Biology subjects

Kastelic, N.

Publications and source records attributed to Kastelic, N..

3 recordsLinked to original sources

bioRxiv2025

Epiproteomic control of the epitranscriptome drives cortical neurogenesis

Neurological conditions are the leading cause of ill health worldwide. Here, we show that the neurodevelopmental disorder-associated ubiquitin ligase UBE3C regulates the cellular composition of the murine cerebral cortex and human brain organoids, with its loss favoring neurogenesis and suppressing glial fate. Using genetic complementation, we demonstrate that disease-associated UBE3C mutations alter its autoubiquitination activity and disrupt cortical lamination. Proteomic profiling of UBE3C-deficient forebrains and organoids identifies Cbll1 as a UBE3C substrate, and we show that the UBE3C-Cbll1 duo drives N6-methyladenosine (m6A) mRNA methylation. Hyperactivation of m6A writers in UBE3C-deficient neural progenitors impairs cell cycle exit, a defect reversible in vivo by the METTL3 inhibitor STM2457. Our findings uncover an epiproteomic mechanism controlling m6A-mediated gene expression and define a regulatory axis linking ubiquitin signaling to epitranscriptomic control of neural fate. This work provides a mechanistic framework for understanding neurodevelopmental disorders and highlights potential therapeutic strategies.

cell biology↗

bioRxiv2024

Sensitive dissection of a genomic regulatory landscape using bulk and targeted single-cell activation

Transcriptional enhancers are non-coding DNA elements that regulate gene transcription in a temporal and tissue-specific manner. Despite advances in computational and experimental methods, identifying enhancers and their target genes essential for specific biological processes remains challenging. Determining target genes for enhancers is also complex and often relies on indirect, low-resolution, and/or assumptive methodologies. To identify and functionally perturb enhancers at their endogenous sites without altering their sequence, we performed a pooled tiling CRISPR activation (CRISPRa) screen surrounding PHOX2B, a master regulator of neuronal cell fate and a key player in neuroblastoma development. This screen allowed the de novo identification of CRISPRa responsive elements (CaREs) that alter cellular growth within the 2 Mb genomic region. To determine CaRE target genes, we developed TESLA-seq (TargEted SingLe cell Activation), which combines CRISPRa screening with targeted single-cell RNA-sequencing and enables the parallel readout of the effect of hundreds of enhancers on all genes in the locus. While most TESLA-revealed CaRE-gene relationships involved neuroblastoma-related regulatory elements already active in the system, we found many CaREs and target connections normally active only in other tissue types or with no previous evidence and induced out of context by CRISPRa. This highlights the power of TESLA-seq to reveal gene regulatory networks, including edges active outside of a given experimental system. HighlightsO_LISystematically perturbed regulatory landscape in a 2 Mb genomic region surrounding PHOX2B to identify hundreds of CRISPRa-responsive elements that affect cellular growth C_LIO_LIDeveloped TESLA-seq as a principled molecular approach to find gene targets of dozens of candidate regulatory elements C_LIO_LIValidated interactions between identified regulatory elements and target genes and characterized their genomic features C_LIO_LIIntegrated a compendium of epigenomic datasets to identify regulatory relationships induced out of context or with no previous evidence C_LI

genomics↗

bioRxiv2018

Quantifying multi-layered expression regulation in response to stress of the endoplasmic reticulum

The mammalian response to endoplasmic reticulum (ER) stress dynamically affects all layers of gene expression regulation. We quantified transcript and protein abundance along with footprints of ribosomes and non-ribosomal proteins for thousands of genes in cervical cancer cells responding to treatment with tunicamycin or hydrogen peroxide over an eight hour time course. We identify shared and stress-specific significant regulatory events at the transcriptional and post-transcriptional level and at different phases of the experiment. ER stress regulators increase transcription and translation at different times supporting an adaptive response. ER stress also induces translation of genes from serine biosynthesis and one-carbon metabolism indicating a shift in energy production. Discordant regulation of DNA repair genes suggests transcriptional priming in which delayed translation fine-tunes the early change in the transcriptome. Finally, case studies on stress-dependent alternative splicing and protein-mRNA binding demonstrate the ability of this resource to generate hypotheses for new regulatory mechanisms.

systems biology↗