The T197A knock-in model of CDKN1B gene to study the effects of p27 restoration in vivo

The CDK inhibitor, p27kip1, encoded by the Cdkn1b gene can negatively modulate cell proliferation. The control of p27 activity during the cell cycle is regulated at multiple levels, including transcription, translation, and protein stability. The last residue of p27 (threonine 198 in human, threonine 197 in mouse) is involved in the control of protein stability. We have generated a murine knock-in model (Cdkn1bT197A) in which threonine 197 is replaced by alanine, which renders p27 protein highly unstable due to a high rate of proteasomal degradation. Expectedly, Cdkn1bT197A/T197A mice present with increased body size and weight, organomegaly, and multiple organ hyperplasia, similar to what is observed in Cdkn1bKO/KO mice. We investigated the effects exerted by the restoration of normal levels of p27 protein in the tissue of Cdkn1bT197A/T197A mice. We found that proteasome inhibition with bortezomib rescues the hyperplasia induced by the lack of p27 expression in Cdkn1bT197A/T197A but not in Cdkn1bKO/KO mice. However, BAY 11-7082, a proteasome inhibitor that stabilizes IκB but not p27, fails to rescue hyperplasia in Cdkn1bT197A/T197A mice. Bortezomib increases p27 half-life and reduces the proliferation in MEFs derived from Cdkn1bT197A/T197A but not from Cdkn1bWT/WT mice, whereas BAY 11-7082 had no effect on the protein levels of p27 and on the proliferation rate of Cdkn1bT197A/T197A MEFs.The results presented here demonstrate that Cdkn1bT197A/T197A mice represent an attractive in vivo model to investigate whether the targeting of p27 degradation machinery might prove beneficial in the treatment of a variety of human proliferative disorders caused by increased turnover of p27 protein.