Biology subjects

Prudovsky, I.

Publications and source records attributed to Prudovsky, I..

6 recordsLinked to original sources

bioRxiv2026

Envelope-Limited Chromatin Sheets (ELCS) Formation in The Nuclear Envelope of HL-60/S4 Cells

Envelope-Limited Chromatin Sheets (ELCS) can be induced in human promyelocytic HL-60/S4 cells by treatment with retinoic acid (RA). After 4 days, the differentiated granulocytes exhibit multilobed nuclei with outgrowths of the nuclear envelope (NE) and associated heterochromatin extending into the surrounding cytoplasm (ELCS). These fascinating structures reveal a periodic meshwork of 30 nm chromatin fibers, when viewed by Cryo-electron microscopy. Genetic and biochemical evidence indicates that RA increases the synthesis of Lamin B Receptor (LBR), which is a key enzyme for Cholesterol biosynthesis and is an essential bridge between the NE and peripheral heterochromatin. This article is in part a review of our microscopic data on the structure of ELCS, and in part a description of related transcription changes that result in the formation of ELCS. In addition, this article contains a structural and biochemical comparison of RA-induced granulocytes with phorbol ester (TPA) induced HL-60/S4 macrophages, which lack nuclear lobulation, do not form ELCS, and exhibit a reduction in LBR and Cholesterol biosynthesis. From our perspective, ELCS can be viewed as "fabric" outgrowths of the nuclear envelope, frequently connecting nuclear lobes and capable of sustaining the twisting and squeezing distortions imposed upon nuclear shape, as the granulocytes traverse narrow tissue channels.

cell biology↗

bioRxiv2026

Nuclear Pore Complexes in Various States of HL-60/S4 Cells

This study is focused upon how the structure and function of interphase nuclear pore complexes (NPCs) respond to various cellular stresses (i.e., cell differentiation, knockdown of Lamin B Receptor [LBR], and cellular dehydration) in a myeloid cell line (HL-60/S4). Each cellular stress was examined by GSEA (Gene Set Enrichment Analysis) to determine how the structure and function of the NPCs were affected. Cell differentiation into granulocytes and into macrophages resulted in widespread decreases in nucleoporin (NUP) transcription levels, affecting NPC structure and NPC transport capability. LBR knockdown (HL-60/sh1 cells) in undifferentiated cells exhibited major increases in NUP transcription, combined with improved NPC structural quality and transport capability, implying that nuclear pore function is not adversely affected by the loss of LBR. In contrast, cell dehydration of the undifferentiated HL-60/S4 cells in hyperosmotic culture medium resulted in disorganized nucleoporin transcription, with evidence of abnormal NPC structure and transport capability. The structural integrity and transport function of nuclear pores are clearly responsive to the various cellular stresses. Future investigations should examine the reversibility and resilience of these cells to such stresses as those described in this study.

cell biology↗

bioRxiv2025

Hyperosmotic Stress of HL-60/S4 Cells: Gene Set Enrichment Analysis and Microscopy Changes

Cellular homeostasis requires that internal cell conditions (e.g., pH, ionic and non-ionic solute concentrations and hydration levels) maintain a dynamic stability. To better understand the mechanisms enforcing this homeostasis, we examined the transcript abundance and cellular morphology of undifferentiated myeloid HL-60/S4 cells exposed to 30 or 60 minutes of hyperosmotic stress, resulting from 300 mM sucrose added to the tissue culture medium. We used Gene Set Expression Analyses (GSEA) to examine various cell functions (Phenotypes) of interest. Microscopy was also employed, where possible, to visualize the nuclear and chromatin physical states. We observed that hyperosmotic stress resulted in congelation of interphase and mitotic chromatin. In parallel, GSEA indicated a reduction of heterochromatin, histone lysine methyltransferase and mRNA transcription. Mitosis ceases during hyperosmotic stress, with the intact tubulin spindle remaining attached to clustered centromeres on the congealed chromosomes. GSEA indicated that ribosome biosynthesis in nucleoli and oxidative activity in mitochondria are significantly increased. In addition, the proteasome phenotype is increased, suggesting that protein synthesis and destruction are both occurring at an increased pace during hyperosmotic stress. Nuclear envelope-associated chromatin also appears to be affected by hyperosmotic stress: LBR and Heterochromatin Protein 1 alpha (CBX5) disperse into the cytoplasm. Evidence is also presented that 300 mM sucrose leads to a reduction of DNA methylation and aberrant cellular localization of MeCP2. The effects of acute hyperosmotic stress on HL-60/S4 cells are very diverse and very profound. In many respects, the stress response resembles a frantic attempt for cell survival in the face of inevitable cell death.

cell biology↗

bioRxiv2025

Phenotypic Changes in Undifferentiated and Retinoic Acid Differentiated HL-60/S4 Cells with-or-without LBR Knockdown

Many functions of the nuclear envelope protein LBR (Lamin B Receptor) have been documented. LBR influences the shape of the nuclear envelope via its connections to underlying heterochromatin and to the lamina. LBR also functions as an essential enzyme in sterol biosynthesis. This study attempts to further define the LBR gene functions in a LBR knockdown cell line (HL-60/sh1), by employing Gene Set Enrichment Analysis (GSEA) to explore: Part 1) the effects of LBR knockdown on cell differentiation to granulocytes induced by retinoic acid (RA); Part 2) the effects of LBR knockdown on phenotypic differences between undifferentiated HL-60/sh1 and undifferentiated control cell lines (HL-60/S4 and HL-60/gfp). These GSEA studies are based upon previously published mRNA transcriptome data. In Part 1, the most significant effects were the increased loss of heterochromatin and the decreased histone methyltransferase activity in RA-differentiated HL-60/sh1 granulocytes, compared to HL-60/S4 and HL-60/gfp granulocytes. In addition, HL-60/sh1 ribosome structural protein transcripts were more increased during granulocyte differentiation, than observed in HL-60/S4 and HL-60/gfp. For many predicted phenotypes (e.g., senescence, migration, chemotaxis, phagocytosis and apoptosis), no significant differences were observed among the three granulocytic cell lines. In Part 2, comparisons were mainly between undifferentiated HL-60/sh1 and HL-60/S4 cells. We noted a significant increase in ribosomal protein and ribosomal RNA synthesis in sh1 0 versus to S4 0 cells. Of further interest, we observed that the position and number of nucleoli per cell appeared to differ between these two undifferentiated cell lines. In addition, GSEA results indicated a significant gain in heterochromatin and nucleosome formation in sh1 0 versus S4 0 cells. Microscopic imaging of undifferentiated sh1 0 cells compared to S4 0 cells indicated increased DAPI stained chromatin condensates surrounding the frequently central nucleoli, possibly reflecting the increased heterochromatin and the decreased LBR. Furthermore, the sh1 0 cell nuclei appeared "rounder" than the S4 0 cell nuclei. Evidence is presented supporting that the LINC Complex ("Linker of Nucleoskeleton and Cytoskeleton") may play a role.

cell biology↗

bioRxiv2024

The Multifaceted Phenotype of Senescent HL-60/S4 Macrophages

Every cell has a multifaceted phenotype. Transcriptional analysis of functionally defined groups of genes can provide insight into this phenotypic complexity. In the present study, the mRNA transcriptome of phorbol ester (TPA) differentiated HL-60/S4 macrophage cells was scrutinized using Gene Set Enrichment Analysis (GSEA), which evaluates the strengths of various cellular phenotypes by examining the enrichment of functionally different gene sets. Employing GSEA, we obtained supporting evidence that HL-60/S4 macrophages are senescent, probably a consequence of enriched TGF{beta} and NOTCH signaling transcripts. There appears to be a reduction of transcripts for heterochromatin, nucleosome formation, and chromatin remodeling phenotypes. In addition, despite upregulated oxidative stress gene transcription, we observed a reduction of DNA damage and repair transcripts. GSEA indicated that transcripts for autophagy, extracellular matrix, and inflammation/inflammasomes are enriched. We also observed that the HL-60/S4 macrophage is enriched for apoptosis gene transcripts, which may promote necrotic death by pyroptosis. The long-term goal of this research direction is to see whether this complex multifaceted phenotypic pattern is shared with other types of macrophages and to determine what mechanisms might exist to coordinate these phenotypic facets within a single cell.

cell biology↗

bioRxiv2019

Single-cell RNA-seq analysis reveals the crucial role of Collagen Triplex Helix Repeat Containing 1 (CTHRC1) cardiac fibroblasts for ventricular remodeling after myocardial infarction

Cardiac fibroblasts have a central role during the ventricular remodeling process associated with different types of cardiac injury. Recent studies have shown that fibroblasts do not respond homogeneously to heart damage, suggesting that the adult myocardium may contain specialized fibroblast subgroups with specific functions. Due to the limited set of bona fide fibroblast markers, a proper characterization of fibroblast population dynamics in response to cardiac damage is still missing. Using single-cell RNA-seq, we identified and characterized a fibroblast subpopulation that emerges in response to myocardial infarction (MI) in a murine model. These activated fibroblasts exhibit a clear pro-fibrotic signature, express high levels of the hormone CTHRC1 and of the immunomodulatory co-receptor CD200 and localize to the injured myocardium. Combining epigenomic profiling with functional assays, we show Sox9 and the non-canonical TGF-{beta} signaling as important regulators mediating their response to cardiac damage. We show that the absence of CTHRC1, in this activated fibroblast subpopulation, results in pronounced lethality due to ventricular rupture in a mouse model of myocardial infarction. Finally, we find evidence for the existence of similar mechanisms in a pig pre-clinical model of MI and establish a correlation between CTHRC1 levels and cardiac function after MI.

cell biology↗