The life cycle of the hepatitis C virus (HCV) can be divided into several stages, including viral entry, protein translation, RNA replication, viral assembly, and release. HCV genomic RNA replication occurs in the replication organelles (RO) and is tightly linked to ER membrane alterations containing replication complexes (proteins NS3 to NS5B). The amplification of HCV genomic RNA could be regulated by the RO biogenesis, the viral RNA structure (i.e., cis-acting replication elements), and both viral and cellular proteins. Studies on HCV replication have led to the development of direct-acting antivirals (DAAs) targeting the replication complex. This review article summarizes the viral and cellular factors involved in regulating HCV genomic RNA replication and the DAAs that inhibit HCV replication.

Hepatitis C virus (HCV) infection is associated with the development of chronic liver diseases, e.g., fibrosis, cirrhosis, even hepatocellular carcinoma, and/or extra-hepatic diseases such as diabetes. As an obligatory intracellular pathogen, HCV absolutely relies on host cells to propagate and is able to modulate host cellular factors in favor of its replication. Indeed, lots of cellular factors, including microRNAs (miRNAs), have been identified to be dysregulated during HCV infection. MiRNAs are small noncoding RNAs that regulate protein synthesis of their targeting mRNAs at the post-transcriptional level, usually by suppressing their target gene expression. The miRNAs dysregulated during HCV infection could directly or indirectly modulate HCV replication and/or induce liver diseases. Regulatory mechanisms of various miRNAs in HCV replication and pathogenesis have been characterized. Some dysregulated miRNAs have been considered as the biomarkers for the detection of HCV infection and/or HCV-related diseases. In this review, we intend to briefly summarize the identified miRNAs functioning at HCV replication and pathogenesis, focusing on the recent developments.

Viruses are obligatorily intracellular pathogens. To generate progeny virus particles, influenza A viruses (IAVs) have to divert the cellular machinery to ensure sufficient translation of viral mRNAs. To this end, several strategies have been exploited by IAVs, such as host gene shutoff, suppression of host innate immune responses, and selective translation of viral mRNAs. Various IAV proteins are responsible for host gene shutoff, e.g., NS1, PA-X, and RdRp, through inhibition of cellular gene transcription, suppression of cellular RNA processing, degradation of cellular RNAs, and blockage of cellular mRNA export from the nucleus. Host shutoff should suppress the innate immune responses and also increase the translation of viral mRNAs indirectly due to the reduced competition from cellular mRNAs for cellular translational machinery. However, many other mechanisms are also responsible for the suppression of innate immune responses by IAV, such as prevention of the detection of the viral RNAs by the RLRs, inhibition of the activities of proteins involved in signaling events of interferon production, and inhibition of the activities of interferon-stimulated genes, mainly through viral NS1, PB1-F2, and PA-X proteins. IAV mRNAs may be selectively translated in favor of cellular mRNAs through interacting with viral and/or cellular proteins, such as NS1, PABPI, and/or IFIT2, in the 5′-UTR of viral mRNAs. This review briefly summarizes the strategies utilized by IAVs to ensure sufficient translation of viral mRNAs focusing on recent developments.

Deoxynucleoside triphosphates (dNTPs) are crucial for the replication and maintenance of genomic information within cells. The balance of the dNTP pool involves several cellular enzymes, including dihydrofolate reductase (DHFR), ribonucleotide reductase (RNR), and SAM and HD domain-containing protein 1 (SAMHD1), among others. DHFR is vital for the de novo synthesis of purines and deoxythymidine monophosphate, which are necessary for DNA synthesis. SAMHD1, a ubiquitously expressed deoxynucleotide triphosphohydrolase, converts dNTPs into deoxynucleosides and inorganic triphosphates. This process counteracts the de novo dNTP synthesis primarily carried out by RNR and cellular deoxynucleoside kinases, which are most active during the S phase of the cell cycle. The intracellular levels of dNTPs can influence various viral infections. This review provides a concise summary of the interactions between different viruses and the genes involved in dNTP metabolism.

The nonstructural protein 1 (NS1) of influenza A virus (IAV) possesses multiple functions, such as the inhibition of the host antiviral immune responses, to facilitate viral infection. To search for cellular proteins interacting with the IAV NS1 protein, the yeast two-hybrid system was adopted. Proteasome family member PSMB4 (proteasome subunit beta type 4) was found to interact with the NS1 protein in this screening experiment. The binding domains of these two proteins were also determined using this system. The physical interactions between the NS1 and cellular PSMB4 proteins were further confirmed by co-immunoprecipitation assay and confocal microscopy in mammalian cells. Neither transiently nor stably expressed NS1 protein affected the PSMB4 expression in cells. In contrast, PSMB4 reduced the NS1 protein expression level, especially in the presence of MG132. As expected, the functions of the NS1 protein, such as inhibition of interferon activity and enhancement of transient gene expression, were suppressed by PSMB4. PSMB4 knockdown enhances IAV replication, while its overexpression attenuates IAV replication. Thus, the results of this study suggest that the cellular PSMB4 protein interacts with and possibly facilitates the degradation of the NS1 protein, which in turn suppresses IAV replication.

In addition to the receptor-binding domain (DII), the C-terminal domain (DIII) of three-domain Cry insecticidal δ-endotoxins from Bacillus thuringiensis has been implicated in target insect specificity, yet its precise mechanistic role remains unclear. Here, the 21 kDa high-purity isolated DIII fragment derived from the Cry4Ba mosquito-specific toxin was achieved via optimized preparative FPLC, allowing direct rendering analyses for binding characteristics toward its target receptor—Aedes aegypti membrane-bound alkaline phosphatase (Aa-mALP). Binding analysis via dotblotting revealed that the Cry4Ba-DIII truncate was capable of specific binding to nitrocellulose-bound Aa-mALP, with a binding signal comparable to its 65 kDa Cry4Ba-R203Q full-length toxin. Further determination of binding affinity via sandwich ELISA revealed that Cry4Ba-DIII exhibited a rather weak binding to Aa-mALP with a dissociation constant (K_d) of ≈1.1 × 10⁻⁷ M as compared with the full-length toxin. Intermolecular docking between the Cry4Ba-R203Q active toxin and Aa-mALP suggested that four Cry4Ba-DIII residues, i.e., Glu⁵²², Asn⁵⁵², Asn⁵⁷⁶, and Leu⁶¹⁵, are potentially involved in such toxin–receptor interactions. Ala substitutions of each residue (E522A, N552A, N576A and L615A) revealed that only the L615A mutant displayed a drastic decrease in biotoxicity against A. aegypti larvae. Additional binding analysis revealed that the L615A-impaired toxin also exhibited a reduction in binding capability to the surface-immobilized Aa-mALP receptor, while two bio-inactive DII-mutant toxins, Y332A and F364A, which almost entirely lost their biotoxicity, apparently retained a higher degree of binding activity. Altogether, our data disclose a functional importance of the C-terminal domain of Cry4Ba for serving as a potential receptor-binding moiety in which DIII-Leu⁶¹⁵ could conceivably be exploited for the binding to Aa-mALP, highlighting its contribution to toxin interactions with such a target receptor in mediating larval toxicity.