Gozali, L. | bioRxiv Science

bioRxiv2022

Differential expression of Nanog 3UTR and coding region mRNA components defines an embryonic stem cell niche

Differential expression of mRNA coding sequences (CDSs) and 3untranslated regions (UTRs) is widespread, yet whether these domains contribute independently to cellular function remains unclear. Here, using Nanog as a model, we find that Nanog mRNA domain usage is spatially organized in mouse and human pluripotent cells and in the blastocyst, with cells enriched in 3UTR transcripts marking colony borders and cells enriched for the Nanog CDS located interior. Functional perturbation reveals a marked asymmetry between mRNA domains. Loss of the Nanog 3UTR leads to defects in colony architecture, cell spreading and morphogenetic behavior, accompanied by decreased extracellular matrix modeling gene expression and ROCK dependent cytoskeletal organization. In contrast, deletion of the Nanog CDS primarily disrupts epithelial polarity-associated transcriptional programs and the expression of chromatin regulators, consistent with a dominant role for the Nanog protein in transcriptional and epigenetic control. These domain specific effects are not redundant but instead reflect separable regulatory activities encoded within a single transcript. Together, these findings demonstrate that distinct regions of a single mRNA can encode separable and asymmetric biological functions, revealing mRNA domain usage as a distinct regulatory layer through which genes can encode multiple biological outputs beyond protein coding capacity.

developmental biology↗

bioRxiv2020

Distinct expression of select and transcriptome-wide isolated 3'UTRs suggests critical roles in development and transition states

Mature mRNA molecules are typically considered to be comprised of a 5UTR, a 3UTR and a coding region (CDS), all attached until degradation. Unexpectedly, however, there have been multiple recent reports of widespread differential expression of mRNA 3UTRs and their cognate coding regions, resulting in the expression of isolated 3UTRs (i3UTRs); these i3UTRs can be highly expressed, often in reciprocal patterns to their cognate CDS. Similar to the role of other lncRNAs, isolated 3UTRs are likely to play an important role in gene regulation but little is known about the contexts in which they are deployed. To begin to parse the functions of i3UTRs, here we carry out in vitro, in vivo and in silico analyses of differential 3UTR/CDS mRNA ratio usage across tissues, development and cell state changes both for a select list of developmentally important genes as well as through unbiased transcriptome-wide analyses. Across two developmental paradigms we find a distinct switch from high i3UTR expression of stem cell related genes in proliferating cells compared to newly differentiated cells. Our unbiased transcriptome analysis across multiple gene sets shows that regardless of tissue, genes with high 3UTR to CDS ratios belong predominantly to gene ontology categories related to cell-type specific functions while in contrast, the gene ontology categories of genes with low 3UTR to CDS ratios are similar and relate to common cellular functions. In addition to these specific findings our data provide critical information from which detailed hypotheses for individual i3UTRs can be tested-with a common theme that i3UTRs appear poised to regulate cell-specific gene expression and state. Significance StatementThe widespread existence and expression of mRNA 3 untranslated sequences in the absence of their cognate coding regions (called isolated 3UTRs or i3UTRs) opens up considerable avenues for gene regulation not previously envisioned. Each isolated 3UTR may still bind and interact with micro RNAs, RNA binding proteins as well as other nucleic acid sequences, all in the absence or low levels of cognate protein production. Here we document the expression, localization and regulation of i3UTRs both within particular biological systems as well as across the transcriptome. As this is an entirely new area of experimental investigation these early studies are seminal to this burgeoning field.

cell biology↗