Transforming growth factor ß2 promotes transcription of COX2 and EP4, leading to a prostaglandin E2-driven autostimulatory loop that enhances virulence of Theileria annulata-transformed macrophages.

Transforming growth factor beta (TGF-ß) is a pleiotropic cytokine known to regulate cell growth, differentiation, and motility and is a potent modulator of immune function. TGF-ß consequently plays a central role in carcinogenesis, and a dampened TGF-ß2 response by Theileria annulata-infected monocytes/macrophages underpins disease resistance to tropical theileriosis. Here, we show that concomitant with the loss of TGF-ß2 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-ß2, which reactivates the expression of COX2 and EP4 while repressing that of protein kinase inhibitor gamma (PKIG) to the levels in virulent macrophages. TGF-ß2 therefore promotes the adhesion and invasiveness of virulent macrophages by modulating COX2, EP4, and PKIG transcription to initiate a prostaglandin E2 (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 4αß1-mediated adhesion of Theileria-infected macrophages.

The level of H2O2 type oxidative stress regulates virulence of Theileria-transformed leukocytes.

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 infected macrophages 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 H2 O2 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.

Stage-specific IFN-induced and IFN gene expression reveal convergence of type I and type II IFN and highlight their role in both acute and chronic stage of pathogenic SIV infection.

Interferons (IFNs) play a major role in controlling viral infections including HIV/SIV infections. Persistent up-regulation of interferon stimulated genes (ISGs) is associated with chronic immune activation and progression in SIV/HIV infections, but the respective contribution of different IFNs is unclear. We analyzed the expression of IFN genes and ISGs in tissues of SIV infected macaques to understand the respective roles of type I and type II IFNs. Both IFN types were induced in lymph nodes during early stage of primary infection and to some extent in rectal biopsies but not in PBMCs. Induction of Type II IFN expression persisted during the chronic phase, in contrast to undetectable induction of type I IFN expression. Global gene expression analysis with a major focus on ISGs revealed that at both acute and chronic infection phases most differentially expressed ISGs were inducible by both type I and type II IFNs and displayed the highest increases, indicating strong convergence and synergy between type I and type II IFNs. The analysis of functional signatures of ISG expression revealed temporal changes in IFN expression patterns identifying phase-specific ISGs. These results suggest that IFN-γ strongly contribute to shape ISG upregulation in addition to type I IFN.

Correction: Stage-specific IFN-induced and IFN gene expression reveal convergence of type I and type II IFN and highlight their role in both acute and chronic stage of pathogenic SIV infection.

[This corrects the article DOI: 10.1371/journal.pone.0190334.].

Engineering attenuated virulence of a Theileria annulata-infected macrophage.

Live attenuated vaccines are used to combat tropical theileriosis in North Africa, the Middle East, India, and China. The attenuation process is empirical and occurs only after many months, sometimes years, of in vitro culture of virulent clinical isolates. During this extensive culturing, attenuated lines lose their vaccine potential. To circumvent this we engineered the rapid ablation of the host cell transcription factor c-Jun, and within only 3 weeks the line engineered for loss of c-Jun activation displayed in vitro correlates of attenuation such as loss of adhesion, reduced MMP9 gelatinase activity, and diminished capacity to traverse Matrigel. Specific ablation of a single infected host cell virulence trait (c-Jun) induced a complete failure of Theileria annulata-transformed macrophages to disseminate, whereas virulent macrophages disseminated to the kidneys, spleen, and lungs of Rag2/γC mice. Thus, in this heterologous mouse model loss of c-Jun expression led to ablation of dissemination of T. annulata-infected and transformed macrophages. The generation of Theileria-infected macrophages genetically engineered for ablation of a specific host cell virulence trait now makes possible experimental vaccination of calves to address how loss of macrophage dissemination impacts the disease pathology of tropical theileriosis.