Biology subjects

Nita-Lazar, A.

Publications and source records attributed to Nita-Lazar, A..

5 recordsLinked to original sources

bioRxiv2026

Quantitative Modeling of TLR Signaling Reveals Missing Negative Feedback Guiding Identification of TANK-IKKε Checkpoint

Toll-like receptor (TLR) signaling must be activated rapidly and then terminated to support host defense without sustained inflammation. We developed a rule-based model of mouse macrophage TLR4 signaling at the molecular-interaction level using measured protein copy numbers, RNA-seq-based abundance estimates, literature- and structure-informed reaction rates, and 979 dynamic experimental constraints. The trained model reproduced much of the TLR4-induced NF-{kappa}B and MAP kinase response but consistently failed to capture deactivation of MyD88, TRAF6-associated species, and IKK/{beta}. The recurrent model failure conveyed important biological information, localizing missing regulation to the proximal MyD88-IRAK-TRAF6 module and guiding experimental evaluation of IKK{varepsilon} and its scaffold TANK. Loss of IKK{varepsilon} enhanced transcriptional, cytokine, MAP kinase, and NF-{kappa}B responses to MyD88-specific TLR ligands. TANK deficiency produced a similar cellular phenotype and abolished stimulus-induced IKK{varepsilon} phosphorylation. Deficiency of either protein increased IRAK1 and TRAF6 ubiquitination without increasing MyD88 ubiquitination, placing the inhibitory checkpoint at or immediately downstream of the IRAK1-TRAF6 ubiquitin-signaling node. Overlapping but non-identical in vivo phenotypes further supported a shared regulatory axis with additional protein-specific functions. Our study presents a model-experiment discovery cycle where quantitative pathway discordance identifies missing biology and reveals a TANK-dependent IKK{varepsilon} checkpoint that restrains MyD88-driven inflammation.

systems biology↗

bioRxiv2025

Sulfide is a keystone metabolite for gut homeostasis and immunity

Hydrogen sulfide is a gaseous, reactive molecule specifically enriched in the gastrointestinal tract. Here, we uncover a non-redundant role for sulfide in the control of both microbial and immune homeostasis of the gut. Notably, depletion of sulfide via both pharmaceutical and dietary interventions led to a profound collapse of CD4 T cells in the ileum of the small intestine lamina propria and significant impact on microbial ecology. As a result, mice with reduced sulfide within the gut were deficient in their ability to mount T cell dependent antibody responses to oral vaccine. Mechanistically, our results support the idea that sulfide could act directly on CD4 T cells via enhanced AP-1 activation, leading to heightened proliferation and cytokine production. This study uncovers sulfides as keystone components in gut ecology and provides mechanistic insight between diet, gut sulfide production and mucosal immunity.

immunology↗

bioRxiv2024

Human ESCRT-I and ALIX function as scaffolding helical filaments in vivo

The Endosomal Sorting Complex Required for Transport (ESCRT) is an evolutionarily conserved machinery that performs reverse-topology membrane scission in cells universally required from cytokinesis to budding of enveloped viruses. Upstream acting ESCRT-I and ALIX control these events and link recruitment of viral and cellular partners to late-acting ESCRT-III CHMP4 through incompletely understood mechanisms. Using structure-function analyses combined with super-resolution imaging, we show that ESCRT-I and ALIX function as distinct helical filaments in vivo. Together, they are essential for optimal structural scaffolding of HIV-1 nascent virions, the retention of viral and human genomes through defined functional interfaces, and recruitment of CHMP4 that itself assembles into corkscrew-like filaments intertwined with ESCRT-I or ALIX helices. Disruption of filament assembly or their conformationally clustered RNA binding interfaces in human cells impaired membrane abscission, resulted in major structural instability and leaked nucleic acid from nascent virions and nuclear envelopes. Thus, ESCRT-I and ALIX function as helical filaments in vivo and serve as both nucleic acid-dependent structural scaffolds as well as ESCRT-III assembly templates. Significance statementWhen cellular membranes are dissolved or breached, ESCRT is rapidly deployed to repair membranes to restore the integrity of intracellular compartments. Membrane sealing is ensured by ESCRT-III filaments assembled on the inner face of membrane; a mechanism termed inverse topology membrane scission. This mechanism, initiated by ESCRT-I and ALIX, is universally necessary for cytokinesis, wound repair, budding of enveloped viruses, and more. We show ESCRT-I and ALIX individually oligomerize into helical filaments that cluster newly discovered nucleic acid-binding interfaces and scaffold-in genomes within nascent virions and nuclear envelopes. These oligomers additionally appear to serve as ideal templates for ESCRT-III polymerization, as helical filaments of CHMP4B were found intertwined ESCRT-I or ALIX filaments in vivo. Similarly, corkscrew-like filaments of ALIX are also interwoven with ESCRT-I, supporting a model of inverse topology membrane scission that is synergistically reinforced by inward double filament scaffolding.

cell biology↗

bioRxiv2024

IL-10 dependent adaptation allows macrophages to adjust inflammatory responses to TLR4 stimulation history

Macrophages must scale inflammation to the trajectories of infections, escalating as pathogen loads rise, resolving as they fall. The cytokine IL-10 and its chromatin-level effector BCL-3 are critically important anti-inflammatory agents, yet how they avoid muting newly recruited cells before those cells read their own pathogen input has been unclear. Applying systematically varied consecutive TLR4 (Kdo2-Lipid A) stimuli to bone marrow-derived macrophages, we identify two coupled features. First, macrophages retain a quantitative memory of prior stimulation: only when a secondary TLR stimulus matches or exceeds a prior one, I{kappa}B degradation, NF-{kappa}B/MAPK activation, and cytokine output increase. Second, IL-10 susceptibility is itself gated by TLR4 history: even a 100-fold IL-10 excess fails to suppress TNF- in weakly primed cells. Both trace to history- and IL-10-dependent BCL-3 recruitment with p65 displacement at the Tnf {kappa}B site. Modeling shows how this licensing logic enables trajectory-aware responses that clear pathogens while limiting tissue damage.

immunology↗

bioRxiv2017

Internal modifications in the CENP-A nucleosome modulate centromeric dynamics

Post-translational modifications (PTMs) of core histones have studied for over 2 decades, and are correlated with changes in transcriptional status, chromatin fiber folding, and nucleosome dynamics. However, within the centromere-specific histone H3 variant CENP-A, few modifications have been reported, and their functions remain largely unexplored. In this multidisciplinary report, we utilize in silico computational and in vivo approaches to dissect lysine 124 of human CENP-A, which was previously reported to be acetylated in advance of replication. Computational modeling demonstrates that acetylation of K124 causes tightening of the histone core, and hinders accessibility to its C-terminus, which in turn diminishes CENP-C binding. Additionally, CENP-A K124ac/H4 K79ac containing nucleosomes are prone to DNA sliding. In vivo experiments using an acetyl or unacetylatable mimic (CENP-A K124Q and K124A respectively) reveal alterations in CENP-C levels, and a modest increase in mitotic errors. Furthermore, mutation of K124 results in alterations in centromeric replication timing, with the permanently acetylated form replicating centromeres early, and the unacetylable form replicating centromeres late. Purification of native CENP-A proteins followed by mass spectrometry analysis reveal that while CENP-A K124 is acetylated at G1/S, it switches to monomethylation during early and mid-S phase. Finally, we provide evidence that the HAT p300 is involved in this cycle. Taken together, our data suggest that cyclical modifications within the CENP-A nucleosome can influence the binding of key kinetochore proteins, the integrity of mitosis and centromeric replication. These data support the emerging paradigm that core modifications in histone variant nucleosomes transduce defined changes to key biological processes.

biochemistry↗