Biology subjects

Van Otterloo, E.

Publications and source records attributed to Van Otterloo, E..

8 recordsLinked to original sources

bioRxiv2024

Epidermal loss of PRMT5 leads to the emergence of an atypical basal keratinocyte-like cell population and defective skin stratification

During skin development, ectoderm-derived cells undergo precisely coordinated proliferation, differentiation, and adhesion to yield stratified epidermis. Disruptions in these processes can result in congenital anomalies including ectodermal dysplasia and harlequin ichthyosis. Protein Arginine Methyl Transferase 5 (PRMT5)--an enzyme responsible for methylating arginine residues in histones and other proteins--maintains progenitor status in germ and limb bud cells. Similarly, in vitro evidence suggests that PRMT5 prevents differentiation of basal keratinocytes, leading us to hypothesize that PRMT5 preserves the stem-cell phenotype of keratinocytes in vivo. To test this possibility, we generated conditional knockout (cKO) mice lacking Prmt5 in early ectoderm (E7.5), impacting the entire epidermis. Prmt5 cKOs exhibited gross skin defects, compromised skin barrier function, and reduced postnatal viability. Histological analyses revealed significant defects in epidermal stratification, without alterations in apoptosis or proliferation. Single-cell RNA and ATAC-seq analysis identified an atypical population of basal keratinocyte-like cells in Prmt5 cKOs, that exhibited a senescence-like program, characterized by increased Cdkn1a (p21), elevated senescence-associated secretory phenotype (SASP) molecules (Igfbp2), and decreased developmental transcription factor (Trp63) expression. Our findings suggest that PRMT5 prevents basal keratinocyte senescence by repressing Cdkn1a, shedding light on the epigenetic regulation of basal keratinocyte maintenance and senescence in congenital skin disorders.

developmental biology↗

bioRxiv2023

Computational identification of key transcription factors for embryonic and postnatal Sox2+ dental epithelial stem cell

While many reptiles can replace their tooth throughout life, human loss the tooth replacement capability after formation of the permanent teeth. It was thought that the difference in tooth regeneration capability depends on the persistence of a specialized dental epithelial structure, the dental lamina that contains dental epithelial stem cells (DESC). Currently, we know very little about DESC such as what genes are expressed or its chromatin accessibility profile. Multiple markers of DESC have been proposed such as Sox2 and Lgr5. Few single cell RNA-seq experiments have been performed previously, but no obvious DESC cluster was identified in these scRNA-seq datasets, possible due to that the expression level of DESC markers such as Sox2 and Lgr5 is too low or the percentage of DESC is too low in whole tooth. We utilize a mouse line Sox2-GFP to enrich Sox2+ DESC and use Smart-Seq2 protocol and ATAC-seq protocol to generate transcriptome profile and chromatin accessibility profile of P2 Sox2+ DESC. Additionally, we generate transcriptome profile and chromatin accessibility profile of E11.5 Sox2+ dental lamina cells. With transcriptome profile and chromatin accessibility profile, we systematically identify potential key transcription factors for E11.5 Sox2+ cells and P2 Sox2+ cells. We identified transcription factors including Pitx2, Id3, Pitx1, Tbx1, Trp63, Nkx2-3, Grhl3, Dlx2, Runx1, Nfix, Zfp536, etc potentially formed the core transcriptional regulatory networks of Sox2+ DESC in both embryonic and postnatal stages.

developmental biology↗

bioRxiv2023

TFAP2 paralogs regulate midfacial development in part through a conserved ALX genetic pathway.

Cranial neural crest development is governed by positional gene regulatory networks (GRNs). Fine-tuning of the GRN components underly facial shape variation, yet how those in the midface are connected and activated remain poorly understood. Here, we show that concerted inactivation of Tfap2a and Tfap2b in the murine neural crest even during the late migratory phase results in a midfacial cleft and skeletal abnormalities. Bulk and single-cell RNA-seq profiling reveal that loss of both Tfap2 members dysregulated numerous midface GRN components involved in midface fusion, patterning, and differentiation. Notably, Alx1/3/4 (Alx) transcript levels are reduced, while ChIP-seq analyses suggest TFAP2 directly and positively regulates Alx gene expression. TFAP2 and ALX co-expression in midfacial neural crest cells of both mouse and zebrafish further implies conservation of this regulatory axis across vertebrates. Consistent with this notion, tfap2a mutant zebrafish present abnormal alx3 expression patterns, and the two genes display a genetic interaction in this species. Together, these data demonstrate a critical role for TFAP2 in regulating vertebrate midfacial development in part through ALX transcription factor gene expression.

developmental biology↗

bioRxiv2022

Transcriptional programs controlling lineages specification of mandibular epithelium during tooth initiation

Tooth development starts with formation of the dental lamina, a localized thickening of the maxillary and mandibular epithelium, at specific location of oral cavity. How this dental lamina is specified within the continuous naive maxillary and mandibular epithelium, remains as a critical and unresolved question. To identify potential genes and transcriptional regulatory networks that controlling dental lamina formation, we utilized single-cell Multiome-seq and Laser Microdissection coupled RNA-seq to profile gene expression and open chromatin of mandibular epithelium during tooth initiation. We comprehensively identified transcription factors (TFs) and signaling pathways that are differentially expressed among different domains (including dental lamina) along dorsal-ventral axis of mandibular epithelium. Specifically, we found Sox2 and Tfap2a/Tfap2b forming two complementary domains along the dorsal-ventral axis of mandibular epithelium. Furthermore, dental lamina specific or enriched TFs such as Pitx1, Pitx2 and Zfp536 are presented around the interface of Sox2 and Tfap2a/Tfap2b. We also identified correlation between signaling pathways and domain specific TFs. Through computational analysis of single-cell Multiome-seq dataset, we identified potential key/driver TFs and core transcriptional regulatory networks (TRNs) for lineages (oral, dental, skin) specification during tooth initiation. Our computational analysis also reveals a potential cross repression interactions between different groups of TFs (i.e. Pitx1/Pitx2 and Tfap2a/Tfap2b) that might be essential for dental lamina specification.

developmental biology↗

bioRxiv2021

TFAP2 paralogs pioneer chromatin access for MITF and directly inhibit genes associated with cell migration

In developing melanocytes and in melanoma cells, multiple paralogs of the Activating-enhancer-binding Protein 2 family of transcription factors (TFAP2) contribute to expression of genes encoding pigmentation regulators, but their interaction with Microphthalmia transcription factor (MITF), a master regulator of these cells, is unclear. Supporting the model that Tfap2 facilitates MITFs ability to activate expression of pigmentation genes, single-cell seq analysis of zebrafish embryos revealed that pigmentation genes are only expressed in the subset of mitfa- expressing cells that also express Tfap2 paralogs. To test this model in SK-MEL-28 melanoma cells we deleted the two TFAP2 paralogs with highest expression, TFAP2A and TFAP2C, creating TFAP2 knockout (TFAP2-KO) cells. We then assessed gene expression, chromatin accessibility, binding of TFAP2A and of MITF, and the chromatin marks H3K27Ac and H3K27Me3 which are characteristic of active enhancers and silenced chromatin, respectively. Integrated analyses of these datasets indicate TFAP2 paralogs directly activate enhancers near genes enriched for roles in pigmentation and proliferation, and directly repress enhancers near genes enriched for roles in cell adhesion. Consistently, compared to WT cells, TFAP2-KO cells proliferate less and adhere to one another more. TFAP2 paralogs and MITF co-operatively activate a subset of enhancers, with the latter necessary for MITF binding and chromatin accessibility. By contrast, TFAP2 paralogs and MITF do not appear to co-operatively inhibit enhancers. These studies reveal a mechanism by which TFAP2 profoundly influences the set of genes activated by MITF, and thereby the phenotype of pigment cells and melanoma cells.

developmental biology↗

bioRxiv2021

AP-2α and AP-2β cooperatively function in the craniofacial surface ectoderm to regulate chromatin and gene expression dynamics during facial development.

The facial surface ectoderm is essential for normal development of the underlying cranial neural crest cell populations, providing signals that direct appropriate growth, patterning, and morphogenesis. Despite the importance of the ectoderm as a signaling center, the molecular cues and genetic programs implemented within this tissue are understudied. Here we show that removal of two members of the AP-2 transcription factor family, AP-2 and AP-2{beta}, within the early embryonic ectoderm leads to major alterations in the mouse craniofacial complex. Significantly, there are clefts in both the upper face and mandible, accompanied by fusion of the upper and lower jaws in the hinge region. Comparison of ATAC-seq and RNA-seq analyses between controls and mutants revealed significant changes in chromatin accessibility and gene expression centered on multiple AP-2 binding motifs associated with enhancer elements within these ectodermal lineages. In particular, loss of these AP-2 proteins affects both skin differentiation as well as multiple signaling pathways, most notably the WNT pathway. The role of reduced Wnt signaling throughput in the mutant phenotype was further confirmed using reporter assays and rescue experiments involving Wnt1 ligand overexpression. Collectively, these findings highlight a conserved ancestral function for AP-2 transcription factors in ectodermal development and signaling, and provide a framework from which to understand the gene regulatory network operating within this tissue that directs vertebrate craniofacial development.

developmental biology↗

bioRxiv2020

Anomalous incisor morphology indicates tissue-specific roles for Tfap2a and Tfap2b in tooth development

Mice possess two types of teeth that differ in their cusp patterns; incisors have one cusp and molars have multiple cusps. The patterning of these two types of teeth relies on fine-tuning of the reciprocal molecular signaling between dental epithelial and mesenchymal tissues during embryonic development. Here we show that the incisors are populated only at early time points by the neural crest, whereas the molars continue to receive contributions at later stages, revealing a temporal difference that could alter epithelial-mesenchymal signaling dynamics between these two types of teeth. The AP-2 transcription factors, particularly Tfap2a and Tfap2b, are essential components of such epithelial-mesenchymal signaling interactions that coordinate craniofacial development in mice and other mammals, but little is known about their roles in the regulation of tooth development and shape. We demonstrate that incisors and molars differ in their temporal and spatial expression of Tfap2a and Tfap2b; in particular, at the bud stage, Tfap2a is expressed in both the epithelium and mesenchyme of the incisors and molars but expression of Tfap2b is restricted to the mesenchyme of the molars. Tissue-specific deletions show that loss of the epithelial domain of Tfap2a and Tfap2b affects the number and spatial arrangement of the incisors, notably resulting in duplicated lower incisors. In contrast, deletion of these two genes in the mesenchymal domain has little effect on tooth development. Collectively these results implicate epithelial expression of Tfap2a and Tfap2b in dorsal-ventral patterning of the incisors and suggest that these genes contribute to morphological differences between anterior (incisor) and posterior (molar) teeth within the mammalian dentition. HighlightsO_LILate-migrating cranial neural crest cells contribute extensively to the developing molar tooth germs but minimally to the incisors. C_LIO_LIDuring tooth development, transcription factors Tfap2a and Tfap2b are expressed in spatially and temporally dynamic patterns and differ between incisor and molar tooth germs. C_LIO_LIEpithelial expression of Tfap2a and Tfap2b is necessary for incisor development, but mesenchymal expression of these genes is not required. C_LI

developmental biology↗

bioRxiv2018

Differential 3’ Processing of Specific Transcripts Expands Regulatory and Protein Diversity Across Neuronal Cell Types

Alternative polyadenylation (APA) regulates mRNA translation, stability, and protein localization. However, it is unclear to what extent APA regulates these processes uniquely in specific cell types. Using a new technique, cTag-PAPERCLIP, we discovered significant differences in APA between the principal types of mouse cerebellar neurons, the Purkinje and granule cells, as well as between proliferating and differentiated granule cells. Transcripts that differed in APA in these comparisons were enriched in key neuronal functions and many differed in coding sequence in addition to 3UTR length. We characterize Memo1, a transcript that shifted from expressing a short 3UTR isoform to a longer one during granule cell differentiation. We show that Memo1 regulates granule cell precursor proliferation and that its long 3UTR isoform is targeted by miR-124, contributing to its downregulation during development. Our findings provide insight into roles for APA in specific cell types and establish a platform for further functional studies.

neuroscience↗