Endoplasmic reticulum (ER) stress activates PKR‐like ER kinase (PERK), increasing phosphorylation of eukaryotic translation initiation factor 2 (p‐eIF2). PERK is one arm of the tripartite Unfolded Protein Response (UPR), which functions to alleviate protein misfolding in the ER. In this study, mice homozygous for the LoxP allele of PERK and expressing albumin‐driven Cre recombinase (liver‐specific PERK null; AlbCre+) and Cre‐negative controls (AlbCre−) were treated with tunicamycin (Tun; 1mg/kg body weight i.p.), an inhibitor of protein glycosylation, or excipient. Livers were harvested 6h after injection. AlbCre− mice treated with Tun demonstrated increased p‐eIF2 in the liver. In contrast, AlbCre+ mice showed no increase in p‐eIF2. Induction of the UPR was monitored using RT‐PCR to evaluate mRNA splicing of the transcription factor, X‐box binding protein‐1 (XBP‐1), and quantitative Real‐Time RT‐PCR to evaluate mRNA expression of C/EBP homologous protein (CHOP) and the molecular chaperone, BiP/grp78. In AlbCre− livers, Tun increased XBP‐1 mRNA splicing 5‐fold and hepatic expression of BiP and CHOP 35‐fold and 350‐fold, respectively. On the other hand, activation of the UPR in the livers of AlbCre+ mice treated with Tun was significantly dampened. Specifically, Tun induced XBP‐1 mRNA splicing 3‐fold whereas BiP and CHOP mRNA expression increased only 7‐fold and 5‐fold, respectively. In summary, liver‐specific PERK null mice demonstrate impaired or delayed UPR activation following Tun injection. These data suggest that PERK is solely responsible for Tun‐mediated p‐eIF2 and serves an important role in early activation of the UPR to ER stress in liver.