Biology subjects

Duffie, R.

Publications and source records attributed to Duffie, R..

5 recordsLinked to original sources

bioRxiv2025

Stemness factor Mex3a times translation and protein trafficking to ensure robust differentiation of olfactory sensory neurons

During the switch from progenitor to differentiated cell, cellular physiology must change to accommodate increased translation and trafficking of membrane-bound proteins. We identify RNA-binding and E3 ubiquitin ligase Mex3a as a key driver of proper neuronal differentiation by regulating mRNA translation and trafficking of cell surface proteins in the context of Unfolded Protein Response (UPR) signaling. Loss of Mex3a in immature olfactory sensory neurons (OSNs) leads to defects in cilia structure, cell surface protein expression, and planar cell polarity in mature OSNs. Proteomics reveal a Mex3a-dependent decrease in proteins related to vesicle transport, lipid metabolism, and ribosome biogenesis. We identify RNA and ubiquitin targets of Mex3a and provide evidence that Mex3a may confer K27 ubiquitin linkage on substrates. Finally, modulating cellular levels of Mex3a changes the recruitment of translation factors Serbp1 and p-eEF2 to ribosomes with possible effects on translation. Our data reveal how a stemness factor regulates development post-transcriptionally and post-translationally to ensure robust differentiation. HighlightsLoss of stemness factor Mex3a in immature olfactory neurons leads to defects in mature olfactory neurons. Translation/Trafficking of cell surface proteins, cilia structure, and planar cell polarity are compromised in the absence of Mex3a. Mex3a may confer K27 ubiquitination on stress granule protein Serbp1 and ribosome protein Rps7. Mex3a levels are associated with Serbp1 and p-eEF2 recruitment to ribosomes.

genetics↗

bioRxiv2025

HP1B and H3K9me3 Regulate Olfactory Receptor Choice and Transcriptional Identity

Diverse epigenetic regulatory mechanisms ensure and regulate cellular diversity. Among others, the histone 3 lysine 9 me3 (H3K9me3) post translational modification participates in silencing lineage-inappropriate genes. H3K9me3 restricts access of transcription factors and other regulatory proteins to cell-fate controlled genes. In mice, olfactory sensory neurons (OSN) express one olfactory receptor (OR) gene out of 2,600 possibilities. This monoallelic and stochastic OR choice happens as OSNs differentiate and undergo dramatic changes in nuclear architecture. OR genes from different chromosomes converge into specialized nuclear bodies and chromatin compartments as H3K9me3 and chromatin binding proteins including heterochromatin protein 1 (HP1) are incorporated. In this work, we have uncovered an unexpected role for HP1{beta} in OR choice and neuronal identity that cannot be rescued by HP1 in vivo. With the use of a conditional knock-in mouse model that replaces HP1{beta} for HP1, we observe changes in H3K9me3 levels, DNA accessibility, and Hi-C contacts over OR gene clusters. These changes alter the expression patterns that partition the mouse olfactory epithelium into five OR expression zones, which results in a reduced OR repertoire leading to a loss of olfactory sensory neuron diversity. We propose that HP1{beta} modulates the competition of OR-promoters for enhancers to promote receptor diversity, by establishing repression gradients in a zonal fashion.

molecular biology↗

bioRxiv2020

Fruitless decommissions regulatory elements to implement cell-type-specific neuronal masculinization

In the fruit fly Drosophila melanogaster, male-specific splicing and translation of the Fruitless transcription factor (FruM) alters the presence, anatomy, and/or connectivity of >60 types of central brain neurons that interconnect to generate male-typical behaviors. While the indispensable function of FruM in sex-specific behavior has been understood for decades, the molecular mechanisms underlying its activity remain unknown. Here, we take a genome-wide, brain-wide approach to identifying regulatory elements whose activity depends on the presence of FruM. We identify 436 high-confidence genomic regions differentially accessible in male fruitless neurons, validate candidate regions as bona-fide, differentially regulated enhancers, and describe the particular cell types in which these enhancers are active. We find that individual enhancers are not activated universally but are dedicated to specific fru+ cell types. Aside from fru itself, genes are not dedicated to or common across the fru circuit; rather, FruM appears to masculinize each cell type differently, by tweaking expression of the same effector genes used in other circuits. Finally, we find FruM motifs enriched among regulatory elements that are open in the female but closed in the male. Together, these results suggest that FruM acts cell-type-specifically to decommission regulatory elements in male fruitless neurons.

neuroscience↗

bioRxiv2018

Antisense lncRNA transcription drives stochastic Protocadherin α promoter choice

Stochastic and combinatorial activation of clustered Protocadherin (Pcdh) , {beta}, and {gamma} gene promoters generates a cell-surface identity code in individual neurons that functions in neural circuit assembly. Here we show that Pcdh promoter choice requires transcription of a long noncoding RNA (lncRNA) initiated from newly identified promoters located in the protein coding sequence of each Pcdh exon. Antisense transcription of the lncRNA through the sense promoter results in its activation and in DNA demethylation of the binding sites for the CCCTC-binding protein, CTCF, located in close proximity to both sense and antisense promoters. Increased CTCF binding promotes the assembly of long-range DNA contacts between the activated promoter and a neuron-specific enhancer, thus locking in the epigenetic state of the stochastically chosen Pcdh promoter. Examination of this hierarchical molecular mechanism in differentiating olfactory sensory neurons, suggests that antisense Pcdh transcription is a key prerequisite for stochastic Pcdh promoter choice in vivo.

genomics↗

bioRxiv2018

Precise temporal regulation of alternative splicing during neural development

Alternative splicing (AS) is a crucial step of gene expression that must be tightly controlled, but the precise timing of dynamic splicing switches during neural development and the underlying regulatory mechanisms are poorly understood. Here we systematically analyzed the temporal regulation of AS in a large number of transcriptome profiles of developing mouse cortices, in vivo purified neuronal subtypes, and neurons differentiated in vitro. Our analysis revealed early- and late-switch exons in genes with distinct functions, and these switches accurately define neuronal maturation stages. Integrative modeling suggests that these switches are under direct and combinatorial regulation by distinct sets of neuronal RNA-binding proteins including Nova, Rbfox, Mbnl and Ptbp. Surprisingly, various neuronal subtypes in the sensory systems lack Nova and/or Rbfox expression. These neurons retain the \"immature\" splicing program in early-switch exons, affecting numerous synaptic genes. These results provide new insights into the organization and regulation of the neurodevelopmental transcriptome.

molecular biology↗