A series of large-scale proteomic studies applying mass-spectrometry have been undertaken to comprehensively identify novel human protein candidates that could interact with the arenavirus proteins [
85,
86,
87,
88,
89]. Special attention has been paid to NP-binding partners due to the multifunctional role of NP in the viral cycle, which involves crucial interactions with L and Z viral proteins [
12,
13,
15,
16,
17,
90], and its ability to hijack host factors to inhibit the antiviral innate immune response [
19,
22,
24,
91]. It is believed that identifying essential NP–host cell protein interactions can pave the way in the rational design of novel strategies to tackle arenavirus infections. One of the LASV NP interactors recently identified in human cells is DDX3, a protein belonging to the DEAD (Asp-Glu-Ala-Asp) box RNA helicase family, which harbors ATPase and RNA helicase activities [
89]. Of note, DDX3 has also emerged in proteomic studies of virus-infected cells, as a novel interacting partner of the OW LCMV and the NW JUNV NPs [
85]. CRISPR/Cas9-mediated deletion of DDX3 gene has been shown to lead to a significant reduction in virus yields of LASV, LCMV or JUNV in cell culture. Subsequently, lentiviral-mediated reconstitution of DDX3 expression resulted in a notable recovery in the infection rate of the three viruses, indicating a relevant role of DDX3 in virus growth as a proviral cellular factor [
89].
DDX3 is known to be involved in multiple steps of RNA metabolism, including RNA transcription and the initiation of translation in host cells [
92,
93,
94]. As other DEAD-box RNA helicases, such as DDX1 and DDX5, DDX3 appears to facilitate replication of different RNA viruses, as the alphavirus Venezuelan equine encephalitis virus and the hepatitis C virus (HCV), among others [
95,
96,
97,
98,
99,
100]. DDX3 is also required for translation of mRNAs containing a long or structured 5′ untranslated region (UTR), such as human immunodeficiency virus type-1 (HIV-1) genomic RNA (gRNA). Indeed, it was reported that DDX3 interacts with the 5′ region of the target mRNA, binds the eukaryotic translation initiation factor 4G (eIF4G) and poly A-binding protein cytoplasmic 1 (PABP), and interacts with HIV-1 Tat protein to facilitate translation of HIV-1 mRNAs [
96,
101]. In reference to arenaviruses, it was demonstrated that translation of a synthetic arenavirus mRNA analog was unaffected in DDX3-deficient cells, indicating no critical engagement of DDX3 in viral mRNA translation initiation [
89]. In contrast, minireplicon assay-based experiments demonstrated that the pro-arenaviral activity of DDX3 strongly depends on DDX3’s ability to promote viral RNA synthesis, involving both previously described DDX3 ATPase and helicase RNA-unwinding activities in this function [
89,
102].
Strikingly, alternative roles have been ascribed to DDX3 in the context of different viral infections [
103,
104]. On the one hand, DDX3 is considered an antiviral factor given that it is involved in the innate immune response against some viruses such as HIV-1, DENV and HCV [
105,
106,
107]. DDX3 has been shown to collaborate in the production of IFN-I, through interaction with components of the RIG-I-mediated IFN-I induction pathway [
108,
109,
110]. However, in contrast to this IFN-I promoting capacity of DDX3, mechanistic analysis has provided evidence that, in the case of LCMV, DDX3 suppresses the IFN-I response at late times of infection, still it remains to be confirmed whether this IFN-I-suppressive role of DDX3 is sustained in the context of an infection with the pathogenic LASV [
89]. Secondly, DDX3 is known to be an essential component for stress granule (SG) assembly, and to interact with other SG proteins, such as eIF4E [
111]. Different proteomic approaches based on mass-spectrometry have singled out new arenavirus NP-binding candidates related to the SG biology; including but not limited to the Ras GTPase-activating protein-binding protein 1 (G3BP1), eIF2α, apoptosis-inducing factor mitochondrion-associated 1 (AIFM1) and PABP, yet none of them have been confirmed as LASV interactors by alternative biochemical methods [
85,
89]. Of note, colocalization experiments have revealed the association of the NW arenavirus TCRV replication–transcription complexes (RTCs), where NP accumulates, with G3BP1 and a non-canonical collection of ribosomal proteins, including the ribosomal proteins RPS6 and RPL10a, as well as translation initiation factors eIF4G and eIF4A [
112]. In this regard, the finding that JUNV infection inhibits SG formation [
113] might be related to the NP-mediated sequestration of DDX3 and other SG-related proteins, resulting in the lack of availability of essential factors needed for SG nucleation. Similarly, in the case of influenza virus infections, it has been hypothesized that the interaction of DDX3 with the viral NS1 protein prevents DDX3 binding to eIF4E and PABP1 as well as DDX3–NP interaction, thus suppressing SG formation, NP recruitment into SGs, and DDX3 antiviral activity [
114]. Therefore, it is possible that in a similar way, LASV NP may counteract DDX3 antiviral function and in turn use DDX3 to enhance its own replication.