taxonomic perturbation in the gut microbiome of caged laying hens. This was related to the negative effects of heat stress in poultry and provided important basic knowledge for heat stress intervention.