Impact of TRIM5α and TRIM22 Genes Expression on the Clinical Course of Coronavirus Disease 2019.

OBJECTIVE: The innate immune response in humans involves a wide variety of factors, including the tripartite motif-containing 5α (TRIM5α) and 22 (TRIM22) as a cluster of genes on chromosome 11 that have exhibited antiviral activity in several viral infections. We analyzed the correlation of the expression of TRIM5α and TRIM22 with the severity of Coronavirus Disease 2019 (COVID-19) in blood samples of 330 patients, divided into two groups of severe and mild disease, versus the healthy individuals who never had contact with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). METHODS: The transcription level of TRIM5α and TRIM22 was determined by quantitative real-time polymerase chain reaction (qPCR). The laboratory values were collected from the patients' records. RESULTS: The expression of both genes was significantly lower in the severe group containing the hospitalized patients than in both the mild group and the control group. However, in the mild group, TRIM22 expression was significantly higher (p <0.0001) than in the control group while TRIM5α expression was not significantly different between these two groups. We found a relationship between the cycle threshold (Ct) value of patients and the expression of the aforementioned genes. CONCLUSION: The results of our study indicated that lower Ct values or higher RNA viral load might be associated with the downregulation of TRIM5α and TRIM22 and the severity of COVID-19. Additional studies are needed to confirm the results of this study.

Mapping of functional SARS-CoV-2 receptors in human lungs establishes differences in variant binding and SLC1A5 as a viral entry modulator of hACE2.

BACKGROUND: The COVID-19 pandemic is an infectious disease caused by SARS-CoV-2. The first step of SARS-CoV-2 infection is the recognition of angiotensin-converting enzyme 2 (ACE2) receptors by the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein. Although the molecular and structural bases of the SARS-CoV-2-RBD/hACE2 interaction have been thoroughly investigated in vitro, the relationship between hACE2 expression and in vivo infection is less understood. METHODS: Here, we developed an efficient SARS-CoV-2-RBD binding assay suitable for super resolution microscopy and simultaneous hACE2 immunodetection and mapped the correlation between hACE2 receptor abundance and SARS-CoV-2-RBD binding, both in vitro and in human lung biopsies. Next, we explored the specific proteome of SARS-CoV-2-RBD/hACE2 through a comparative mass spectrometry approach. FINDINGS: We found that only a minority of hACE2 positive spots are actually SARS-CoV-2-RBD binding sites, and that the relationship between SARS-CoV-2-RBD binding and hACE2 presence is variable, suggesting the existence of additional factors. Indeed, we found several interactors that are involved in receptor localization and viral entry and characterized one of them: SLC1A5, an amino acid transporter. High-resolution receptor-binding studies showed that co-expression of membrane-bound SLC1A5 with hACE2 predicted SARS-CoV-2 binding and entry better than hACE2 expression alone. SLC1A5 depletion reduces SARS-CoV-2 binding and entry. Notably, the Omicron variant is more efficient in binding hACE2 sites, but equally sensitive to SLC1A5 downregulation. INTERPRETATION: We propose a method for mapping functional SARS-CoV-2 receptors in vivo. We confirm the existence of hACE2 co-factors that may contribute to differential sensitivity of cells to infection. FUNDING: This work was supported by an unrestricted grant from "Fondazione Romeo ed Enrica Invernizzi" to Stefano Biffo and by AIRC under MFAG 2021 - ID. 26178 project - P.I. Manfrini Nicola.

The emerging multifunctional roles of ERAP1, ERAP2 and IRAP between antigen processing and renin-angiotensin system modulation.

The Endoplasmic Reticulum Aminopeptidase 1 and 2 (ERAP1 and ERAP2) and Insulin Regulated Aminopeptidase (IRAP) are three M1 zinc metalloproteases whose role in antigen processing is the refining of peptidome either in the Endoplasmic reticulum (ERAP1 and ERAP2), or in the endosomes (IRAP). However, other novel and distinct functions are emerging. Here, we focus specifically on ERAP2. This gene has a peculiar evolutionary history, being absent in rodents and undergoing in humans to a balanced selection of two haplotypes, one of which not expressing the full length ERAP2. These observations suggest that its role in antigen presentation is not essential. An additional, less investigated role is in the regulation of the Renin Angiotensin System (RAS). ERAP1 and ERAP2 cleave Angiotensin II (Ang II) into Ang III and IV, which counteract the action of Ang II whereas IRAP is itself the receptor for Ang IV. We have recently reported that macrophages, independently from the haplotype, express and release a N-terminus ERAP2 "short" form which directly binds IRAP and the two molecules are co-expressed in the endosomes and on the cell membrane. This new evidence suggests that the maintenance of the ERAP2 gene in humans could be due to its activity in the regulation of the RAS system, possibly as an Ang IV agonist. Its role in the immune-mediated diseases as well as in disorders more specifically related to an imbalance of the RAS system, including hypertension, pre-eclampsia but also viral infections such as COVID-19, is discussed here.

ERAP1 and ERAP2 Enzymes: A Protective Shield for RAS against COVID-19?

Patients with coronavirus disease 2019 (COVID-19) have a wide variety of clinical outcomes ranging from asymptomatic to severe respiratory syndrome that can progress to life-threatening lung lesions. The identification of prognostic factors can help to improve the risk stratification of patients by promptly defining for each the most effective therapy to resolve the disease. The etiological agent causing COVID-19 is a new coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that enters cells via the ACE2 receptor. SARS-CoV-2 infection causes a reduction in ACE2 levels, leading to an imbalance in the renin-angiotensin system (RAS), and consequently, in blood pressure and systemic vascular resistance. ERAP1 and ERAP2 are two RAS regulators and key components of MHC class I antigen processing. Their polymorphisms have been associated with autoimmune and inflammatory conditions, hypertension, and cancer. Based on their involvement in the RAS, we believe that the dysfunctional status of ERAP1 and ERAP2 enzymes may exacerbate the effect of SARS-CoV-2 infection, aggravating the symptomatology and clinical outcome of the disease. In this review, we discuss this hypothesis.

Low temperature bacterial expression of the neutral amino acid transporters SLC1A5 (ASCT2), and SLC6A19 (B0AT1).

It is well established that Escherichia coli represents a powerful tool for the over-expression of human proteins for structure/function studies. In many cases, such as for membrane transporters, the bacterial toxicity or the aggregation of the target protein hamper the expression limiting the application of this tool. The aim of this study was finding the appropriate conditions for the expression of reluctant proteins that is the human neutral amino acid transporters ASCT2 and B0AT1, that have great relevance to human health in cancer therapy and in COVID-19 research, respectively. The cDNAs coding for the proteins of interest were cloned in the pCOLD I vector and different E. coli strains (BL21 codon plus RIL, and RosettaGami2) were cultured in absence or in presence of glucose (0.5-1%), at low temperature (15 °C), and low inducer concentrations (10-100 µM). Cell growth and protein production were monitored by optical density measurements and western blotting assay, respectively. Even though in different conditions, the expression of both amino acid transporters was obtained.Reducing the growth rate of specific E. coli strains by lowering the temperature and the IPTG concentration, together with the addition of glucose, two reluctant human neutral amino acid transporters have been expressed in E. coli. The results have a potentially great interest in drug discovery since ASCT2 is an acknowledged target of anticancer therapy, and B0AT1 together with ACE2 is part of a receptor for the SARS-Cov-2 RBD proteins.

MAIT Cells in COVID-19: Heroes, Villains, or Both?

Mucosa-associated invariant T cells (MAIT cells) are unconventional, innate-like T lymphocytes with remarkable effector and immunoregulatory functions. They are abundant in the human peripheral blood and also enriched in mucosal layers and in the lungs, SARS-CoV-2's main ports of entry. Once activated, MAIT cells produce inflammatory cytokines and cytolytic effector molecules quickly and copiously. MAIT cells are best known for their antibacterial and antifungal properties. However, they are also activated during viral infections, typically in a cytokine-dependent manner, which may promote antiviral immunity. On the other hand, it is plausible to assume active roles for MAIT cells in infection-provoked cytokine storms and tissue damage. SARS-CoV-2 infection may be asymptomatic, mild, severe, or even fatal, depending on sex, age, the presence of preexisting morbidities, and the individual's immunological competence, or lack thereof, among other factors. Based on the available literature, I propose that MAIT cells regulate the host response to SARS-CoV-2 and constitute attractive targets in the prevention or clinical management of coronavirus disease 19 (COVID-19) and some of its complications. Unlike mainstream T cells, MAIT cells are restricted by a monomorphic antigen-presenting molecule called MHC-related protein 1 (MR1). Therefore, MR1 ligands should modify MAIT cell functions relatively uniformly in genetically diverse subjects and may be tested as immunotherapeutic agents or vaccine adjuvants in future studies.