Biology subjects

Llano, M.

Publications and source records attributed to Llano, M..

4 recordsLinked to original sources

bioRxiv2025

NF-κB-Driven HIV-1 Gene Expression in Human Cells Is Independent of Poly(ADP-ribose) polymerase-1 Function.

The cellular enzyme poly (ADP-ribose) polymerase-1 (PARP-1) is required for NF-{kappa}B to activate inflammatory and immune response gene expression. NF-{kappa}B is also an important transcription factor in HIV-1 gene expression during active replication and latency reactivation. Therefore, enhancing NF-{kappa}B signaling is an alternative for HIV-1 latency reactivation, but significant systemic side effects related to the NF-{kappa}B role in inflammatory and immune responses are predictable. To verify this prediction, we determined whether PARP-1 is required in NF-{kappa}B-dependent HIV-1 gene expression in a human CD4+ T lymphoblastoid cell line (SUP-T1) and HEK 293T cells. Our findings indicated that PARP-1 knockout does not impair HIV-1 infection or gene expression. Specifically, NF-{kappa}B-dependent HIV-1 gene expression was not impaired by PARP-1 deficiency, highlighting an important transcriptional regulatory difference between HIV-1 and inflammatory and immune activation genes. Our findings define a negligible role of PARP-1 in HIV-1 gene expression, suggesting that PARP-1 antagonism could ameliorate the expected inflammatory response with latency-reactivating agents that act through the NF-{kappa}B signaling pathway. ImportancePARP-1 is required for NF-{kappa}B to activate the expression of inflammatory and immune response genes. NF-{kappa}B is also an important transcription factor in HIV-1 gene expression during active replication and latency reactivation. Enhancing NF-{kappa}B signaling is expected to cause HIV-1 latency reactivation, but significant systemic side effects related to the NF-{kappa}B role in inflammatory and immune responses are predictable. The role of PARP-1 in NF-{kappa}B-mediated activation of HIV-1 gene expression and in viral infection has not been determined in the context of HIV-1 infection of CD4+ T cells. Our data indicate that PARP-1 is dispensable for NF-{kappa}B-mediated activation of HIV-1 gene expression in a human CD4+ T lymphoblastoid cell line. These findings suggest that the pharmacological antagonism of PARP-1 could diminish the inflammatory effects of latency-reactivating agents that activate NF-{kappa}B signaling without impairing their effect on HIV-1 gene expression.

molecular biology↗

bioRxiv2024

Poly (ADP-ribose) polymerase-1 regulates HIV-1 replication in human CD4+ T cells

The cellular enzyme poly (ADP-ribose) polymerase-1 (PARP-1) regulates multiple processes that are potentially implicated in HIV-1 infection. However, the role of PARP-1 in HIV-1 infection remains controversial, with reports indicating or excluding that PARP-1 influence early steps of the HIV-1 life cycle. Most of these studies have been conducted with Vesicular Stomatitis virus Glycoprotein G (VSV-G)-pseudotyped, single-round infection HIV-1; limiting our understanding of the role of PARP-1 in HIV-1 replication. Therefore, we evaluated the effect of PARP-1 deficiency or inhibition in HIV-1 replication in human CD4+ T cells. Our data showed that PARP-1 knockout increased viral replication in SUP-T1 cells. Similarly, a PARP-1 inhibitor that targets PARP-1 DNA-binding activity enhanced HIV-1 replication. In contrast, inhibitors affecting the catalytic activity of the enzyme were inactive. In correspondence with the pharmacological studies, mutagenesis analysis indicated that the DNA-binding domain was required for the PARP-1 anti-HIV-1 activity, but the poly-ADP-ribosylation activity was dispensable. Our results also demonstrated that PARP-1 acts at the production phase of the viral life cycle since HIV-1 produced in cells lacking PARP-1 was more infectious than control viruses. The effect of PARP-1 on HIV-1 infectivity required Env, as PARP-1 deficiency or inhibition did not modify the infectivity of Env-deleted, VSV-G-pseudotyped HIV-1. Furthermore, virion-associated Env was more abundant in sucrose cushion-purified virions produced in cells lacking the enzyme. However, PARP-1 did not affect Env expression or processing in the producer cells. In summary, our data indicate that PARP-1 antagonism enhances HIV-1 infectivity and increases levels of virion-associated Env. ImportanceDifferent cellular processes counteract viral replication. A better understanding of these interfering mechanisms will enhance our ability to control viral infections. We have discovered a novel, antagonist effect of the cellular enzyme poly (ADP-ribose) polymerase-1 (PARP-1) in HIV-1 replication. Our data indicate that PARP-1 deficiency or inhibition augment HIV-1 infectivity in human CD4+ T cells, the main HIV-1 target cell in vivo. Analysis of the mechanism of action suggested that PARP-1 antagonism increases in the virus the amounts of the viral protein mediating viral entry to the target cells. These findings identify for the first time PARP-1 as a host factor that regulates HIV-1 infectivity, and could be relevant to better understand HIV-1 transmission and to facilitate vaccine development.

microbiology↗

bioRxiv2023

Schlafen14 Regulates Gene Expression Depending on Codon Usage

Schlafen (SLFN) is a family of proteins upregulated by type I interferons with a regulatory role in translation. Intriguingly, SLFN14 associates with the ribosome and can degrade rRNA, tRNA, and mRNA in vitro, but a role in translation is still unknown. Ribosomes are important regulatory hubs during translation elongation of mRNAs rich in rare codons. Therefore, we evaluated the potential role of SLFN14 in the expression of mRNAs enriched in rare codons, using HIV-1 genes as a model. We found that SLFN14 regulates in a variety of cell types, including primary immune cells, the expression of HIV-1 and non-viral genes based on their codon adaptation index, a measurement of the synonymous codon usage bias; consequently, SLFN14 inhibited the replication of HIV-1. The potent inhibitory effect of SLFN14 on the expression of the rare codon-rich transcript HIV-1 Gag was minimized by codon optimization. Mechanistically, we found that the endoribonuclease activity of SLFN14 is required, and that ribosomal, but not messenger, RNA degradation is associated with the transcript selectivity of SLFN14. Therefore, we propose that SLFN14 impairs the expression of transcripts rich in rare codons, in a catalytic-dependent manner.

molecular biology↗

bioRxiv2018

Schlafen 11 Restricts Flavivirus Replication

Schlafen 11 (Slfn11) is a ubiquitously expressed interferon stimulating gene (ISG) that controls synthesis of proteins by regulating tRNA abundance. Likely through this mechanism, Slfn11 has previously been shown to impair human immunodeficiency virus 1 (HIV-1) infection and the expression of codon-biased open reading frames. Because replication of positive-sense single-stranded RNA [(+)ssRNA viruses] requires the immediate translation of the incoming viral genome whereas negative sense, single stranded [(-)ssRNA] viruses carry at infection an RNA replicase that makes multiple translation competent copies of the incoming viral genome, we reasoned that (+)ssRNA viruses will be more sensitive to the effect of Slfn11 on protein synthesis than (-)ssRNA viruses. To evaluate this hypothesis, we tested the effects of Slfn11 on the replication of a panel of ssRNA viruses in the human glioblastoma cell line A172, which naturally expresses Slfn11. Depletion of Slfn11 in this cell line significantly increased the replication of (+)ssRNA viruses from the Flavivirus family, including West Nile (WNV), dengue (DENV), and Zika virus (ZIKV) but had no significant effect on the replication of the (-)ssRNA viruses vesicular stomatitis (VSV, Rhabdoviridae family) and Rift Valley fever (RVFV, Phenuiviridae family). Despite that WNV titers in Slfn11-deficient cells were almost 100-fold higher than in cells expressing this protein; they produced approximately two-fold less viral particles, as determined by PCR-based quantification of virion-associated WNV RNA in the cell culture supernatant. These data indicated that Slfn11 impairs WNV fitness but does not affect other steps of the viral life cycle including entry, viral RNA replication and translation, and budding. Similarly to the proposed anti-HIV-1 mechanism of Slfn11, this protein prevented WNV-induced down-regulation of a subset of tRNAs implicated in the translation of 19% of the viral polyprotein. Importantly, we provided evidence suggesting that the broad anti-viral activity of Slfn11 requires other cellular proteins, since overexpression of Slfn11 in cells that naturally lack the expression of this protein, did not impair WNV or HIV-1 infection. In summary, this study demonstrates that Slfn11 restricts flaviviruses replication by impairing viral fitness.\n\nAUTHOR SUMMARYThe host targets mechanisms that viruses have evolved to optimize replication. We provide evidence that the cellular protein Schlafen 11 (Slf11) impairs replication of flaviviruses, including West Nile (WNV), dengue (DENV), and Zika virus (ZIKV). However, replication of single-stranded, negative RNA viruses was not affected. Specifically, Slf11 decreases the fitness of WNV potentially by preventing virus-induced modifications of the host tRNA repertoire that could lead to enhanced viral protein folding. Furthermore, we demonstrated that Slf11 is not the limiting factor of this novel broad anti-viral pathway.

microbiology↗