The objective of this study was to elucidate the immune system response to heat stress in chickens. In this study, mRNA-seq was conducted on the spleen and bursa of experimental chickens, six differentially expressed genes associated with immunity were present in the spleen following immunization. Following exposure to heat stress, 15 differentially expressed genes related to immune and heat shock proteins were identified. Furthermore, the expression levels of DUSP1 and HSPA5 were significantly lower in the non-stressed group. With regard to the mechanism, overexpression of DUSP1 or HSPA5 resulted in no significant difference in MHC-I, MHC-II, and CD80 mRNA expression. However, following stimulation with LPS, mRNA expression of MHC-II, CD80, CD86, CD1C, IL1B, and TLR4 was significantly increased. Furthermore, the enhancement was observed to occur at an earlier stage than when LPS was stimulated alone, thereby facilitating the recognition of LPS by HD11. Following the inhibition of DUSP1 or HSPA5 and the stimulation of LPS, no significant alterations were detected. However, CD1C expression was notably diminished. In conclusion, DUSP1 and HSPA5 have been demonstrated to play important roles in immunity to heat stress by affecting antigen presentation. The present study provides a theoretical basis for the regulation mechanism of disease resistance in poultry.