Biology subjects

Lodewijk, G. A.

Publications and source records attributed to Lodewijk, G. A..

4 recordsLinked to original sources

bioRxiv2025

E2F1 induces a G0-G1 reentry transcriptional program without changing chromatin accessibility

Quiescent cells actively repress cell-cycle genes via chromatin-based mechanisms to maintain a non-dividing state, yet remain poised to reenter upon stimulation. E2F1, a canonical activator of cell-cycle genes, is sufficient to induce reentry from quiescence, but how it overcomes chromatin-mediated repression remains unclear. Here, we show that inducible E2F1 expression triggers exit from quiescence and progression through the cycle without changes in chromatin accessibility, by harnessing regulatory elements with limited, pre-existing accessibility. Using time-resolved transcriptomics, we demonstrate that E2F1 induces an accelerated transcriptional program compared to serum. Unlike serum, which triggers broad chromatin remodeling, E2F1-induced activation occurs in a context of limited accessibility. ChIP-seq reveals that E2F1 directly binds target sites in quiescent cells to upregulate canonical genes. Biochemical reconstitution shows that E2F1 binds nucleosomes and accesses internal E2F sites within histone-wrapped DNA. These findings suggest that E2F1 can engage nucleosome-associated DNA and initiate transcription without major chromatin reorganization, redefining transcription factor-chromatin dynamics during cell fate transitions and establishing E2F1 as a potent regulator of cell-cycle reentry.

cell biology↗

bioRxiv2024

Self-organization of embryonic stem cells into a reproducible embryo model through epigenome editing

Embryonic stem cells (ESCs) can self-organize in vitro into developmental patterns with spatial organization and molecular similarity to that of early embryonic stages. This self-organization of ESCs requires transmission of signaling cues, via addition of small molecule chemicals or recombinant proteins, to induce distinct embryonic cellular fates and subsequent assembly into structures that can mimic aspects of early embryonic development. During natural embryonic development, different embryonic cell types co-develop together, where each cell type expresses specific fate-inducing transcription factors through activation of non-coding regulatory elements and interactions with neighboring cells. However, previous studies have not fully explored the possibility of engineering endogenous regulatory elements to shape self-organization of ESCs into spatially-ordered embryo models. Here, we hypothesized that cell-intrinsic activation of a minimum number of such endogenous regulatory elements is sufficient to self-organize ESCs into early embryonic models. Our results show that CRISPR-based activation (CRISPRa) of only two endogenous regulatory elements in the genome of pluripotent stem cells is sufficient to generate embryonic patterns that show spatial and molecular resemblance to that of pre-gastrulation mouse embryonic development. Quantitative single-cell live fluorescent imaging showed that the emergence of spatially-ordered embryonic patterns happens through the intrinsic induction of cell fate that leads to an orchestrated collective cellular motion. Based on these results, we propose a straightforward approach to efficiently form 3D embryo models through intrinsic CRISPRa-based epigenome editing and independent of external signaling cues. CRISPRa-Programmed Embryo Models (CPEMs) show highly consistent composition of major embryonic cell types that are spatially-organized, with nearly 80% of the structures forming an embryonic cavity. Single cell transcriptomics confirmed the presence of main embryonic cell types in CPEMs with transcriptional similarity to pre-gastrulation mouse embryos and revealed novel signaling communication links between different embryonic cell types. Our findings offer a programmable embryo model and demonstrate that minimum intrinsic epigenome editing is sufficient to self-organize ESCs into highly consistent pre-gastrulation embryo models

developmental biology↗

bioRxiv2021

DeepSea: An efficient deep learning model for automated cell segmentation and tracking

Dynamics and non-genetic heterogeneity are two fundamental characteristics of basic processes of life such as cell division or differentiation. Time-lapse microscopy is the only method that can directly capture the dynamics and heterogeneity of fundamental cellular processes at the singlecell level with high temporal resolution. Successful application of single-cell time-lapse microscopy requires automated segmentation and tracking of hundreds of individual cells over several time points. Recently, deep learning models have ushered in a new era in the quantitative analysis of microscopy images. However, integrated segmentation and tracking of single cells remain challenges for the analysis of time-lapse microscopy images. This work presents a versatile and trainable deep-learning software, termed DeepSea, that allows for both segmentation and tracking of single cells in sequences of phase-contrast live microscopy images. Our segmentation model can easily be trained to segment phase-contrast images of different cell types with higher precision than existing models. Our tracking model allows for quantification of dynamics of several cell biological features of individual cells, such as cell division cycle, mitosis, cell morphology, and cell size, with high precision using phase-contrast images. We showcase the application of DeepSea by analyzing cell size regulation in embryonic stem cells. Our findings show that embryonic stem cells exhibit cell size control in the G1 phase of the cell cycle despite their unusual fast division cycle. Our training dataset, user-friendly software, and code are available here https://deepseas.org.

systems biology↗

bioRxiv2017

Human-specific NOTCH-like genes in a region linked to neurodevelopmental disorders affect cortical neurogenesis

Genetic changes causing dramatic brain size expansion in human evolution have remained elusive. Notch signaling is essential for radial glia stem cell proliferation and a determinant of neuronal number in the mammalian cortex. We find three paralogs of human-specific NOTCH2NL are highly expressed in radial glia cells. Functional analysis reveals different alleles of NOTCH2NL have varying potencies to enhance Notch signaling by interacting directly with NOTCH receptors. Consistent with a role in Notch signaling, NOTCH2NL ectopic expression delays differentiation of neuronal progenitors, while deletion accelerates differentiation. NOTCH2NL genes provide the breakpoints in typical cases of 1q21.1 distal deletion/duplication syndrome, where duplications are associated with macrocephaly and autism, and deletions with microcephaly and schizophrenia. Thus, the emergence of hominin-specific NOTCH2NL genes may have contributed to the rapid evolution of the larger hominin neocortex accompanied by loss of genomic stability at the 1q21. 1 locus and a resulting recurrent neurodevelopmental disorder.

neuroscience↗