Transforming growth factor beta (TGF-␤) is a pleiotropic cytokine known to regulate cell growth, differentiation, and motility and is a potent modulator of immune function. TGF-␤ consequently plays a central role in carcinogenesis, and a dampened TGF-␤2 response by Theileria annulata-infected monocytes/macrophages underpins disease resistance to tropical theileriosis. Here, we show that concomitant with the loss of TGF-␤2 production, there is ablated expression of COX2 and EP4, which leads to a drop in cyclic AMP (cAMP) levels and, consequently, reduced activation of protein kinase A (PKA) and EPAC. This ablated phenotype can be rescued in attenuated macrophages by the addition of exogenous TGF-␤2, which reactivates the expression of COX2 and EP4 while repressing that of protein kinase inhibitor gamma (PKIG) to the levels in virulent macrophages. TGF-␤2 therefore promotes the adhesion and invasiveness of virulent macrophages by modulating COX2, EP4, and PKIG transcription to initiate a prostaglandin E 2 (PGE2)-driven autostimulatory loop that augments PKA and EPAC activities. A virulence phenotype stemming from the double activation of PKA and EPAC is the induction of a CREB-mediated transcriptional program and the upregulation of JAM-L-and integrin 4␣␤1-mediated adhesion of Theileria-infected macrophages.T heileria annulata is a tick-borne apicomplexan parasite and the causative agent of the cattle disease tropical theileriosis, which is of major economic importance in countries in Northern Africa and Asia. T. annulata transforms B cells and monocytes/macrophages, and infected leukocytes display many characteristics of cancer cells, such as heightened migratory and invasive capacities (1, 2). However, and importantly, the tumor-like phenotype is reversed upon drug-induced parasite death (3). Moreover, virulent macrophages can be attenuated by multiple in vitro passages, and upon attenuation, they lose both adhesion and invasiveness (4). The similarities in tumor hyperinvasiveness between Theileria-transformed leukocytes and human leukemias argue that studying Theileria-induced transformation can give insights into generally applicable mechanisms involved in tumor virulence.Transforming growth factor beta (TGF-␤) is a pleiotropic cytokine involved in numerous critical processes such as cell proliferation, differentiation, and survival or apoptosis. In normal cells and cells in early stages of cancer, TGF-␤ acts as a tumor suppressor and inhibits cell growth, while in late stages of cancer, it plays a contrasting role, acting as a tumor promoter enhancing metastasis and invasion. TGF-␤2 is strongly expressed in highly aggressive and malignant tumors (5). We have demonstrated that following infection, Theileria-transformed Holstein-Friesian (HF) (Bos taurus) macrophages produce more TGF-␤2 than do infected Sahiwal (Bos indicus) macrophages, and augmented TGF-␤2 output underpins the greater invasive capacity of infected HF macrophages. Moreover, attenuated Ode HF macrophages used as a live vaccine against tropical ...

Theileria annulata infects predominantly macrophages, and to a lesser extent B cells, and causes a widespread disease of cattle called tropical theileriosis. Disease-causing infected macrophages are aggressively invasive, but this virulence trait can be attenuated by long-term culture. Attenuated macrophages are used as live vaccines against tropical theileriosis and via their characterization one gains insights into what host cell trait is altered concomitant with loss of virulence. We established that sporozoite infection of monocytes rapidly induces hif1-α transcription and that constitutive induction of HIF-1α in transformed leukocytes is parasite-dependent. In both infectedmacrophages and B cells induction of HIF-1α activates transcription of its target genes that drive host cells to perform Warburg-like glycolysis. We propose that Theileria-infected leukocytes maintain a HIF-1α-driven transcriptional programme typical of Warburg glycolysis in order to reduce as much as possible host cell H2O2 type oxidative stress. However, in attenuated macrophages H2O2 production increases and HIF-1α levels consequently remained high, even though adhesion and aggressive invasiveness diminished. This indicates that Theileria infection generates a host leukocytes hypoxic response that if not properly controlled leads to loss of virulence.