CDC37, an essential gene in Saccharomyces cerevisiae, interacts genetically with multiple protein kinases and is required for production of Cdc28p/cyclin complexes through an unknown mechanism. We have identified mammalian p50Cdc37 as a protein kinase-targeting subunit of the molecular chaperone Hsp90. Previously, p50 was observed in complexes with pp60v-src and Raf-1, but its identity and function have remained elusive. In mouse fibroblasts, a primary target of Cdc37 is Cdk4. This kinase is activated by D-type cyclins and functions in passage through G1. In insect cells, Cdc37 is sufficient to target Hsp90 to Cdk4 and both in vitro and in vivo, Cdc37/Hsp90 associates preferentially with the fraction of Cdk4 not bound to D-type cyclins. Cdc37 is coexpressed with cyclin Dl in cells undergoing programmed proliferation in vivo, consistent with a positive role in cell cycle progression. Pharmacological inactivation of Cdc37/Hsp90 function decreases the half-life of newly synthesized Cdk4, indicating a role for Cdc37/Hsp90 in Cdk4 stabilization. This study suggests a general role for p50Cdc37 in signaling pathways dependent on intrinsically unstable protein kinases and reveals a previously unrecognized chaperone-dependent step in the production of Cdk4/cyclin D complexes.

The Cdc37 gene encodes a 50 kDa protein which targets intrinsically unstable oncoprotein kinases such as Cdk4, Raf-1, and src to the molecular chaperone Hsp90. This activity is thought to play an important role in the establishment of signaling pathways controlling cell proliferation. The budding yeast Cdc37 homolog is required for cell division and mammalian Cdc37 is expressed in proliferative zones during embryonic development and in adult tissues, consistent with a positive role in proliferation. Here we report that human prostatic tumors, neoplasias and certain pre-malignant lesions display increased Cdc37 expression, suggesting an important and early role for Cdc37 in prostatic transformation. To test the consequences of increased Cdc37 levels, transgenic mice expressing Cdc37 in the prostate were generated. These mice displayed a wide range of growth-related abnormalities including prostatic epithelial cell hyperplasia and dysplasia. These data suggest that the expression of Cdc37 may promote inappropriate proliferation and may be an important early step in the development of human prostate cancer.

CDC37 encodes a 50-kDa protein that targets intrinsically unstable oncoprotein kinases including Cdk4, Raf-1, and v-src to the molecular chaperone Hsp90, an interaction that is thought to be important for the establishment of signaling pathways. CDC37 is required for proliferation in budding yeast and is coexpressed with cyclin D1 in proliferative zones during mouse development, a finding consistent with a positive role in cell proliferation. CDC37 expression may not only be required to support proliferation in cells that are developmentally programmed to proliferate but may also be required in cells that are inappropriately induced to initiate proliferation by oncogenes. Here we report that mouse mammary tumor virus (MMTV)-CDC37 transgenic mice develop mammary gland tumors at a rate comparable to that observed previously in MMTVcyclin D1 mice. Moreover, CDC37 was found to collaborate with MMTV-c-myc in the transformation of multiple tissues, including mammary and salivary glands in females and testis in males, and also collaborates with cyclin D1 to transform the female mammary gland. These data indicate that CDC37 can function as an oncogene in mice and suggests that the establishment of protein kinase pathways mediated by Cdc37-Hsp90 can be a rate-limiting event in epithelial cell transformation.

It has long been known that prolonged culture or serial transplantation leads to the loss of hematopoietic stem cells (HSCs); however, the mechanisms for this loss are not well understood. We hypothesized that expression of p16 Ink4a or p19 Arf or both may play a role in the loss of HSCs during conditions of enhanced proliferation, either in vitro or in vivo. Arf was not expressed in freshly isolated HSCs from adult mice but was induced in phenotypically primitive cells after 10 to 12 days in culture. When cultured bone marrow cells from either Arf ؊/؊ or Ink4a-Arf ؊/؊ mice were compared to wild-type cells in a competitive repopulation assay, no significant differences in HSC activity were seen. We then evaluated the role of p19 Arf IntroductionRapid turnover of mature hematopoietic cells requires production of tens of billions of cells every day during the life span of mammals. Hierarchical organization of the hematopoietic system helps to ensure the lifelong production of blood cells of different lineages. Hematopoietic stem cells (HSCs) have the ability to self-renew, differentiate into all hematopoietic lineages, and repopulate lethally irradiated hosts. Proliferative activity of HSCs is tightly regulated by incompletely understood mechanisms. HSCs are relatively quiescent and generally cytokine-resistant, with a long life span, whereas more committed progeny have a robust proliferation potential, are highly cytokine-responsive, and have a more limited life span. Although the low proliferation potential of HSCs may play a role in maintenance of HSCs over time, it is an obstacle in gene therapy applications and in clinical strategies to expand HSCs in vitro. 1 Several candidate genes have recently been implicated in the HSC self-renewal process. It has recently been demonstrated that HSC entry into the cell cycle is regulated by a cyclin-dependent kinase inhibitor (CKI), p21 Cip1 , whereas proliferation of progenitors is regulated by another cell-cycle inhibitor, p27 Kip1 . 2,3 Deletion of an early G 1 -phase CKI, p18(INK4C), results in increased self-renewal of the primitive hematopoietic cells and improved long-term engraftment. 4 Low levels of telomerase in HSCs have been implicated in the replicative senescence of HSCs 5 ; however, overexpression of TERT, a catalytic reverse transcriptase component of telomerase, does not lead to enhancement of HSC self-renewal. 6 Overexpression of the HOXB4 homeodomain transcription factor is able to induce expansion of HSCs both in vivo 7 and in vitro. 8 Overexpression of P-glycoprotein can also cause an HSC expansion, later followed by development of a myeloproliferative disorder. 9,10 HSC expansion was also noted in murine HSCs expressing the acute myeloid leukemia 1-eight-twenty-one (AML1-ETO) fusion protein, a product of a chromosomal translocation common in human AML. 11,12 Bmi1, a polycomb group repressor, is essential for the selfrenewal of neuronal and hematopoietic stem cells. Bmi1 is expressed in stem cells of neuronal and hematopoietic origin, and deletion...