Biology subjects

Novotny, J.

Publications and source records attributed to Novotny, J..

4 recordsLinked to original sources

bioRxiv2025

PAF15-PCNA assembly exhaustion governs lagging strand replication and replisome integrity

Genome replication in eukaryotic cells is surveyed by the S-phase checkpoint, which orchestrates sequential replication origin activation to avoid exhaustion of hitherto poorly defined rate-limiting replisome components. Here, we find that excessive activation of replication origins depletes chromatin-bound PCNA and lagging strand components, thereby limiting additional PCNA loading at new origins when checkpoint control is disrupted. PAF15 (PCNA-associated factor 15) emerges as a dosage-sensitive regulator of PCNA, delineating the dynamic range of global genome duplication and defining distinct roles for PCNA on the leading and lagging strands. Through its high-affinity PIP motif and interaction within the DNA encircling channel of PCNA, PAF15 stabilizes PCNA exclusively on the lagging strand, optimizing and rate-limiting lagging strand processing. On the other hand, misregulation of PAF15--whether by overexpression or mislocalization to the leading strand--impairs replication fork progression and leads to cell death. These defects are mitigated by TIMELESS and CLASPIN, which restrain PAF15-PCNA interactions beyond the lagging strand. E2F4-mediated repression orchestrates PAF15 expression in normal and cancer cells, maintaining its optimal dosage for lagging strand-specific interactions with PCNA. Thus, the S-phase checkpoint functions in concert to restrict origin activation when lagging strand PAF15-PCNA assembly is exhausted, linking a previously concealed strand-specific rate limitation to overall replication dynamics.

biochemistry↗

bioRxiv2024

A high lipid diet leads to greater pathology and lower tolerance during infection

Despite the clear effect of food resources on wildlife disease dynamics, few studies have investigated how shifts in diet, specifically macronutrient content, shape vertebrate host responses to infection. While each individual macronutrient plays a vital role in physiological processes necessary for pathogen defense, it is often difficult to disentangle the role of an individual macronutrient on disease pathology. We explored the effects of dietary macronutrient composition on disease pathology and feeding behavior of canaries (Serinus canaria domestica) infected with Mycoplasma gallisepticum (MG). First, we provided canaries with isocaloric diets comprised of identical ingredients that varied in macronutrient content (high-protein or high-lipid) then MG- or sham-inoculated birds. We then offered both diets to canaries before and after MG- and sham-inoculation. In experiment one, high protein diet birds consumed more food than high lipid diet birds and experienced a more pronounced decrease in food intake after infection. High protein diet birds were more tolerant to MG infection, exhibiting reduced pathology when compared to high lipid diet birds, despite the two treatments having similar levels of MG-specific antibodies and MG loads. When birds had access to both diets, they consumed more of the high protein diet and experienced pathology for less time than lipid or protein restricted birds. These results highlight that macronutrient makeup of the diet can shape host tolerance and pathology and thus affect host-pathogen transmission dynamics. Summary StatementMacronutrient composition of the diet shapes tolerance in an avian host pathogen system. Birds that had greater access to protein exhibited reduced pathology despite similar pathogen loads when compared with birds with greater access to lipids.

ecology↗

bioRxiv2020

Phenotypic and clonal stability of antigen-inexperienced memory-like T cells across the genetic background, hygienic status, and aging

Antigen-inexperienced memory-like T (AIMT) cells are functionally unique T cells representing one of the two largest subsets of murine CD8+ T cells. However, differences between laboratory inbred strains, insufficient data from germ-free mice, a complete lack of data from feral mice, and unclear relationship between AIMT cells formation during aging represent major barriers for better understanding of their biology. We performed a thorough characterization of AIMT cells from mice of different genetic background, age, and hygienic status by flow cytometry and multi-omics approaches including analyses of gene expression, TCR repertoire, and microbial colonization. Our data showed that AIMT cells are steadily present in mice independently of their genetic background and hygienic status. Despite differences in their gene expression profiles, young and aged AIMT cells originate from identical clones. We identified that CD122 discriminates two major subsets of AIMT cells in a strain-independent manner. Whereas thymic CD122LOW AIMT cells (innate memory) prevail only in young animals with high thymic IL-4 production, peripheral CD122HIGH AIMT cells (virtual memory) dominate in aged mice. Co-housing with feral mice changed the bacterial colonization of laboratory strains, but had only minimal effects on the CD8+ T-cell compartment including AIMT cells.

immunology↗

bioRxiv2018

The day/night difference in the circadian clock’s response to acute lipopolysaccharide and the rhythmic Stat3 expression in the rat suprachiasmatic nucleus

The circadian clock in the suprachiasmatic nucleus (SCN) regulates daily rhythms in physiology and behaviour and is an important part of the mammalian homeostatic system. Previously, we have shown that systemic inflammatory stimulation with lipopolysaccharide (LPS) induced the daytime-dependent phosphorylation of STAT3 in the SCN. Here, we demonstrate the LPS-induced Stat3 mRNA expression in the SCN and show also the circadian rhythm in Stat3 expression in the SCN, with high levels during the day. Moreover, we examined the effects of LPS (1mg/kg), applied either during the day or the night, on the rhythm in locomotor activity of male Wistar rats. We observed that recovery of normal locomotor activity patterns took longer when the animals were injected during the night. The clock genes Per1, Per2 and Nr1d1, and phosphorylation of kinases ERK1/2 and GSK3{beta} are sensitive to external cues and function as the molecular entry for external signals into the circadian clockwork. We also studied the immediate changes in these clock genes expressions and the phosphorylation of ERK1/2 and GSK3{beta} in the suprachiasmatic nucleus in response to daytime or night-time inflammatory stimulation. We revealed mild and transient changes with respect to the controls. Our data stress the role of STAT3 in the circadian clock response to the LPS and provide further evidence of the interaction between the circadian clock and immune system.

neuroscience↗