Biology subjects

Porat, J.

Publications and source records attributed to Porat, J..

8 recordsLinked to original sources

bioRxiv2026

ERV-derived dsRNAs induce stress granules that drive neural crest fate to initiate melanoma

Melanoma arises when melanocytes are oncogenically reprogrammed into neural crest-like cells, but what initiates this switch is unknown. Here we show that endogenous retrovirus (ERV)-derived double stranded RNAs (dsRNAs) induce stress granules (SGs) that drive this fate change. In zebrafish, melanoma progresses through a stage-specific ERV switch: zferv marks precursor cells before crestin activates in tumor cells. Both elements generate dsRNA that triggers eif2ak2-dependent SGs within precursor lesions and through transformation. SGs are tumorigenic as G3BP1 overexpression accelerates melanoma onset, whereas mosaic, melanocyte-specific g3bp1/2 knockout delays it. Small molecule-mediated inhibition of G3BP1/2 eliminates precursor lesions and reduces tumor burden. Similarly, blocking dsRNA sensing prevents SG formation and lowers melanoma incidence. Human nevi and melanomas show analogous, stage-specific ERV programs and SG assembly. In human melanoma cells, SGs sequester and translationally repress SOX10-regulating mRNAs, including the tumor suppressor KMT2D. Mosaic knockout of kmt2d in vivo enhances SOX10-driven neural crest reactivation. Thus, an ERV-SG axis establishes an oncogenic translatome that enables melanoma fate, with diagnostic and therapeutic potential.

cancer biology↗

bioRxiv2026

Stochasticity in Metabolism: Oxidative Stress Response Phenotypes As Emergent Consequences of Network Topology

Reactive oxygen species (ROS) are ubiquitous yet elusive due to their reactivity and short half-lives, making simulation essential for mechanistic insight. Traditional ROS modeling has been confined to small ODE frameworks limited by kinetic-parameter scarcity. Here, we recast ROS dynamics as a flux-balance analysis (FBA) optimization problem using COBRA and CPLEX, augmenting the Escherichia coli iJO1366 genome-scale model with 84 spontaneous oxidation, 14 redox, and 9 detoxification reactions, targeting three ROS species: hydroxyl radicals, superoxide, and hydrogen peroxide (H2O2). We also formalize five fundamental rules of oxidation biology in the form of mixed-integer linear constraints, allowing us to dynamically incorporate reaction rate constants and let complex behavior emerge from first principles. Among these phenomena is the bimodal killing pattern under H2O2 stress, first announced over 40 years ago and never computationally recapitulated until now. We observe it by stochastically sampling our models with GapSplit, simulating bet-hedging and partitioning solutions into growers, persisters, and non-survivors. In our simulations, this killing effect arises solely from the metabolic network topology during random exploration of alternative metabolic configurations. The conservation of steady state creates a stochastic bias for detoxification when central carbon metabolism reaches its metabolic capacity to deal with ROS, causing the famous survival recovery phase. The recovery phase is thus part of Mode 1 metabolism, characterized by spare detoxification capacity and fulfilled by the Ahp reaction in E. coli. Mode 2 has no spare detoxification. We anticipate this to be an overlooked universal principle.

systems biology↗

bioRxiv2026

Unbiased, systematic surfaceome profiling of Acute Leukemia to identify novel immunotherapy targets

Cell surface proteomics provides a direct topological assessment of the outer membrane of cells and enables the capture of low abundance proteins that may be missed by whole cell proteomics. Here we present an unbiased atlas of the whole cell and surface proteomes of 25 commonly used leukemic cell lines, encompassing both lymphoid and myeloid lineages, and a variety of driver mutations. Paired-wise analysis highlights recurrent surface proteins that are not detected by whole cell proteomics. Coupling this dataset to RNA-sequencing, we also discovered genes where protein and RNA abundances are discordant. In KMT2A-rearranged AML, CD70 expression was increased across cell lines and validated in primary patient samples, supporting CD70 as a candidate therapeutic target in this disease. Several proteins are enriched in the surface proteomes but lack surface annotation, adding to the growing list of potential non-canonical cell surface proteins. These findings reveal a substantial pool of proteins absent from conventional surface annotations, including RNA-binding proteins, an emerging class of candidate immunotherapeutic targets.

molecular biology↗

bioRxiv2025

DNASE1L3 surveils mitochondrial DNA on the surface of distinct mammalian cells

The extracellular space is a critical environment for discriminating self versus non-self nucleic acids and initiating the appropriate immune responses through signaling cascades to relay information about extracellular nucleic acids. Here, we provide evidence that oxidized mitochondrial DNA is tethered to the surface of select mammalian cells through cell surface proteins and heparan sulfate proteoglycans. We demonstrate that cell surface DNA accumulates in large clusters that partially overlap with domains enriched in RNA binding proteins. Finally, we show that human and murine B cell surfaces contain DNA that can be cleared by the secreted nuclease DNASE1L3, and that patients with a DNASE1L3 missense variant associated with increased risk for autoimmune disease harbor increased levels of surface DNA on B and T cells. Taken together, this work expands the scope of cell surface nucleic acid biology and provides a mechanistic link between cell surface molecules and DNA targeting in autoimmune disease.

cell biology↗

bioRxiv2024

O-glycosylation contributes to mammalian glycoRNA biogenesis

There is an increasing appreciation for the role of cell surface glycans in modulating interactions with extracellular ligands and participating in intercellular communication. We recently reported the existence of sialoglycoRNAs, where mammalian small RNAs are covalently linked to N-glycans through the modified base acp3U and trafficked to the cell surface. However, little is currently known about the role for O-glycosylation, another major class of carbohydrate polymer modifications. Here, we use parallel genetic, enzymatic, and mass spectrometry approaches to demonstrate that O-linked glycan biosynthesis is responsible for the majority of sialoglycoRNA levels. By examining the O-glycans associated with RNA from cell lines and colon organoids we find known and previously unreported O-linked glycan structures. Further, we find that O-linked glycans released from small RNA from organoids derived from ulcerative colitis patients exhibit higher levels of sialylation than glycans from healthy organoids. Together, our work provides flexible tools to interrogate O-linked glycoRNAs (O-glycoRNA) and suggests that they may be modulated in human disease.

molecular biology↗

bioRxiv2023

The tRNA methyltransferase Trm1 promotes eukaryotic pre-tRNA maturation through catalytic and non-catalytic activities

tRNAs undergo an extensive maturation process involving post-transcriptional modifications often associated with tRNA structural stability and promoting the native fold. Impaired post-transcriptional modification has been linked to human disease, likely through defects in translation, mitochondrial function, and increased susceptibility to degradation by various tRNA decay pathways. More recently, evidence has emerged that bacterial tRNA modification enzymes can act as tRNA chaperones to guide tRNA folding in a manner independent from catalytic activity. Here, we provide evidence that the fission yeast tRNA methyltransferase Trm1, which dimethylates nuclear- and mitochondrial-encoded tRNAs at G26, can also promote tRNA functionality in the absence of catalysis. We show that wild type and catalytic-dead Trm1 are active in an in vivo tRNA-mediated suppression assay and possess RNA strand annealing and dissociation activity in vitro, similar to previously characterized RNA chaperones. Trm1 and the RNA chaperone La have previously been proposed to function synergistically in promoting tRNA maturation, yet we surprisingly demonstrate that La binding to nascent pre-tRNAs decreases Trm1 tRNA dimethylation in vivo and in vitro. Collectively, these results support the hypothesis for tRNA modification enzymes that combine catalytic and non-catalytic activities to promote tRNA maturation, as well as expand our understanding of how La function can influence tRNA modification.

molecular biology↗

bioRxiv2023

The fission yeast methylphosphate capping enzyme Bmc1/Bin3 promotes 2'-O- methylation of U6 and pre-mRNA splicing

Splicing requires the tight coordination of dynamic spliceosomal RNAs and proteins. U6 is the only spliceosomal RNA transcribed by RNA Polymerase III and undergoes an extensive maturation process. In humans and fission yeast, this includes addition of a 5 {gamma}-monomethyl phosphate cap by members of the Bin3/MePCE family. Previously, we have shown that the Bin3/MePCE homolog Bmc1 is recruited to the S. pombe telomerase holoenzyme by the LARP7 family protein Pof8, where it acts in a catalytic-independent manner to protect the telomerase RNA and facilitate holoenzyme assembly. Here, we show that Bmc1 and Pof8 also interact in a U6-containing snRNP. We demonstrate that Bmc1 and Pof8 promote 2-O-methylation of U6 and identify and characterize a non-canonical snoRNA that guides this methylation. Further, we show that fission yeast strains deleted of Bmc1 or Pof8 show altered U6 snRNP assembly patterns, supporting a more general role for these factors in guiding noncoding RNP assembly beyond the telomerase RNP. These results are thus consistent with a novel role for Bmc1/MePCE family members in stimulating U6 post-transcriptional modifications.

molecular biology↗

bioRxiv2021

The methyl phosphate capping enzyme Bin3 is a stable component of the fission yeast telomerase holoenzyme

The telomerase holoenzyme is critical for maintaining eukaryotic genome integrity. In addition to a reverse transcriptase and an RNA template, telomerase contains additional proteins that protect the telomerase RNA and promote holoenzyme assembly. Here we report that the methyl phosphate capping enzyme (MePCE) Bin3 is a stable component of the S. pombe telomerase holoenzyme. Bin3 associates with the telomerase holoenzyme and U6 snRNA through an interaction with the recently described LARP7 family member Pof8, and we demonstrate that these two factors are evolutionarily linked in fungi. Our data suggest that the association of Bin3 with telomerase is independent of its methyltransferase activity, but rather that Bin3 functions in telomerase holoenzyme assembly by promoting TER1 accumulation and Pof8 recruitment to TER1. Taken together, this work yields new insight into the composition, assembly, and regulation of the telomerase holoenzyme in fission yeast as well as the breadth of its evolutionary conservation.

molecular biology↗