Biology subjects

Ruiz-Perez, F.

Publications and source records attributed to Ruiz-Perez, F..

3 recordsLinked to original sources

bioRxiv2026

A plasmid-encoded H-NS protein selectively binds its own plasmid

H-NS is an abundant nucleoid-associated protein found in Enterobacterales species. Some conjugative plasmids encode H-NS homologues, which are thought to facilitate plasmid maintenance and reduce the fitness costs associated with plasmid carriage. Here, we characterize HppXCROD2, an H-NS homologue encoded by the IncX4 plasmid pCROD2 of Citrobacter rodentium. Our data indicate that HppXCROD2 has a strong preference for binding pCROD2 over the chromosome or other plasmids. By contrast, chromosomally encoded H-NS displays no preference for plasmid sequence. When expressed from a heterologous plasmid in Escherichia coli, HppXCROD2 showed similar DNA-sequence preference to chromosomally encoded H-NS. Moreover, HppXCROD2 binding to a sequence from pCROD2 was much lower when that sequence was cloned in a laboratory plasmid. Thus, HppXCROD2 preferentially binds DNA in the context of the plasmid where it is encoded, a phenomenon we term "cognate plasmid specificity". We propose that cognate plasmid specificity occurs through recognition of plasmid-specific DNA topology generated by plasmid-encoded topoisomerases. Cognate plasmid specificity may insulate regulation of plasmid genes from the effects of host DNA, while minimizing disruption of host chromosome regulation due to plasmid carriage. IMPORTANCEMany bacteria carry conjugative plasmids, mobile DNA molecules that spread traits such as antibiotic resistance. Some conjugative plasmids encode proteins related to the bacterial DNA-binding protein H-NS. We show that an H-NS-like protein from the IncX4 plasmid pCROD2 binds almost exclusively to the plasmid from which it originates, while largely ignoring the host chromosome. Our findings reveal a previously unrecognized mechanism that allows plasmids to regulate their own genes with high specificity while minimizing interference with host gene expression.

microbiology↗

bioRxiv2025

Bright Cells, and Inefficient Enzymatic Activity in a Renin Hypomorph Mouse

Juxtaglomerular (JG) cells are crucial regulators of blood pressure and fluid-electrolyte homeostasis. Under normal conditions, renin secretion by JG cells is sufficient to maintain homeostasis. However, under physiological stress such as narrowing of one of the renal arteries, heart failure, dehydration, or chronic administration of renin-angiotensin system (RAS) inhibitors, additional cells along the renal arterioles are transformed to the renin phenotype to meet the demands for renin and regain homeostasis. In cases of prolonged and persistent stimulation of renin cells, concentric arteriolar hypertrophy develops. The study of renin cell identity, plasticity and function often requires the isolation of this rare cell type. Here, we report on the generation of a mouse model to label renin-expressing cells with a bright fluorescent reporter under control of the Ren1c locus for the tracking and isolation of renin cells. Kidneys from adult heterozygous (Het) Ren1ctdTomato/+ mice showed tdTomato signal confined to the JG area under basal conditions, and extending along the afferent arterioles and in the intraglomerular mesangium upon treatment with captopril + low-salt diet to induce the endocrine transformation of renin cells. Unexpectedly, homozygous (Homo) Ren1ctdTomato/tdTomato mice exhibited increased tdTomato signal that extended along the afferent arterioles and into the mesangium even under normal physiological conditions, with progressive thickening of the kidney arterioles with age. Despite reduced renin immunostaining in the renal cortex, Ren1ctdTomato/tdTomato Homo mice exhibited significantly higher kidney Ren1 mRNA and circulating renin levels when compared to Het controls. Moreover, Homo mice showed significantly lower blood pressure measured under anesthesia and angiotensin I (Ang I) plasma levels, indicating compromised renin activity. In addition, Homo mice developed interstitial fibrosis and compromised kidney function. The concentric arteriolar hypertrophy phenotype observed in these mice is identical to that described when RAS is genetically or pharmacologically inhibited, including the presence of mutations in the renin gene. Unlike mice with global deletion of renin, these animals did not require neonatal saline injections to survive and did not develop other kidney abnormalities, indicating that the bicistronic approach rendered a renin hypomorphic mouse. Ren1ctdTomato mice constitute an excellent model for the bright and strong labeling of renin-expressing cells and for the study of the mechanisms involved in the development of concentric vascular hypertrophy under RAS inhibition. In addition, this model may provide a better understanding of factors controlling renin protein folding, stability, packaging, and release.

physiology↗

bioRxiv2025

CD45-mediated apoptosis and IL2R downregulation by serine proteases from diarrheagenic bacteria

Most enteropathogens secrete one or more members of the serine protease autotransporters of Enterobacteriaceae (SPATE). We previously demonstrated that SPATE cleaves various O-linked glycoproteins on leukocytes, including the tyrosine phosphatase CD45RO. SPATE impairs leukocyte functions and triggers apoptosis in activated T cells in vitro. Here, we show that SPATE produced by pathogenic E. coli, Shigella, and the mouse pathogen Citrobacter rodentium cleaves not only CD45RO but also CD45 isoforms containing exons A and B. We found that the cleavage of CD45 in primary T cells from both human and murine sources correlated with decreased IL2RA (CD25) surface expression in a concentration-dependent manner. SPATE did not cleave CD25 or affect T cell activation. However, SPATE requires CD45 expression for the depletion of CD25 in activated T cells, as SPATE did not significantly impact CD25 in the Jurkat J45.01 cell line, which lacks CD45. More importantly, we discovered that J45.01 cells resisted SPATE-mediated apoptosis, whereas apoptotic wild-type Jurkat cells exhibited decreased surface expression of CD25. Furthermore, we observed that mice infected with C. rodentium lacking SPATE displayed lower mortality, delayed intestinal colonization, reduced inflammatory cytokines, and decreased leukocyte infiltration in the lamina propria while having a higher number of CD25+ T cells compared to mice infected with wild-type CR or the CR SPATE mutant expressing Crc2 in trans. Our data suggest that SPATE-producing pathogens trigger T-cell apoptosis through CD45 via a mechanism akin to IL2 deprivation, demonstrating that SPATE can act as immunomodulators at various levels of the immune system. SIGNIFICANCEWe have demonstrated for the first time that serine proteases (C2S) from clinically relevant pathogens, such as E. coli pathotypes and Shigella, can cleave leukocyte glycoproteins, including the tyrosine phosphatase CD45, which play crucial roles in cellular and immune functions. In this study, we discovered that C2S induces apoptosis in activated T cells through a previously unknown mechanism resembling IL-2 deprivation, mediated by CD45. Furthermore, we found that C2S is essential for bacterial virulence in vivo. This suggests that pathogens producing C2S may possess previously undescribed immunoregulatory functions that enhance their survival in the host and contribute to the disease process by eliminating T cells through the targeting of CD45 and the IL-2 receptor.

microbiology↗