Inhibition of Calmodulin-Dependent Kinase Kinase Blocks Human Cytomegalovirus-Induced Glycolytic Activation and Severely Attenuates Production of Viral Progeny

Abstract

Viruses depend on the host cell to provide the energy and biomolecular subunits necessary for production of viral progeny. We have previously reported that human cytomegalovirus (HCMV) infection induces dramatic changes to central carbon metabolism, including glycolysis, the tricarboxylic acid (TCA) cycle, fatty acid biosynthesis, and nucleotide biosynthesis. Here, we explore the mechanisms involved in HCMV-mediated glycolytic activation. We find that HCMV virion binding and tegument protein delivery are insufficient for HCMV-mediated activation of glycolysis. Viral DNA replication and late-gene expression, however, are not required. To narrow down the list of cellular pathways important for HCMV-medicated activation of glycolysis, we utilized pharmaceutical inhibitors to block pathways reported to be both involved in metabolic control and activated by HCMV infection. We find that inhibition of calmodulin-dependent kinase kinase (CaMKK), but not calmodulin-dependent kinase II (CaMKII) or protein kinase A (PKA), blocks HCMV-mediated activation of glycolysis. HCMV infection was also found to target calmodulin-dependent kinase kinase 1 (CaMKK1) expression, increasing the levels of CaMKK1 mRNA and protein. Our results indicate that inhibition of CaMKK has a negligible impact on immediate-early-protein accumulation yet severely attenuates production of HCMV viral progeny, reduces expression of at least one early gene, and blocks viral DNA replication. Inhibition of CaMKK did not affect the glycolytic activation induced by another herpes virus, herpes simplex virus type 1 (HSV-1). Furthermore, inhibition of CaMKK had a much smaller impact on HSV-1 replication than on that of HCMV. These data suggest that the role of CaMKK during the viral life cycle is, in this regard, HCMV specific. Taken together, our results suggest that CaMKK is an important factor for HCMV replication and HCMV-mediated glycolytic activation.It has long been known that infection with numerous evolutionarily divergent viruses results in a general activation of host cell metabolism (4,9,10,17,21,25,30). Furthermore, this metabolic activation can be clinically helpful. For example, a wide variety of antiviral compounds target specific nucleotide metabolic activities to treat numerous different viral infections, such as those caused by hepatitis B virus, HIV, human cytomegalovirus (HCMV), and herpes simplex virus (HSV) (1,6,12,20). While in some instances these activities have proven to be therapeutically beneficial, the identity of most of the specific metabolic activities induced by viral infection and the mechanisms through which they are activated are unclear. The identification of these activities and their associated mechanisms may highlight novel targets for therapeutic intervention given the viral dependence on the host cell metabolic network for the production of viral progeny.We have previously found that infection with HCMV induces substantial changes to the host cell metabolic network (22,23). HCMV is a betaherpesvirus containing a large d...

Vaccinia Virus Requires Glutamine but Not Glucose for Efficient Replication

“…As our laboratory and others have previously described, many different viruses activate glycolysis (1, 4-6, 29, 30), indicating an important role for this metabolic pathway during virus infection. In agreement with this, inhibition of glycolysis has been shown to attenuate both HCMV and HSV-1 replication as well as to induce apoptosis in cells latently infected with KSHV (29)(30)(31)(32). Recently, it was reported that the VACV protein C16 stabilizes hypoxia-induced factor 1␣ (HIF-1␣), resulting in the transcriptional upregulation of HIF-responsive genes, including two genes involved in glucose metabolism (33).…”

Dengue Virus Induces and Requires Glycolysis for Optimal Replication

“…As our laboratory and others have previously reported, many human viruses activate glycolysis (1,3,5,8,32,41). Moreover, inhibition of the glycolytic pathway has been shown to restrict HCMV and HSV-1 replication and induce apoptosis in cells latently infected with KSHV (32,(41)(42)(43).…”

Viral activation of cellular metabolism

“…Subsequent studies examined the mechanism of HCMV induction of glucose uptake and glycolysis. Viral protein expression is required for HCMV induced glycolysis, as cyclohexamide blocks induction (McArdle et al, 2011). It appears that early genes are necessary for HCMV induction of glyocolysis as phosphonoacetic acid, an inhibitor of viral replication that prevents the expression of late viral genes, has no effect on HCMV induced glycolysis.…”

“…This study also analyzed a cellular pathway know to be important for increased glycolysis in cancer cells, the calmodulin-dependent protein kinase family. An inhibitor of CAMKK, but not CaMKII, blocks HCMV induced glycolytic flux (McArdle et al, 2011). Additionally, the CAMKK inhibitor blocks viral replication and late gene synthesis and the production of viral progeny.…”