Biology subjects

Iyyathurai, J.

Publications and source records attributed to Iyyathurai, J..

3 recordsLinked to original sources

bioRxiv2023

SARS-CoV-2 Mpro protease variants of concern display altered viral and host target processing but retain potency towards antivirals

Main protease of SARS-CoV-2 (Mpro) is the most promising drug target against coronaviruses due to its essential role in virus replication. With newly emerging variants there is a concern that mutations in Mpro may alter structural and functional properties of protease and subsequently the potency of existing and potential antivirals. We explored the effect of 31 mutations belonging to 5 variants of concern (VOC) on catalytic parameters and substrate specificity, which revealed changes in substrate binding and rate of cleavage of a viral peptide. Crystal structures of 11 Mpro mutants provided structural insight into their altered functionality. Additionally, we show Mpro mutations influence proteolysis of an immunomodulatory host protein Galectin-8 (Gal-8) and subsequent significant decrease in cytokine secretion, providing evidence for alterations in escape of host-antiviral mechanisms. Accordingly, mutations associated with the highly virulent Delta VOC resulted in significant increase in Gal-8 cleavage. Importantly, IC50s of nirmatrelvir (Pfizer) and our irreversible inhibitor AVI-8053 demonstrated no changes in potency for both drugs for all mutants, suggesting Mpro will remain a high-priority antiviral drug candidate as SARS-CoV-2 evolves.

biochemistry↗

bioRxiv2022

Early immunomodulatory program triggered by pro-tolerogenic Bifidobacterium pseudolongum drives cardiac transplant outcomes

BackgroundDespite ongoing improvements in regimens to prevent allograft rejection, most cardiac and other organ grafts eventually succumb to chronic vasculopathy, interstitial fibrosis, or endothelial changes, and eventually graft failure. The events leading to chronic rejection are still poorly understood and the gut microbiota is a known driving force in immune dysfunction. We previously showed that gut microbiota dysbiosis profoundly influences the outcome of vascularized cardiac allografts and subsequently identified biomarker species associated with these differential graft outcomes. MethodsIn this study, we further detailed the multifaceted immunomodulatory properties of pro-tolerogenic and pro-inflammatory bacterial species over time, using our clinically relevant model of allogenic heart transplantation. ResultsIn addition to tracing longitudinal changes in the recipient gut microbiome over time, we observed that Bifidobacterium pseudolongum (Bifido) induced an early anti-inflammatory phenotype within 7 days, while Desulfovibrio desulfuricans (Desulfo) resulted in a pro-inflammatory phenotype, defined by alterations in leukocyte distribution and lymph node (LN) structure. Indeed, in vitro results showed that Bifido and Desulfo acted directly on primary innate immune cells. However, by 40 days after treatment, these two bacterial strains were associated with mixed effects in their impact on LN architecture and immune cell composition and loss of colonization within gut microbiota, despite protection of allografts from inflammation with Bifido treatment. ConclusionsThese dynamic effects suggest a critical role for early microbiota-triggered immunological events such as innate immune cell engagement, T cell differentiation, and LN architectural changes in the subsequent modulation of pro-tolerant versus pro-inflammatory immune responses in organ transplant recipients.

immunology↗

bioRxiv2019

Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

BackgroundCT1 is a 25 amino acid therapeutic peptide incorporating the Zonula Occludens-1 (ZO-1)-binding domain of connexin43 (Cx43) that is currently in Phase III clinical testing for healing chronic skin wounds. In preclinical studies in mice, we reported that CT1 reduces arrhythmias and improves ventricular function following cardiac injury, effects that were accompanied by increases in PKC{varepsilon} phosphorylation of Cx43 at serine 368 (pS368). In this study, we undertake a systematic characterization of the molecular mode-of-action of CT1 in mitigating the effects of ischemia reperfusion injury on ventricular contractile function.\n\nMethods and ResultsTo determine the basis of CT1-mediated increases in pS368 we undertook tandem mass spectrometry of reactants in an in vitro assay of PKC{varepsilon} phosphorylation, identifying an interaction between negatively charged amino acids in the CT1 Asp-Asp-Leu-Glu-Iso sequence and positively charged lysines (Lys345, Lys346) in a short -helical sequence (H2) within the Cx43 CT domain. In silico modeling provided further support of the specificity of this interaction, leading us to conclude that CT1 has potential to directly interact with both Cx43 and ZO-1. Using surface plasmon resonance, thermal shift and phosphorylation assays, we characterized a series of CT1 variant peptides, identifying sequences competent to interact with either ZO-1 PDZ2 or the Cx43 CT, but with limited or no ability to bind both polypeptides. Based on this analysis, it was found that only those peptides competent to interact with Cx43, but not ZO-1 alone, resulted in increased pS368 phosphorylation in vitro and in vivo. Moreover, in a mouse model of global ischemia reperfusion injury we determined that pre-ischemic infusion only with those peptides competent to bind Cx43 preserved left ventricular (LV) contractile function following injury. Interestingly, a short 9 amino acid (MW=1110) Cx43-binding variant of the original 25 amino acid CT1 sequence demonstrated potent LV-protecting effects when infused either before or after ischemic injury.\n\nConclusionsInteraction of CT1 with the Cx43 CT, but not ZO-1 PDZ2, explains cardioprotection mediated by this therapeutic peptide. Pharmacophores targeting the Cx43 carboxyl terminus could provide a novel translational approach to preservation of ventricular function following ischemic injury.

cell biology↗