Chronic ethanol ingestion leads to inhibition of proteasomal activity. As a consequence, proteins accumulate in liver cells. Cytokeratin accumulation as seen in alcoholic hepatitis could lead to the formation of Mallory bodies. In order to study the phenomenon of cytokeratin accumulation in liver cells, rats were fed ethanol or dextrose for 1 month and some were given the proteasome inhibitor, PS-341, to augment the inhibitory effect of ethanol feeding. This was done to study the involvement of proteasome inhibition in the process of cytokeratin accumulation. There was a marked increase in the accumulation of polyubiquitinated proteins, and heat shock proteins (hsp) 25 and 70 in the liver of rats treated with PS-341. Similarly, cytokeratin-8 (CK-8) levels were markedly increased in the liver homogenates of rats fed ethanol when given PS-341. When normal mouse cultured hepatocytes were transfected with cytokeratin-18 (CK-18) tagged with red fluorescent protein (RFP), CK-18 aggresomes formed because proteasome was overloaded. These data provide new evidence that proteasome inhibition is involved in cytokeratin accumulation, when aggresomes are formed in tissue culture. Accumulation of cytokeratin in this way may ultimately lead to Mallory body formation as seen in alcoholic hepatitis.