Proteo-genomic analysis of SARS-CoV-2: A clinical landscape of SNPs, COVID-19 proteome and host responses

A novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of COVID-19 and continues to be a global health challenge. To understand viral disease biology, we have carried out proteo-genomic analysis using next generation sequencing (NGS) and mass-spectrometry on nasopharyngeal swabs of COVID-19 patients to examine clinical genome and proteome. Our study confirms the hyper mutability of SARS-CoV-2 showing multiple SNPs. NGS analysis detected 27 mutations of which 14 are synonymous, 11 are missense and 2 are extragenic in nature. Phylogenetic analysis of SARS-CoV-2 isolates indicated their close relation to Bangladesh isolate and multiple origins of isolates within a country. Our proteomic analysis, for the first time identified 13 different SARS-CoV-2 proteins from the clinical swabs. Of the total 41 peptides captured by HRMS, 8 matched to nucleocapsid protein, 2 to ORF9b, 1 to spike glycoprotein and ORF3a, with remaining mapping to ORF1ab polyprotein. Additionally, host proteome analysis revealed several key host proteins to be uniquely expressed in COVID-19 patients. Pathway analysis of these proteins points towards modulation in immune response, especially involving neutrophil and IL-12 mediated signaling. Besides revealing the aspects of host-virus pathogenesis, our study opens new avenues to develop better diagnostic markers and therapeutics.

Tear Fluid Protein Changes in Dry Eye Syndrome Associated with Rheumatoid Arthritis: A Proteomic Approach.

PURPOSE: Sjögren syndrome (SS) secondary to rheumatoid arthritis (RA) affects lacrimal and salivary glands, and therefore dry eye syndrome (DES) is more prevalent in patients with RA. This study used a proteomic approach to identify potential biomarkers in tear of DES secondary to RA (DES-RA). METHODS: Tear specimens were collected with Schirmer strips from patients with DES with RA, patients with other types of dry eye (namely, primary Sjögrens and non-Sjögrens [NSS]), and age-matched controls. Tear proteins were subjected to 2D-differential gel electrophoresis (2D-DIGE), and the differentially expressed proteins were identified using nano ESI-LC-MS/MS analysis. RESULTS: Among the differentially regulated proteins of DES-RA that were identified, lactotransferrin isoform 1 precursor was found to be d own-regulated in 100% cases and SHC transforming 1 isoform in 63% of the cases, while proteins such as ribonuclease p protein subunit 20, protocadherin, and heterogeneous nuclear ribonucleoprotein Q isoform 6 were down-regulated in over 80% of the cases. Proteins such as Ecto-ADP ribosyltransferase 5 precursor, Rho-related GTP-binding protein, and RhoJ precursor were up-regulated in 80% of the cases. CONCLUSION: Functional annotation revealed that these proteins have roles in regulation, antimicrobial activity, immune, metabolic, and cellular processes. The study observed characteristic marker proteins differentially expressed in DES-RA that are previously unreported. Further validation is needed.

Two dimensional electrophoretic analysis of human tears: collection method in dry eye syndrome.

Tear proteomics, by 2-DE, can give a fingerprint of the protein profile, which is well suited in clinical proteomics for biomarker identification and in diagnostics. The mode of tear collection can influence the representation of the proteins in the tear and therefore it is important to use the appropriate method. In this study, capillary and Schirmer mode of tear collection was done in the healthy controls and the Schirmer method was validated in dry eye syndrome conditions. 2-D PAGE of normal and dry eye tear was performed using pH 3-10 linear IPG strips followed by 13% SDS-PAGE. The spot intensity was analyzed by the PD quest software. The two methods were compared using Bland-Altman statistical tool. The 2-D map of capillary and Schirmer tear showed 147 ± 8 spots and 145 ± 7 spots respectively. Both the collection methods were in agreement with each other and were comparable. Dry eye tear protein showed differential expression of proteins as observed in 25-35 kDa region. One of the significantly reduced protein was identified as proline-rich 4 protein. Schirmer method of tear collection is reliable in patients with dry eye, which can display the differential protein expression and help in biomarker identification.