Anoxic Fibroblasts Activate a Replication Checkpoint That Is Bypassed By E1a

Abstract

Little is known about cell cycle regulation in hypoxic cells, despite its significance. We utilized an experimentally tractable model to study the proliferative responses of rat fibroblasts when rendered hypoxic (0.5% oxygen) or anoxic (<0.01% oxygen). Hypoxic cells underwent G 1 arrest, whereas anoxic cells also demonstrated S-phase arrest due to suppression of DNA initiation. Upon reoxygenation, only those cells arrested in G 1 were able to resume proliferation. The oncoprotein E1a induced p53-independent apoptosis in anoxic cells, which when suppressed by Bcl-2 permitted proliferation despite anoxia. E1a expression led to marked increases in the transcription factor E2F, and overexpression of E2F-1 allowed proliferation in hypoxic cells, although it had minimal effect on the anoxic suppression of DNA initiation. We thus demonstrate two distinct cell cycle responses to low oxygen and suggest that alterations that lead to increased E2F can overcome hypoxic G 1 arrest but that additional alterations, promoted by E1a expression, are necessary for neoplastic cells to proliferate despite anoxia.Cellular hypoxia is common in many physiological and pathophysiological states, including cancer. Poor and disordered vascularization and rapid tumor cell proliferation lead to areas of significant hypoxia in the tumor microenvironment. Direct measurements of oxygen tension reveal that oxygenation may vary widely in tumors, with some areas approaching anoxia (55). Hypoxic tumors are poorly responsive to radiation and chemotherapy and appear to be more aggressive than nonhypoxic tumors (30). This may be partly related to the observation that some oncogenes, such as c-myc, selectively promote apoptosis in hypoxic p53 wild-type cells; thus, hypoxia can select for cells with mutant p53 (24, 25, 52). The role of hypoxia in tumor proliferation may also have important consequences in therapy resistance and tumor progression. However, the regulation of growth in hypoxia, and how oncogenes may affect this proliferation, is still poorly understood despite its importance in tumor biology and cancer treatment.Normal cell cycle progression is regulated by the coordinated actions of cyclins, cyclin-dependent kinases (CDKs), cyclin-dependent kinase inhibitors (CDKIs), and the E2F family of transcription factors (22). Exit from G 1 is mediated by CDK-mediated phosphorylation of the retinoblastoma protein (Rb) and its related pocket proteins, including p107. This phosphorylation releases members of the E2F family, which may then promote the transcription of genes necessary for S phase, including cyclin E enzymes necessary for deoxynucleotide synthesis and members of the DNA initiation complex, CDC6 and MCM6 (5, 11, 40). CDK2-mediated phosphorylation is important for other steps required for DNA synthesis, including the assembly and activation of the DNA initiation complex (11,12,40). CDK2 activity is dependent on the formation of cyclin A or cyclin E complexes and can be inhibited by the CDKIs p27 and p21. Many neoplasms have deregulated cel...

Hypoxia reduces adenoviral replication in cancer cells by downregulation of viral protein expression

“…5,26 Further, it has been previously demonstrated that hypoxia causes cell cycle growth arrest in G1 in nontransformed cells. [16][17][18] Our data, however, show that the reduced viral yield under hypoxic conditions was not due to cell cycle arrest of tumor cells. Although as anticipated, hypoxia was able to growth arrest normal HUVEC, hypoxia did not growth arrest the cancer cells.…”

“…[16][17][18] Although the exact mechanism is still debated, hypoxia generally reduces the activity of cyclin-dependent kinases, which leads to hypophosphorylation of the Rb protein and growth arrest in G1. [16][17][18] Moreover, the presence of cells in S-phase may be important to enable adenoviral replication. 19,20 To Figure 1 HIF-1a is expressed under hypoxic conditions.…”

Hypoxic Regulation of Id-1 and Activation of the Unfolded Protein Response Are Aberrant in Neuroblastoma

“…For example, most normal cells and many neoplastic cells undergo a growth arrest when hypoxic, whereas some stem cells and neoplastic cells continue to proliferate under hypoxic conditions (7,8). Studies have also suggested that hemangioblasts, renal tubular cells, and embryonic stem cells all differentiate when hypoxic, while hypoxic adipocytes and hematopoietic stem cells are resistant to differentiation (9 -12).…”

Staphylococcus aureus Induces Hypoxia and Cellular Damage in Porcine Dermal Explants

“…For example, fibroblasts can replicate at oxygen levels below 25 mm Hg (ϳ1.13 mg/liter), but their capacity for hydroxylation of proline and lysine, which is required for collagen synthesis, is impaired. Cell replication is arrested at oxygen levels below 0.5% (approximately 0.16 mg/liter) (17). Furthermore, angiogenesis in chronic wounds also requires oxygen.…”