The mRNA encoding the JUND tumor suppressor detains nuclear RNA-binding proteins to assemble polysomes that are unaffected by mTOR

Abstract

One long-standing knowledge gap is the role of nuclear proteins in mRNA translation. Nuclear RNA helicase A (DHX9/RHA) is necessary for the translation of the mRNAs of JunD proto-oncogene AP-1 transcription factor subunit (JUND) and HIV-1 genes, and nuclear cap-binding protein 1 (NCBP1)/CBP80 is a component of HIV-1 polysomes. The protein kinase mTOR activates canonical messenger ribonucleoproteins (mRNPs) by posttranslationally downregulating the eIF4E inhibitory protein, 4E-BP1. We posited here that NCBP1 and DHX9/RHA (RHA) support a translation pathway of JUND RNA that is independent of mTOR. We present evidence from reciprocal immunoprecipitation experiments indicating that NCBP1 and RHA both are components of mRNPs in several cell types. Moreover, tandem affinity and RT-qPCR results revealed that JUND mRNA is a component of a previously unknown ribonucleoprotein complex. Results from the tandem IP indicated that another component of the JUND-containing ribonucleoprotein complex is NCBP3, a recently identified ortholog of NCBP2/CBP20. We also found that NCBP1, NCBP3, and RHA, but not NCBP2, are components of JUND-containing polysomes. Mutational analysis uncovered two dsRNA-binding domains of RHA that are necessary to tether JUND-NCBP1/NCBP3 to polysomes. We also found that JUND translation is unaffected by inhibition of mTOR, unless RHA was down-regulated by siRNA. These findings uncover a noncanonical cap-binding complex consisting of NCBP1/NCBP3 and indicate that RHA substitutes for eukaryotic translation initiation factor 4E (eIF4E) and eIF4G in activating mTOR-independent translation of the mRNA encoding the tumor suppressor JUND.

HIV-1 hypermethylated guanosine cap licenses specialized translation unaffected by mTOR

“…Indeed, the polysomes of HIV-1 Rev/RRE-dependent RNAs have failed to enrich eIF4E and instead engage a CBP80 mRNP ( 27 ). Recently, NCBP3/CBP80 heterodimer has been shown to assemble JUND polysomes for cap-dependent translation without eIF4E ( 21 ). To test whether TMG-capped HIV-1 mRNPs enriched eIF4E or NCBP3, the messenger ribonucleoproteins (mRNPs) were collected from HIV-infected, primary CD4 + T lymphocytes and subjected to RT-qPCR, as summarized in Fig.…”

“…Torin-1 is a selective catalytic site mTOR inhibitor that up-regulates hypophosphorylated 4E-BP1 ( 21 , 28 , 29 ). Lymphocytes were infected with HIV NL4-3 for 6 h (multiplicity of infection [MOI] = 1), washed, and continuously cultured with or without Torin-1.…”

“…The junD mRNA maintains translation during mTOR inhibition yet fails conventional tests for cap-independent translation by an internal ribosome entry mechanism ( 20 ). Recently, the heterodimeric CBP80/NCBP3 has been shown to assemble JUND polysomes ( 21 ). The CBP80-NCBP3 heterodimer had been characterized in conditions of metabolic stress and viral infection and has been assigned roles in antiviral defense and RNA export that remain under investigation ( 22 , 23 ).…”

A New Approach to 3D Modeling of Inhomogeneous Populations of Viral Regulatory RNA

“…The HIV late RNAs have been shown to specifically retain nuclear cap-binding complex (CBC) in polysomes and virions, instead of exchanging to eIF4E [ 56 ]. Recently, polysomal CBC RNPs were shown to activate eIF4E-independent translation of stress response proteins, overcoming AP-1 protein translation attenuation by mTOR inhibition [ 57 ]. Structural malleability near the HIV 5′-Cap may be beneficial to regulate cap-exchange and antagonize antiviral mTOR inhibition.…”

Circular RNAs Are Regulators of Diverse Animal Transcriptomes: One Health Perspective

“…The deposition of nuclear factors to pre-mRNA is necessary for nucleo-cytoplasmic transport and subsequent commitment to decay, storage or efficient translation to protein (Le Hir et al, 2001;Singh et al, 2020). circRNAs that are derived from introns (circular intronic RNAs, ciRNAs), remain in the nucleus and may regulate transcription of parental genes (Zhang et al, 2013;Zang et al, 2020) (Figure 1).…”