Biology subjects

Granot, Z.

Publications and source records attributed to Granot, Z..

6 recordsLinked to original sources

bioRxiv2026

Targeting the 3 splice site by a decoy oligonucleotide attenuates U2AF1 splicing activity and inhibits leukemia

Recurrent mutations in spliceosomal genes are a hallmark of myeloid malignancies, with SF3B1, SRSF2, U2AF1 and ZRSR2 among the most frequently affected. These alterations are typically heterozygous, mutually exclusive missense mutations targeting highly conserved residues, reflecting a selective pressure to maintain a dysregulated yet essential splicing machinery. This constraint suggests that leukemic cells remain dependent on residual splicing activity, exposing a potential therapeutic vulnerability that extends beyond genetically defined subsets. For example, a previously developed therapeutic, Pladienolide B, is a potent cancer cell growth inhibitor targeting the SF3B1 subunit of the spliceosome1,2. Here we present an RNA decoy-based strategy to disrupt 3' splice site recognition by competitively engaging components of the spliceosomal machinery. We engineered a chemically stabilized RNA decoy that mimics the 3 splice site (3'SS decoy), thereby sequestering proteins involved in 3 splice site recognition from endogenous pre-mRNA targets. Although the decoy is expected to engage multiple components of the 3' splice site recognition complex, U2AF1 was used as the primary molecular readout to assess target engagement and downstream effects. To enable intracellular delivery, decoys were encapsulated in lipid nanoparticles (LNPs), facilitating efficient uptake in leukemic systems. We show that LNP-encapsulated decoys are efficiently delivered into leukemic cells, including established cell lines and patient-derived blasts, and directly engage components of the splicing machinery. Decoy treatment induces widespread alterations in RNA splicing programs and impairs leukemic cell fitness in vitro. Importantly, systemic administration of the 3SS decoy significantly reduces leukemia burden in an in vivo xenograft model. Notably, these effects are observed independently of spliceosomal mutational status, supporting a broader dependency of leukemic cells on intact splicing factor function. Together, our findings establish decoy-mediated disruption of splicing factor activity as a mechanistically targeted therapeutic strategy and identify LNPs as an effective platform for the delivery of RNA-based modulators of essential RNA-protein interactions in myeloid malignancies.

molecular biology↗

bioRxiv2021

RNA interference mediates RNA toxicity with parent-of-origin effects in C. elegans expressing CTG repeats

Nucleotide repeat expansions are a hallmark of over 40 neurodegenerative diseases. These repeats cause RNA toxicity and trigger multisystemic symptoms that worsen with age. RNA toxicity can trigger, through an unclear mechanism, severe disease manifestation in infants that inherited repeats from their mothers. Here we show in Caenorhabditis elegans how RNA interference machinery causes intergenerational toxicity through inheritance of siRNAs derived from CUG repeats. The maternal repeat-derived small RNAs cause transcriptomic changes in the offspring, reduce motility and shorten lifespan. However, the toxicity phenotypes in the offspring can be rescued by perturbing the RNAi machinery in affected mothers. This points to a novel mechanism linking maternal bias and the RNAi machinery and suggests that toxic RNA is transmitted to offspring and causes disease phenotypes through intergenerational epigenetic inheritance.

molecular biology↗

bioRxiv2021

Neutrophils impose strong selective pressure against PfEMP1 variants implicated in cerebral malaria

Plasmodium falciparum, the deadliest form of human malaria, remains one of the major threats to human health in endemic regions. Its virulence is attributed to its ability to modify infected red blood cells (iRBC) to adhere to endothelial receptors by placing variable antigens known as PfEMP1 on the surface of the red cell. PfEMP1 expression on the red cell surface determines the cytoadhesive properties of the iRBCs and is implicated in severe manifestations of malaria. To evade antibody mediated responses the parasite undergoes continuous switches of expression between different PfEMP1 variants. Recently it became clear that in addition to antibody mediated responses, PfEMP1 triggers an innate immune response, however, the role of neutrophils, the most abundant white blood cells in the human circulation, in malaria remains elusive. Here we show that neutrophils recognize and kill blood stages of several P. falciparum isolates, and we identify neutrophil ICAM-1 and specific PfEMP1s implicated in cerebral malaria as the key molecules involved in this killing. Our data provide mechanistic insight into the interactions between neutrophils and iRBCs and demonstrate the important influence of PfEMP1 on the selective innate response to cerebral malaria.

microbiology↗

bioRxiv2021

Mapping cellular subpopulations within triple negative breast cancer tumors provides a tool for cancer sensitization to radiotherapy

Triple negative breast cancer (TNBC) is an aggressive type of cancer that is known to be resistant to radiotherapy (RT). Evidence is accumulating that is indicative of the plasticity of TNBC, where one cancer subtype switches to another in response to various treatments, including RT. In this study we aim to overcome tumor resistance by designing TNBC-sensitizing targeted therapies that exploit the plasticity occurring due to radiation exposure. Using single cell analysis of molecular changes occurring in irradiated TNBC tumors, we identified two initially undetected distinct subpopulations, represented by overexpressed Her2 and cMet, expanding post-RT and persisting in surviving tumors. Using murine cancer models and patient-derived TNBC tumors, we showed that only simultaneous targeting of Her2 and cMet was successful in sensitizing TNBC to RT and preventing its regrowth. The strategy presented herein holds the potential to be broadly applicable in clinical use. HighlightsO_LISensitization of TNBC to radiotherapy (RT) is a clinically unmet need C_LIO_LISingle cell strategy creates a precise map of subpopulations expanding post-RT C_LIO_LIEvolution of intra-tumor heterogeneity is turned into a therapeutic advantage C_LIO_LISimultaneous targeting of expanding subpopulations sensitizes TNBC to radiotherapy C_LI

cancer biology↗

bioRxiv2020

The PD-L1/PD-1 Axis Blocks Neutrophil Cytotoxicity In Cancer

The PD-L1/PD-1 axis was shown to promote tumor growth and progression via the inhibition of anti-tumor immunity. Blocking this axis was shown to be beneficial in maintaining the anti-tumor functions of the adaptive immune system. Still, the consequences of blocking the PD-L1/PD-1 axis on innate immune responses remain largely unexplored. In this context, neutrophils were shown to consist of different subpopulations, which possess either pro- and anti-tumor properties. PD-L1-expressing neutrophils are considered pro-tumor as they can suppress cytotoxic T-cells. That said, we found that PD-L1 expression is not limited to tumor promoting neutrophils but is also evident in anti-tumor neutrophils. We show that neutrophil cytotoxicity is effectively and efficiently blocked by tumor cell expressed PD-1. Furthermore blocking of either neutrophil PD-L1 or tumor cell PD-1 maintains neutrophil cytotoxicity resulting in enhanced tumor cell apoptosis. Importantly, we show that tumor cell PD-1 blocks neutrophil cytotoxicity and promotes tumor growth via a mechanism independent of adaptive immunity. Taken together, these findings highlight the therapeutic potential of enhancing anti-tumor innate immune responses via blocking of the PD-L1/PD-1 axis.

immunology↗

bioRxiv2019

IRE1-UBE2D3 signaling controls the recruitment of myeloid cells to glioblastoma.

Tumor cells are exposed to intrinsic and environmental challenges that trigger endoplasmic reticulum (ER) homeostasis alteration, in turn leading to ER stress. To cope with this, tumor cells engage an adaptive signaling pathway, the unfolded protein response (UPR) thus promoting the acquisition of malignant features. As such, glioblastoma multiforme (GBM), the most aggressive primary brain tumors, exhibit constitutive UPR signals to sustain growth. Herein, we showed that signaling elicited by one of the UPR sensors, IRE1, promotes GBM tumor invasion, angiogenesis and infiltration by macrophages. Hence, high IRE1 activity in tumors predicts worse outcome. We further dissect IRE1-dependent mechanisms that shape the brain tumor immune microenvironment towards myeloid cells. We identify an IRE1-dependent signaling pathway that directly controls the expression/release of proinflammatory chemokines (CXCL2, IL6, IL8) leading to tumor cell-mediated chemoattraction of neutrophils and macrophages. This pathway requires XBP1 non-conventional mRNA splicing and XBP1s-dependent expression of the E2 ubiquitin enzyme UBE2D3. The latter contributes to the degradation of the NF{kappa}B inhibitor I{kappa}B, leading to the up-regulation of proinflammatory chemokines. Our work identifies a novel IRE1/UBE2D3 proinflammatory signaling axis instrumental to pro-tumoral immune regulation of GBM.

cancer biology↗