Biology subjects

Feldman, B.

Publications and source records attributed to Feldman, B..

3 recordsLinked to original sources

bioRxiv2025

Developmental regulation of intestinal best4+ cells

best4+/CFTR-high expressing cells are a recently described intestinal epithelial cell type potentially altered in inflammatory bowel disease and colorectal cancer. However, their developmental origin, developmental regulation, and functions remain undefined. This study identifies their conserved transcriptional program and uses zebrafish to dissect their developmental regulation in vivo. Lineage tracing identified that best4+ cells arise from atoh1b+ secretory progenitors. We identify that Notch signaling, mediated by dll4, specifies best4+ cells at the expense of enterochromaffin cells. Downstream of Notch, meis1b confers best4+ cell identity. best4+ cells then exhibit regionalized gene expression, regulated by pbx3a. Additionally, this study demonstrates a system where best4+ cells can be manipulated, observed, and removed in an organismal context. Live imaging and electron microscopy of best4+ cells identified dynamic cellular projections, suggesting a sensory or communicative function. Removal of best4+ cells in vivo eliminated previously proposed functions: they are not required to restore intestinal pH following acidic challenge and do not absorb nutrients. However, we identify region-specific intracellular pH differences that suggest potential functional heterogeneity. Altogether, this study presents a comprehensive description of best4+ cell development from birth to spatial regulation that will be instrumental to understand how best4+ cells change in disease or might be therapeutically manipulated and presents the tools to dissect their function in vivo.

developmental biology↗

bioRxiv2025

Elevated body temperature exacerbates arrhythmia and seizure-like activity in a zebrafish model of Timothy syndrome

Timothy syndrome (TS) is a multisystem disorder with autistic-like features, seizures and arrhythmia as the main symptoms. Most TS cases are caused by a de novo single amino acid substitution G406R in the CACNA1C gene that encodes the pore-forming subunit of the voltage-gated L-type calcium channel CaV1.2. We generated a zebrafish model of TS with a homologous amino acid substitution in the cacna1c-encoded protein. Unlike patients, heterozygous mutants showed only mild impairments with no changes in mortality. However, homozygous mutants showed increased mortality, arrhythmias, neural activity and sensitivity to pentylenetetrazole-induced seizure-like behavior. Mutants also exhibited microcephaly, cerebellar hypotrophy and abnormal development of GABAergic neuron populations, and transcriptomic analysis revealed dysregulated expression of neuropeptide genes (including bdnf and vgf). Consistent with the idea that CaV1.2 channels activate during fever, we found that heterozygous larvae manifested arrhythmia and seizure-like behavior when exposed to elevated environmental temperature, and homozygous larvae switched from bradycardia to tachycardia. These data provide a basis for using zebrafish to study the etiology of TS abnormalities and suggest t hat fever may be a particular risk for TS patients.

genetics↗

bioRxiv2021

Detection of precisely edited CRISPR/Cas9 alleles through cointroduced restriction-fragment length polymorphisms

CRISPR/Cas9 is a powerful tool for producing genomic insertions and deletions (indels) to interrogate gene function. Modified CRISPR/Cas9 protocols can produce targeted genetic changes that are more precise than indels, but founder recovery is less efficient. Focusing on producing missense mutations in zebrafish using single-stranded oligo deoxynucleotide (ssODN) donor templates, we pioneered a strategy of adding synonymous changes to create novel restriction-enzyme (RE) sites, allowing detection of rare precise edits in a modified fluorescent-PCR fragment assay. We have named this process TIARS (test for incorporation of added recognition sites). Aided by TIARS, we induced two distinct amino-acid substitutions (T979I and P1387S) in the atp7a gene among somatic tissues of CRISPR-Cas9-treated F0 zebrafish. One of these F0s transmitted the allele to atp7aT979I/+ F1 progeny, and trans-heterozygosity of this allele against a null atp7a allele causes hypopigmentation, consistent with more severe pigment deficits in zebrafish or humans carrying only null mutations in atp7a/ATP7A. Design of ssODNs with novel RE recognition sites is labor-intensive, so we developed an in silico tool, TIARS Designer, and performed bioinformatic validation indicating that TIARS should be generalizable to other genes and experimental systems that employ donor template DNA.

genetics↗