Biology subjects

Peterson, K. A.

Publications and source records attributed to Peterson, K. A..

8 recordsLinked to original sources

bioRxiv2026

Targeting SUFU-mediated GLI activity alleviates ciliopathic phenotypes

Ciliopathies are incurable, pleiotropic diseases that result from defective cilia. Several ciliopathic phenotypes arise from improper processing and activity of GLI transcription factors, yet attempts to rescue these phenotypes by overexpressing activator or repressor GLI isoforms have proven unsuccessful. Here, we demonstrate that modulating SUFU-GLI interactions restores GLI transcriptional activity and rescues ciliopathic phenotypes in vivo. As previous work identified increased accumulation of SUFU in ciliopathic conditions, we hypothesized that exploiting SUFU's epistatic relationship to the cilium could provide a novel therapeutic strategy for ciliopathies. Genetic introduction of either a single null allele (Sufu+/-) or point mutation allele (SufuD159R), which modulates SUFU-GLI binding, into a ciliopathic mouse model revealed that both interventions improved midfacial phenotypes; however, SufuD159R provided a more complete rescue by significantly improving mandibular phenotypes as well. Molecular examination of this genetic modification revealed restored GLI3 binding to DNA, GLI3 interaction with transcriptional cofactors, and GLI3-directed transcription of target genes, collectively driving the rescue of numerous ciliopathic phenotypes. These findings reveal cilium-independent functions for SUFU and establish that modulating SUFU-GLI interactions bypasses the need for ciliary GLI processing. Together, these data provide a promising therapeutic approach for ameliorating ciliopathic pathologies.

developmental biology↗

bioRxiv2025

A dual role for GLI3 signaling in neural crest development

Neural crest cells (NCCs) are a multipotent cell population that undergo specification, epithelial-to-mesenchymal transition, migration, and differentiation into a plethora of cell types. A wealth of studies across various embryonic model systems have established dogma as to the molecular mechanisms and signaling cascades that contribute to NCC development. While Wnt, FGF, and BMP signaling pathways have well-established and essential roles in several aspects of NCC development, the Hedgehog (HH) signaling pathway has received limited attention for any specific role in this process. Herein, we propose two distinct, temporal roles for the transcription factor GLI3 in NCC development. Gli3, and other members of the HH pathway, were robustly co-expressed with established NCC induction and specification markers in chick, mouse, and human embryonic stem cell derived NCCs. Early knock-down of GLI3 reduced expression of key markers of NCC specification and conditional knock-out of Gli3 post-specification specifically impaired the ability of cranial NCCs to differentiate into ectomesenchymal derivatives. Together, these results demonstrate dual, novel roles for GLI3 in early NCC specification and later in cranial NCC differentiation. Summary StatementGLI3 plays distinct temporal roles in neural crest specification and differentiation into ectomesenchymal derivatives.

developmental biology↗

bioRxiv2025

Interaction of Wnt and SHH gradients synchronizes cell cycle 1 exit and differentiation

Classic models of the French flag problem depict sharp cell-type boundaries emerging from threshold responses to morphogen gradients. Yet, how such boundaries arise during dynamic cell-state transitions remains unclear. We use hair follicle dermal condensates to study a sharp cell-type transition in which proliferative progenitors synchronously undergo cell-cycle exit and molecular differentiation. Using genetic and genomic approaches, we show that Wnt and Hedgehog signaling interact to coordinate the timing of these processes. When their activities are temporally aligned, intermediate transitional states are compressed through cell-cycle exit, producing a sharp cell-type boundary; when misaligned, transitional states expand, yielding fuzzy borders. Mechanistically, elevated Wnt activity promotes cell-cycle exit by regulating chromatin binding of the Hedgehog mediator GLI3. Hedgehog signaling induces differentiation genes in a Wnt-dependent manner and simultaneously elevates Wnt activity, aligning arrest and differentiation in time. These findings reveal a mechanism wherein morphogen interactions coordinate transitions to generate precise cell-type boundaries.

developmental biology↗

bioRxiv2025

Generation and Characterization of a Knockout Mouse of an Enhancer of EBF3

Genomic studies of autism and other neurodevelopmental disorders have identified several relevant protein-coding and noncoding variants. One gene with an excess of protein-coding de novo variants is EBF3 that also is the gene underlying the Hypotonia, Ataxia, and Delayed Development Syndrome (HADDS). In previous work, we have identified noncoding de novo variants in an enhancer of EBF3 called hs737 and further showed that there was an enrichment of deletions of this enhancer in individuals with neurodevelopmental disorders. In this present study, we generated a novel mouse line that deletes the highly conserved, orthologous mouse region of hs737 within the Rr169617 regulatory region, and characterized the molecular and phenotypic aspects of this mouse model. This line contains a 1,160 bp deletion within Rr169617 and through heterozygous crosses we found a deviation from Mendelian expectation (p = 0.02) with a significant depletion of the deletion allele (p = 5.8 x 10-4). Rr169617+/- mice had a reduction of Ebf3 expression by 10% and Rr169617-/- mice had a reduction of Ebf3 expression by 20%. Differential expression analyses in E12.5 forebrain, midbrain, and hindbrain in Rr169617+/+versus Rr169617-/- mice identified dysregulated genes including histone genes (i.e., Hist1h1e, Hist1h2bk, Hist1h3i, Hist1h2ao) and other brain development related genes (e.g., Chd5, Ntng1). A priori phenotyping analysis (open field, hole board and light/dark transition) identified sex-specific differences in behavioral traits when comparing Rr169617-/- males versus females; whereby, males were observed to be less mobile, move slower, and spend more time in the dark. Furthermore, both sexes when homozygous for the enhancer deletion displayed body composition differences when compared to wild-type mice. Overall, we show that deletion within Rr169617 reduces the expression of Ebf3 and results in phenotypic outcomes consistent with potential sex specific behavioral differences. This enhancer deletion line provides a valuable resource for others interested in noncoding regions in neurodevelopmental disorders and/or those interested in the gene regulatory network downstream of Ebf3.

genetics↗

bioRxiv2021

The production of 4,182 mouse lines identifies experimental and biological variables impacting Cas9-mediated mutant mouse line production

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3,313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. A systematic evaluation of the variables impacting success rates identified gene essentiality as the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for using Cas9 to generate alleles in mouse essential genes, many of which are orthologs of genes linked to human disease.

genetics↗

bioRxiv2021

Whole genome analysis for 163 guide RNAs in Cas9 edited mice reveals minimal off-target activity

Introductory ParagraphThe Knockout Mouse Phenotyping Program (KOMP2) uses CRISRPR/Cas9 for high-throughput mouse line production to generate null alleles in the inbred C57BL/6N strain for broad-based in vivo phenotyping. In order to assess the risk of spurious S. pyogenes Cas9-induced off-target mutagenesis, we applied whole genome sequencing to compare the genomes of 50 Cas9-derived founder mice representing 163 different gRNAs to 28 untreated inbred control mice. Our analysis pipeline detected 28 off-target sequence variants associated with 21 guides. These potential off-targets were identified in 18/50 (36%) founders with 9/28 (32%) independently validated corresponding to 8 founder animals. In total, only 4.9% (8/163) of all guides exhibited off-target activity resulting in a rate of 0.16 Cas9 off-target mutations per founder analyzed. In comparison, we observed ~1225 unique variants in each mouse regardless of whether or not it was exposed to Cas9. These findings indicate that Cas9-mediated off-target mutagenesis is rare in founder knockout mice generated using guide RNAs designed to minimize off-target risk. Overall, bona fide off-target variants comprise a small fraction of the genetic heterogeneity found in carefully maintained colonies of inbred strains.

genetics↗

bioRxiv2020

Gli3 utilizes Hand2 to synergistically regulate tissue-specific transcriptional networks

Despite a common understanding that Gli TFs are utilized to reiterate a Hh morphogen gradient, genetic analyses suggest craniofacial development does not completely fit this paradigm. We demonstrated that rather than being driven by a Hh threshold, robust Gli3 transcriptional activity during skeletal and glossal development required interaction with the bHLH TF Hand2. Not only did genetic and expression data support a co-factorial relationship, but genomic analysis further revealed that Gli3 and Hand2 were enriched at regulatory elements for genes essential for mandibular patterning and development. Interestingly, motif analysis at sites co-occupied by Gli3 and Hand2 uncovered mandibular-specific, low-affinity, divergent Gli binding motifs (dGBMs). Functional validation revealed these dGBMs conveyed synergistic activation of Gli targets essential for mandibular patterning and development. In summary, this work elucidates a novel, sequence-dependent mechanism for Gli transcriptional activity within the craniofacial complex that is independent of a graded Hh signal.

developmental biology↗

bioRxiv2019

Human and mouse essentiality screens as a resource for disease gene discovery

Although genomic sequencing has been transformative in the study of rare genetic diseases, identifying causal variants remains a considerable challenge that can be addressed in part by new gene-specific knowledge. Here, we integrate measures of how essential a gene is to supporting life, as inferred from the comprehensive viability and phenotyping screens performed on knockout mice by the International Mouse Phenotyping Consortium and from human cell line essentiality screens. We propose a novel, cross-species gene classification across the Full Spectrum of Intolerance to Loss-of-function (FUSIL) and demonstrate that genes in five mutually exclusive FUSIL categories have differing characteristics in the biological processes they regulate, tissue expression levels and human mutation rates. Most notably, Mendelian disease genes, particularly those associated with developmental disorders, are highly overrepresented in the developmental lethal category, representing genes not essential for cell survival but required for organism development. Exploiting this finding, we have screened developmental disorder cases from three independent disease sequencing consortia and identified potentially pathogenic, de novo variants shared in different patients for several developmental lethal genes that have not previously been associated with rare disease. We therefore propose FUSIL as an efficient resource for disease gene discovery.

genomics↗