Biology subjects

Sarshad, A.

Publications and source records attributed to Sarshad, A..

5 recordsLinked to original sources

bioRxiv2025

RBProximity-CLIP Enables Subcellular Mapping of RNA-Binding Protein Interactions at Nucleotide Resolution

RNA-binding proteins (RBPs) enable post-transcriptional gene regulation (PTGR) through specific interactions with RNA molecules, influencing processes ranging from nuclear processing and export to cytoplasmic localization, translation, storage and degradation. A key determinant of PTGR processes is the subcellular compartmentalization of RBPs, which dictates RNA targets they can access and the regulation performed in that environment. To characterize RBP-RNA interactions at subcellular resolution, we developed RBProximity-CLIP. RBProximity-CLIP enables compartment-specific isolation and profiling of individual RBP-RNA interactions by combining APEX2-based proximity labeling and 4-thiouridine-enhanced RNA-protein crosslinking, with sequential RBP- and biotin-affinity purifications. Using this approach, we profiled the RNA targets of three RBPs, AGO2, YBX1, and ELAVL1, across the cytoplasmic, nuclear, and nucleolar compartments, revealing nucleus-specific miRNA-mediated AGO2 targets, as well as subsets of YBX1 and ELAVL1 targets that differ by compartment, yet share identical binding motifs. RBProximity-CLIP enables specific and sensitive detection of compartment-specific RBP-RNA interactomes, thereby providing new insight into spatial gene regulation by RBPs.

cell biology↗

bioRxiv2025

Extended nuclear glycosylation is a common post-translational modification

In eukaryotes, glycans modify proteins in the secretory pathway and the extracellular space. Aside from nucleocytoplasmic O-GlcNAc, glycosylation is not considered a relevant post-translational modification in other cellular compartments. Here, we challenge this long-standing paradigm by showing that extended O-glycans are commonly found on intranuclear proteins. Through comprehensive genetic and biochemical analyses, we conclusively demonstrate that these O-glycans stem from the secretory pathway, yet are found on nuclear proteins across mammalian cell lines and primary cells. Using knock-out cell lines, we show mechanistically that nuclear glycans are shuttled to the nucleus via active vesicular transport. We identify several of these intranuclear glycoproteins as RNA-binding proteins, including KHSRP/FUBP2, RBM12, and RPP30. Lastly, we show that site-specific glycosylation of RPP30 is crucial for effective tRNA processing. Overall, our findings suggest a much broader role for glycosylation in regulating cellular functions and open up investigation into the role of glycans in more biological processes.

molecular biology↗

bioRxiv2025

SARS-CoV-2 Exploits Host Translation and Immune Evasion Pathways via Viral RNA-Host Protein Interactions

RNA viruses, including SARS-CoV-2, Influenza A Virus (IAV), Zika Virus, and Dengue Virus (DENV) pose serious global health challenges by manipulating host cellular mechanisms. SARS-COV-2, in particular exploits host translational machinery to enhance replication and evade immune response. Here, we investigated how SARS-CoV-2 circumvents host immune defenses through RNA - host protein interactions. By integrating multiple datasets, ClusterProfiler, KEGG, Reactome, WikiPathways, and Gene Ontology, we performed functional enrichment analyses on host protein interactions with SARS-CoV-2 RNA. Our results identified key pathways involved in viral replication, translation regulation, and immune evasion. Comparing SARS-CoV-2 interactomes from IAV, Zika, and DENV, we uncovered a subset of 275 common host proteins serving as promising targets for broad-spectrum antiviral strategies. Network analysis highlighted critical translation factors (EEF1A1, EIF4A1, EIF3H) and RNA-binding proteins (NCL, ILF3) as key nodes in viral replication. These findings provide insights into RNA virus pathogenesis and support the development of targeted therapeutics.

immunology↗

bioRxiv2024

Influenza A Virus H7 nanobody recognizes a conserved immunodominant epitope on hemagglutinin head and confers heterosubtypic protection

Influenza remains a persistent global health challenge, largely due to the virus continuous antigenic drift and shift, which impede the development of a universal vaccine. To address this, the identification of broadly neutralizing antibodies and their epitopes is crucial. Nanobodies, with their unique characteristics and binding capacity, offer a promising avenue to identify such epitopes. Here, we isolated and purified a hemagglutinin (HA)-specific nanobody that recognizes an H7 subtype of influenza A virus. Notably, the nanobody, named E10, exhibited broad-spectrum binding, cross-group neutralization and in vivo protection across various influenza A subtypes. Through phage display and in vitro characterization, we demonstrated that E10 specifically targets an epitope on HA head. This epitope is part of the conserved lateral patch of HA head and proved to be highly immunodominant upon H7 infection. Importantly, immunization with a peptide including the E10 epitope elicited cross-reactive antibodies and mediated partial protection from lethal viral challenge. Our data highlight the potential of E10 and its associated epitope as a candidate for future influenza prevention strategies.

microbiology↗

bioRxiv2024

Nuclear RNAi Modulates Influenza A Virus Infectivity By Downregulating Type-I Interferon Response

The role of Argonaute (AGO) proteins and the RNA interference (RNAi) machinery in mammalian antiviral response has been debated. Therefore, we set out to investigate how mammalian RNAi impacts influenza A virus (IAV) infection. We reveal that IAV infection triggers nuclear accumulation of AGO2, which is directly facilitated by p53 activation. Mechanistically, we show that IAV induces nuclear AGO2 targeting of TRIM71, a proposed AGO2 E3 ligase, and type-I interferon-pathway genes for silencing. Accordingly, Tp53-/- mice do not accumulate nuclear AGO2 and demonstrate decreased susceptibility to IAV infection. Hence, the RNAi machinery is highjacked by the virus to evade the immune system and support viral replication. Furthermore, the FDA approved drug arsenic trioxide, which prevents p53 tetramerization and nuclear translocation, increases interferon response and decreases viral replication in vitro and in a mouse model in vivo. Our data indicates that targeting the AGO2:p53-mediated silencing of innate immunity may offer a promising strategy to mitigate viral infections.

biochemistry↗