In the work presented here, by analyzing expression levels of all host genes in HIV-1-infected cells by RNA-seq, we identified nine genes functionally associated with the nucleolus and whose expression was down-regulated in infected samples. A closer look at these genes showed that they all encode for proteins that play critical roles in ribosome biogenesis. Down-regulation of genes involved in ribosome biogenesis was validated by RT-qPCR analysis using total RNA of Jurkat cells. Moreover, these results were further confirmed in infected primary CD4+ T lymphocytes. Importantly, the same effect was observed in independent RNA-Seq datasets, where we found an even larger number of downregulated genes with the same functional role and cellular localization. By performing a Northern Blot analysis using the same total RNA subjected to the RNA-seq experiments, we were able to show an impairment of pre-rRNA processing in HIV-1-infected samples compared to mock-infected samples, resulting in a marked decrease in the accumulation of the 30S rRNA precursor. These results demonstrated that, indeed, HIV-1 infection affects at least one of the steps leading to ribosome biogenesis, uncovering a novel mechanism of regulation of host gene expression mediated by the virus. In support of this hypothesis, it has been previously reported that other viruses can either promote, as hepatitis C virus, or inhibit, as poliovirus, the host pre-rRNA synthesis and, notably, herpes simplex virus type 1 can affect pre-rRNA processing without affecting pre-rRNA transcription. Concerning HIV-1, it has recently been reported that expression of the viral Tat TH-302 CYP17 inhibitor protein in Drosophila oocytes leads to a dramatic reduction of cytoplasmic ribosomes, probably caused by an impairment of pre-rRNA maturation. The Nef protein, in turn, has been found associated to components of the 40S ribosomal subunit, in particular the 18S rRNA and the RPS10 protein. In addition, it was previously published that HIV-1 infected CEMx174 T cells displayed a subtle decrease in soluble and membrane-associated polyribosomes compared to mock-infected control. Moreover, infected cells showed suppression of host mRNAs translation mediated by the action of the viral protein Vpr while the translation of viral structural protein is sustained. In light of our results, it can be hypothesized that alteration of ribosome biogenesis mediated by HIV-1 may allow the virus to alter the protein translation machinery or to induce stress signals, thus inhibiting host protein synthesis and, ultimately, inducing apoptotic pathways. This HIV-1 strategy will most likely come into action in the late steps of viral replication, once progeny virions have been massively produced and cell survival has become dispensable, if not deleterious, for the virus. Further studies will be necessary to dissect the link between ribosome biogenesis alteration and viral infection, possibly unraveling a novel intricate network of interactions between HIV-1 and host cell. PCMV can remain latent in adult pigs, but active infection causes fatal systemic failure in piglets less than 3 weeks of age.