Biology subjects

Hashem, A. M.

Publications and source records attributed to Hashem, A. M..

5 recordsLinked to original sources

bioRxiv2026

Phosphorylation of UBE2J1 at serine residue S184 contributes towards infection and cellular syncytialization by Vesicular Stomatitis Virus

The ubiquitin-conjugating enzyme UBE2J1 functions in the proteasomal degradation of proteins at the ER. Existing evidence suggests that it plays a role during viral infection, with elevated UBE2J1 levels generally associated with increased infection. This is particularly relevant for some RNA viruses; however, the regulation of UBE2J1 during infection has not been well studied. Here, we used a BHK21 cell model to demonstrate that UBE2J1 overexpression promotes the replication of Vesicular Stomatitis Virus (VSV), as indicated by a significant increase in viral titres. To better understand the underlying molecular processes, cells were co-transfected to express the VSV-G protein and wild-type UBE2J1 protein, and we observed a significant increase in the syncytial fusion area. This effect was not observed when catalytically inactive (C91S) or phospho-deficient (S184A) forms of the protein were used. Interestingly, overexpression of a truncated, non-ER localized form of UBE2J1 ({Delta}TM) led to the largest increase in the syncytial fusion area. This arose as a result of increased syncytia size, and may indicate an enhanced cellular role if soluble forms of UBE2J1 are not anchored to the ER. Additional studies using truncated, mutated and wild-type proteins confirmed that UBE2J1 increases VSV viral replication and is associated with an increase in the number of infection plaques. Considering the emerging evidence for UBE2J1 involvement in viral infection, our finding should help in understanding the role of this protein in viral pathogenesis and cellular processes linked to syncytialization.

biochemistry↗

bioRxiv2026

Protein kinase A regulates phosphorylation of UBE2J1 at serine residue S266 in response to glucagon signalling

The ubiquitin conjugating enzyme UBE2J1/Ubc6e localizes to the endoplasmic reticulum where it mediates the ubiquitination and proteasomal degradation of terminally misfolded proteins. Although the protein is known to undergo phosphorylation at serine S184, we have considered modification at an additional site and used a bespoke anti-phospho antibody to confirm phosphorylation also at serine residue S266. Despite the well-described role of UBE2J1 in ER associated degradation (ERAD), we found no evidence for regulation at S266 during Unfolded Protein Response (UPR) induction by thapsigargin. Instead, our studies suggest that phosphorylation occurs independently at the S184 and S266 sites, with mutation at one site failing to disrupt basal phosphorylation at the second. We identified several contexts in which these two phosphorylations were differentially regulated. For example, ER localization, which is important for phosphorylation at S184, was not required for modification at S266, and sensitivity to proteasome inhibitors, which is regarded as a distinguishing feature of the S184 phospho-variant, was unaltered by the S266A mutation. Regarding regulation at S266 on the other hand, we found that pharmacological activation of protein kinase A resulted in rapid phosphorylation, with differential use of phospho-specific antibodies confirming that phosphorylation at S184 was unchanged by this treatment. Hormonal stimulation by glucagon resulted in a similar pattern of UBE2J1 phosphorylation, which occurred exclusively at S266 and could be inhibited by H89. The differential regulation demonstrated in these studies extends our understanding of the UBE2J1 enzyme, and may indicate a role in the integration of energy metabolism with environmental stress conditions.

biochemistry↗

bioRxiv2025

Genomic surveillance of Methicillin Resistant Staphylococcus aureus in Saudi Arabia.

Methicillin-resistant Staphylococcus aureus (MRSA) surveillance in regions with mass gatherings presents unique challenges for public health systems. Saudi Arabia, hosting millions of pilgrims annually, provides a distinctive setting for studying how human mobility shapes bacterial populations, yet comprehensive genomic surveillance data from this region remains limited. Here, we present an integrated analysis of S. aureus isolates collected across seven Saudi Arabian regions, combining whole-genome sequencing with extensive antimicrobial susceptibility testing and standardized metadata following FAIR data principles. Our analysis revealed striking differences between pilgrimage and non-pilgrimage cities. Pilgrimage cities showed significantly higher genetic diversity and antimicrobial resistance rates, harboring numerous international strains including recognized clones from diverse geographic origins. Reported lineage dynamics is changing, expanding toward community clones. While genomic prediction of antimicrobial resistance showed high accuracy for some antibiotics, particularly beta-lactams, with varying performance for others, highlighting the necessity for phenotypic testing in clinical settings. Our findings demonstrate how mass gatherings drive bacterial population structures and emphasize the importance of integrated surveillance approaches in regions with significant global connectivity and travel. ImportanceGenomic data enables the tracking of pathogens by revealing clonal expansions within populations and identifying successful lineages. However, comprehensive national-level data from Saudi Arabia remains limited on a large scale. The adoption of FAIR principles and reproducible workflows ensures robust, consistent analysis, fostering effective data sharing. The OneHealth approachs success depends on the integration and collaboration across diverse domains in todays digital landscape.

genomics↗

bioRxiv2023

Proteome profiling of nasopharynx reveals pathophysiological signature of COVID-19 disease severity

An aberrant innate immune system caused by the beta coronavirus SARS-CoV-2 is a characteristic manifestation of severe coronavirus disease 2019 (COVID-19). Here, we performed proteome profiling of nasopharyngeal (NP) swabs from 273 hospitalized patients with mild and severe COVID-19 symptoms, including non-survivors. We identified depletion in STAT1-mediated type I interferon response, retinol metabolism and NRF2 antioxidant system that are associated with disease severity in our patient demography. We found that the dysregulation of glucocorticoid signaling and renin-angiotensin-aldosterone system (RAAS) contribute to the pathophysiology of COVID-19 fatality. Hyperactivation of host innate immune system was observed in severe patients, marked by elevated proteins involved in neutrophil degranulation and platelet aggregation. Our study using high-throughput proteomics on the nasopharynx of COVID-19 patients provides additional evidence on the SARS-CoV-2-induced pathophysiological signatures of disease severity and fatality.

immunology↗

bioRxiv2021

Enhanced immunogenicity of a synthetic DNA vaccine expressing consensus SARS-CoV-2 Spike protein using needle-free immunization

The ongoing global pandemic of Coronavirus Disease 2019 (COVID-19) calls for an urgent development of effective and safe prophylactic and therapeutic measures. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) glycoprotein is a major immunogenic and protective protein, and plays a crucial role in viral pathogenesis. In this study, we successfully constructed a synthetic codon-optimized DNA-based vaccine as a countermeasure against SARS-CoV-2; denoted as VIU-1005. The design was based on the synthesis of codon-optimized coding sequence for optimal mammalian expression of a consensus full-length S glycoprotein. The successful construction of the vaccine was confirmed by restriction digestion and sequencing, and the protein expression of the S protein was confirmed by western blot and immunofluorescence staining in mammalian cells. The immunogenicity of the vaccine was tested in two mouse models (BALB/c and C57BL/6J). Th1-skewed systemic S-specific IgG antibodies and neutralizing antibodies (nAbs) were significantly induced in both models four weeks post three injections with 100 g of the VIU-1005 vaccine via intramuscular needle injection but not intradermal or subcutaneous routes. Importantly, such immunization induced long-lasting IgG response in mice that lasted for at least 6 months. Interestingly, using a needle-free system, we showed an enhanced immunogenicity of VIU-1005 in which lower doses such as 25-50 g or less number of doses were able to elicit significantly high levels of Th1-biased systemic S-specific IgG antibodies and nAbs via intramuscular immunization compared to needle immunization. Compared to the intradermal needle injection which failed to induce any significant immune response, intradermal needle-free immunization elicited robust Th1-biased humoral response similar to that observed with intramuscular immunization. Furthermore, immunization with VIU-1005 induced potent S-specific cellular response as demonstrated by the significantly high levels of IFN-{gamma}, TNF and IL-2 cytokines production in memory CD8+ and CD4+ T cells in BALB/c mice. Together, our results demonstrate that the synthetic VIU-1005 candidate DNA vaccine is highly immunogenic and capable of inducing long-lasting and Th1-skewed immune response in mice. Furthermore, we show that the use of needle-free system could enhance the immunogenicity and minimize doses needed to induce protective immunity in mice, supporting further preclinical and clinical testing of this candidate vaccine.

immunology↗