The Cyclin-Dependent Kinase 5 Inhibitor Peptide Inhibits Herpes Simplex Virus Type 1 Replication

Abstract

In order to evaluate the influence of CDK5 inhibitory peptide (CIP) on Human alphaherpesvirus 1 (HSV-1) replication, we constructed two recombinant adeno-associated-virus 2 (rAAV2) vectors encoding CIP fused with cyan-fluorescent-protein (CFP), with or without nuclear localization signal. A third vector encoding non-fused CIP and CFP was also constructed. HeLa and HEK 293T cells were infected with the rAAV-CIP vectors at multiplicity of infection (MOI) of 5000, in the absence or presence of a recombinant HSV-1 that encodes a yellow-fluorescent-protein (rHSV48Y; MOI = 1). Cells co-infected with rHSV48Y and rAAV vectors that did not express the CIP gene (rAAV-CFP-Neo) served as controls. At 24 h after infection, the effect of CIP on rHSV48Y replication was assessed by PCR, qRT-PCR, Western-blot, flow-cytometry, epifluorescence and confocal microscopy. We show that in cultures co-infected with rAAV-CFP-Neo, 27% of the CFP-positive cells present rHSV48Y replication compartments. By contrast, in cultures co-infected with CIP-encoding rAAV2 vectors and rHSV48Y only 6–20% of the cells positive for CIP showed rHSV48Y replication compartments, depending on the CIP variant. Flow-cytometry showed that less than 40% of the rHSV48Y/rAAV-CIP, and more than 75% of rHSV48Y/rAAV-CFP-Neo co-infected cells were positive for both transgene products. The microscopy and flow-cytometry data support the hypothesis that CIP is inhibiting HSV-1 replication.

Long noncoding RNA U90926 is crucial for herpes simplex virus type 1 proliferation in murine retinal photoreceptor cells

“…Whole DNA was extracted from HSV-1-infected 661W cells using the NucleoSpin RNA kit with the NucleoSpin RNA/DNA buffer set (Macherey-Nagel). HSV-1 DNA was quantified by qPCR using specific primers for the ICP27 gene of HSV-1, and the results were normalised against Gapdh 33 . The relative HSV-1 DNA amount was calculated based on a 1 h post HSV-1 infection sample.…”

The genetic architecture of human infectious diseases and pathogen-induced cellular phenotypes

“…The A and B chains of the CDK5-p25 complex ( Figure 4C for structure diagram (Tarricone et al, 2001)) are required for cytoskeletal protein binding (CDK5), whereas the D and E chains (p25) are involved in actin regulation and kinase function, all molecular processes implicated in our pathway analysis. Intriguingly, blocking CDK5 can have a substantial impact on the outcome of inflammatory diseases including sepsis (Pfänder et al, 2019), enhancing the anti-inflammatory potential of immunosuppressive treatments, and has been shown to attenuate herpes virus replication (Man et al, 2019), suggesting that modulation of this complex is important for viral pathogenesis.…”

Human DNA Virus Exploitation of the MAPK-ERK Cascade

“…Through the implementation of dominant-negative ERK1 constructs, Colao et al demonstrated that ERK1 activity is necessary for cell cycle control during HSV-1 infection, which is in turn required for viral replication [145]. Interestingly, HSV-1 infection also requires cyclin E and the cyclin-dependent kinases (CDK) 2 and 5 for infection, suggesting further viral influence over the cell cycle and viral replication [145,148,149]. HSV-2 also utilizes the MAPK-ERK pathway to dysregulate the cell cycle, specifically through ribonucleotide reductase R1 protein (ICP10), to induce a proliferative cellular state [141,150,151], demonstrating the conserved utilization of the MAPK-ERK pathway for cell cycle dysregulation between these human herpesviruses.…”