Hiroaki Kiyokawa, MD, PhD Associate Professor Molecular Pharmacology & Biological Chemistry Roles of cell cycle-regulatory proteins in differentiation, senescence and tumorigenesis Curricula: Cancer Biology Molecular Biology and Genetics E-mail:   kiyokawa@northwestern.edu

(1) Roles of cell cycle-regulatory proteins in differentiation, senescence and tumorigenesis

(2) Cell cycle control in endocrine and reproductive organs

Molecular events determining cell fate are initiated during the G1 phase of the cell cycle. G1 progression is governed by the kinase activity of cyclin D/CDK4 and cyclin E/CDK2. Generally, when the activity of cyclin D/CDK4 is high, the cell will continue proliferation, whereas inactivation of this kinase complex will recruit the cell to quiescence and differentiation. CDK inhibitor proteins, including p16, p21 and p27, function to inactivate these CDKs.

We have been investigating the G1-CDK regulatory system, by generating novel "knockout" mice. p27-knockout mice display gigantism with multi-organ hyperplasia, suggesting that p27 limits the replicative capacity of differentiating cells. p27-knockout mice also develop pituitary adenomas, indicating that p27 has a tumor suppressor function. In contrast, CDK4-knockout mice exhibit growth retardation and infertility associated with hypoplastic gonads. These apparently opposite phenotypes imply that p27 and CDK4 counteract each other in developmental control of cell fate. p27-knockout mice exhibit high susceptibility to carcinogen- or g -irradiation-induced tumorigenesis, while CDK4-knockout mice are resistant to carcinogens. Thus, activation of the G1-CDKs forms one of the rate-limiting steps for carcinogenesis.

Our recently investigations on CDK4-null mice revealed unique regulation of cell cycle progression in neuroendocrine tissues such as pancreatic islets and pituitary glands.  These cell types undergo extremely slow proliferation with full characteristics of differentiation, which can speed up in response to altered extracellular environments, e.g., chronic hyperglycemia and pregnancy-associated hyperestrogenemia.  We have shown that in this particular type of cell cycle, Cdk4 plays an indispensable and rate-limiting role.  Efforts are ongoing to further characterize the “adaptive” cell cycle in neuroendocrine tissues.

Furthermore, we have demonstrated that CDC25A phosphatase, which activates CDK2 and CDK1, is an oncogene that plays a rate-limiting role in initiation and progression of various tumors, including breast cancer.  CDC25A protein is overexpressed in a majority of breast cancer tissues at early stages.  Our data using in vivo mouse models and in vitro biochemical systems indicate that proteosome-mediated degradation of CDC25A protein is impaired in cancer cells, and stabilization of the protein can result in not only accelerated cell proliferation but also increased cell survival under stressful conditions.

We are currently investigating roles of the cell cycle machinery in differentiation, tumorigenesis and apoptosis, by combinations of mouse models and molecular analyses.

Publications:

Tsutsui T, Hesabi B, Moons D, Pandolfi PP, Hansel K, Koff A, Kiyokawa H. Targeted disruption of CDK4 delays cell cycle entry with enhanced p27Kip1 activity.  Mol Cell Biol 19: 7011-7019, 1999.

Zou X, Tsutsui T, Ray D, Blomquist JF, Ichijo H, Ucker DS, Kiyokawa H.  Cell cycle-regulatory CDC25A phosphatase inhibits apoptosis signal-regulating kinase 1.  Mol Cell Biol 21: 4818-4828, 2001.

Moons DS, Jirawatnotai S, Parlow AF, Gibori G, Kineman RD, Kiyokawa H. Pituitary hypoplasia and lactotroph dysfunction in mice deficient for Cyclin-dependent kinase-4 (Cdk4).  Endocrinology 143, 3001-3008, 2002.

Zou X, Ray D, Aziyu A, Christov K, Boiko A, Gudkov A, Kiyokawa H.  Cdk4 disruption renders primary mouse cells resistant to oncogenic transformation leading to Arf/p53-independent senescence.  Genes Dev 16: 2923-2934, 2002.

Jirawatnotai S, Moons DS, Stocco CO, Franks R, Hales DB, Gibori G, Kiyokawa H.  The Cdk inhibitors p21^Cip1/Waf1 and p27^Kip1 cooprerate to restrict proliferative life span in differentiating ovarian cells.  J Biol Chem, 278: 17021-17027, 2003.

Jirawatnotai S, Aziyu A, Osmundson E, Moons DS, Zou X, Kineman R, Kiyokawa H.  Cdk4 is indispensable specifically for postnatal proliferation of somato/lactotrophs of the murine pituitary.   J Biol Chem, 279: 51100-51106, 2004.

Ray D, Terao Y, Donzelli M, Ghosh AK, Tsutsui T, Zou X, Varga J, Draetta GF, Kiyokawa H.  The Smad3-mediated signaling of TGF-beta facilitates β-TrCP-dependent degradation of Cdc25A.  Mol Cell Biol, 25:3338-47, 2005

Aleem E, Kiyokawa H, Kaldis P.  Cdk2/cyclin E complexes regulate the G1/S phase transition.  Nat Cell Biol, 7: 831-836, 2005.

Ray D, Osmundson EC, Kiyokawa H. Constitutive and UV-induced degradation of fibronectin is a ubiquitination-dependent process controlled by β-TrCP.  J Biol Chem, 281:23060-23065, 2006.

Ray D, Terao Y, Fuhrken PG, Ma ZQ, DeMayo FJ, Christov KT, Heerema NA, Franks R, Tsai SY, Papoutsakis ET, Kiyokawa H. Deregulated CDC25A expression promotes mammary tumorigenesis with genomic instability.  Cancer Res, 67: 984-991, 2007.