High temperatures severely affect plant growth and pose a threat to global crop production. Heat causes the accumulation of misfolded proteins in the ER, and triggers the heat shock response (HSR) in the cytosol and the unfolded protein response (UPR) in the ER. Excessive misfolded proteins undergo further degradation through ER associated degradation (ERAD). Although much work has been done on the plant heat stress response, the regulation of ER-localized proteins has not been well studied thus far. We isolated the microsome fraction from heat treated and untreated maize seedlings and performed proteomic and ubtiquitylome analyses. Out of 8306 total proteins detected by proteomics, 1675 proteins were significantly unregulated, and 708 proteins were significantly downregulated. Global ubiquitination analysis discovers 1780 proteins with at least one ubiquitination site in each protein. Motif analysis reveals that alanine and glycine are preferred upstream and downstream of the ubiquitinated lysine. ERAD components are found to be hyper-ubiquitinated after heat treatment, implying the feedback regulation of ERAD activity through protein degradation.