Histone acetylation is an essential mechanism regulating chromatin conformation and gene transcription during cell homeostasis, proliferation and differentiation. Using inducible cre-recombinase and CRISPR/Cas9 deletion of the gene encoding the histone lysine acetyltransferase TIP60 (KAT5/HTATIP) we investigated the biological role of TIP60 (Kamine et al., 1996) in human and mouse cells and mouse embryos. We found that TIP60 depletion causes complete cell growth arrest in human HEK293 and U2OS cells and mouse fibroblasts. Remarkably, the growth arrest was independent of the cell cycle regulatory proteins INK4A-ARF and p53. Tip60 null cells display cell cycle arrest in either G1 or M phase and an increase in endoreplication, but no change in cell survival. Comprehensive western blot analysis revealed that TIP60 is essential for over 85% of pan-H2AZ acetylation, and solely responsible for acetylating H2AZK7. H2AZ acetylation has been correlated with transcriptionally active genes (Valdes-Mora et al., 2012). RNA -sequencing showed that the expression levels 6236 human and 8848 mouse genes were dependent on TIP60, supporting a role for TIP60 as a core transcriptional co-regulator. In conclusion, we propose a primary role of TIP60 in H2AZK7 acetylation and an essential role in cell proliferation. The ability to arrest growth independently of the major tumour suppressors p53 and INK4A-ARF flags TIP60 a potential anti-cancer drug target, as the genes encoding these tumour suppressors are commonly mutated or deleted in cancer.