Efficient transmission of herpesviruses is essential for dissemination in host populations; however, little is known about the viral genes that mediate transmission, mostly due to a lack of natural virus-host model systems. Marek’s disease is a devastating herpesviral disease of chickens caused by Marek’s disease virus (MDV) and an excellent natural model to study skin-tropic herpesviruses and transmission. Like varicella zoster virus that causes chicken pox in humans, the only site where infectious cell-free MD virions are efficiently produced is in epithelial skin cells, a requirement for host-to-host transmission. Here, we enriched for heavily infected feather follicle epithelial skin cells of live chickens to measure both viral transcription and protein expression using combined short- and long-read RNA sequencing and LC/MS-MS bottom-up proteomics. Enrichment produced a previously unseen breadth and depth of viral peptide sequencing. We confirmed protein translation for 84 viral genes at high confidence (1% FDR) and correlated relative protein abundance with RNA expression levels. Using a proteogenomic approach, we confirmed translation of most well-characterized spliced viral transcripts and identified a novel, abundant isoform of the 14 kDa transcript family via IsoSeq transcripts, short-read intron-spanning sequencing reads, and a high-quality junction-spanning peptide identification. We identified peptides representing alternative start codon usage in several genes and putative novel microORFs at the 5’ ends of two core herpesviral genes, pUL47 and ICP4, along with strong evidence of independent transcription and translation of the capsid scaffold protein pUL26.5. Using a natural animal host model system to examine viral gene expression provides a robust, efficient, and meaningful way of validating results gathered from cell culture systems. In the natural host, the transcriptome and proteome of many herpesviruses are poorly defined. Here, we evaluated the viral transcriptome and proteome in feather follicle epithelial skin cells of chickens infected with Marek’s disease virus (MDV), an important poultry pathogen as well as an excellent model for skin-tropic human alphaherpesvirus replication in skin cells. Using fluorescently tagged virus, we significantly enriched the number of infected cells sampled from live chickens, greatly enhancing the detection of viral transcripts and proteins within a host background. We carefully defined the in vivo transcriptome using both short-read RNA-Seq and long-read full-length transcript sequencing, and deep MS/MS-based proteomics from the enriched virus-infected feather follicle samples confirmed the translation of most transcripts and identified novel expressed peptides supportive of an increasingly complex translational and regulatory viral landscape. The demonstrated deep peptide sequencing capability can serve as a template for future in vivo work in herpesviral proteomics.

The increasing global prevalence of diabetic nephropathy poses substantial health and economic burdens. Currently, effective anti-fibrotic therapies for managing kidney fibrosis associated with chronic kidney disease are lacking. This study reveals corisin, a microbiota-derived peptide, as a central driver in the progression of diabetic kidney fibrosis. Corisin levels were found to be markedly elevated in the serum of diabetic chronic kidney disease patients relative to healthy controls, with strong correlations to advanced disease stages and declining renal function. In a murine model of kidney fibrosis, corisin levels were similarly heightened, directly contributing to increased inflammation and worsening fibrosis and renal impairment. Notably, the use of a monoclonal anti-corisin antibody significantly reduced nephropathy severity in diabetic mice. Through molecular dynamics simulations and experimental validation, we demonstrated that corisin interacts with human serum albumin, potentially enhancing its renal accumulation and pathological impact. The pathogenic mechanism of corisin involves the acceleration of cellular senescence and the induction of epithelial-mesenchymal transition and apoptosis in kidney cells. These findings underscore the critical role of corisin in progressive diabetic nephropathy and suggest a promising new target for therapeutic intervention.

Conserved Herpesviridae protein kinases (CHPK) are conserved among all members of the Herpesviridae. Herpesviruses lacking CHPK propagate in cell culture at varying degrees, depending on the virus and cell culture system. CHPK is dispensable for Marek’s disease herpesvirus (MDV) replication in cell culture and experimental infection in chickens; however, CHPK—particularly its kinase activity—is essential for horizontal transmission in chickens, also known as natural infection. To address the importance of CHPK during natural infection in chickens, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) based proteomics of samples collected from live chickens. Comparing modification of viral proteins in feather follicle epithelial (FFE) cells infected with wildtype or a CHPK-null virus, we identified the US10 protein (pUS10) as a potential target for CHPK in vivo. When expression of pUS10 was evaluated in cell culture and in FFE skin cells during in vivo infection, pUS10 was severely reduced or abrogated in cells infected with CHPK mutant or CHPK-null viruses, respectively, indicating a potential role for pUS10 in transmission. To test this hypothesis, US10 was deleted from the MDV genome, and the reconstituted virus was tested for replication, horizontal transmission, and disease induction. Our results showed that removal of US10 had no effect on the ability of MDV to transmit in experimentally infected chickens, but disease induction in naturally infected chickens was significantly reduced. These results show CHPK is necessary for pUS10 expression both in cell culture and in the host, and pUS10 is important for disease induction during natural infection.

Peritoneal fluid (PF) is intimately associated with the gastrointestinal tract, and changes in the PF may directly reflect abdominal pathology. We aimed to quantify differences in the PF proteome between intestinal lesion type (ischemic vs. non-ischemic) and location (small vs. large intestine). PF samples were collected at hospital admission from horses presenting for abdominal pain (colic). Cases were clinically categorized by lesion type and location after resolution (10 per group). PF proteins were extracted and quantified by label-free liquid chromatography-tandem mass spectroscopy. Data were analyzed in Perseus and R, with functional annotation by UniProtKB and interaction visualization in STRING. Sixteen proteins unique to ischemic lesions and twelve unique to small intestinal lesions had significant network enrichment with functions related to inflammatory and immune responses. Identified proteins related to ischemic and small intestinal lesions included calprotectin, lactotransferrin, alpha 2 macroglobulin, and serine proteases/protease inhibitors, as well as apolipoprotein B and lipid metabolism pathways not previously described in relation to ischemic intestinal disease. While no single biomarker is expected to adequately diagnose or predict the outcome of equine colic, the proteins identified here should be considered as candidates for further study in a larger population.