By interacting with key regulatory proteins such as the pRb family, cyclins, cyclin-dependent kinases and p300/CBP of host cells, adenoviral E1A interferes with various cellular processes to provide a suitable environment for the replication of viruses. E1A may promote DNA synthesis and cell cycle progression, immortalize rodent cells in culture and transform cultured cells in cooperation with E1B, Ras, or other oncoproteins. Both extreme N terminus and conserved region 1 of E1A are required for the immortalization and the transformation of rodent cells, transcriptional repression and specific induction of the expression of cellular genes such as the proliferating cell nuclear antigen (PCNA) and heat shock protein 70 (HSP70). Although the molecular mechanisms of these functions of E1A are not fully understood, it is believed that protein-protein interactions may play essential roles. In this communication, we report that a new set of cellular proteins with apparent molecular weight of 200, 90, 45, 30, and 28 specifically associate with the extreme N terminus of E1A. Further analysis demonstrate that these associations do not depend on E1A's association with p300 or pRB. Neither the 30 kDa nor the 28 kDa polypeptide is identical to Cdc2 or Cdk2. The region of E1A required for the protein interaction is also required for the recently identified N-terminal transactivation activity of E1A. Our observations suggest that in addition to p300/CBP, the new set of cellular proteins may be involved in the functional complexity of the N terminus of E1A, thus predicting a p300/CBP independent pathway.