Abstract
BackgroundThe 3′ untranslated regions (UTRs) of mRNAs play a major role in post-transcriptional regulation of gene expression. Selection of transcript cleavage and polyadenylation sites is a dynamic process that produces multiple transcript isoforms for the same gene within and across different cell types. Using LITE-Seq, a new quantitative method to capture transcript 3′ ends expressed in vivo, we have characterized sex- and cell type-specific transcriptome-wide changes in gene expression and 3′UTR diversity in Caenorhabditis elegans germline cells undergoing proliferation and differentiation.ResultsWe show that nearly half of germline transcripts are alternatively polyadenylated, that differential regulation of endogenous 3′UTR variants is common, and that alternative isoforms direct distinct spatiotemporal protein expression patterns in vivo. Dynamic expression profiling also reveals temporal regulation of X-linked gene expression, selective stabilization of transcripts, and strong evidence for a novel developmental program that promotes nucleolar dissolution in oocytes. We show that the RNA-binding protein NCL-1/Brat is a posttranscriptional regulator of numerous ribosome-related transcripts that acts through specific U-rich binding motifs to down-regulate mRNAs encoding ribosomal protein subunits, rRNA processing factors, and tRNA synthetases.ConclusionsThese results highlight the pervasive nature and functional potential of patterned gene and isoform expression during early animal development.
Highlights
The 3′ untranslated regions (UTRs) of messenger RNA (mRNA) play a major role in post-transcriptional regulation of gene expression
An mRNA’s fate depends on internal cis-regulatory sequences that are most commonly located in their 3′ untranslated regions (3′UTRs), and mutations within these have been associated with cancers and other diseases [9]
A Quantitative 3′-end Capture Sequencing Method for Low Input RNA Samples To monitor gene expression and 3′UTR dynamics using small amounts of RNA isolated from manually dissected germline tissue, we developed a new Low-Input 3′-Terminal sequencing method (LITE-Seq) that is both highly sensitive and quantitative (Fig. 1)
Summary
The 3′ untranslated regions (UTRs) of mRNAs play a major role in post-transcriptional regulation of gene expression. Germ cells are produced in assembly-line fashion in the gonad, progressing from mitotic proliferation in the distal region through meiosis and differentiation into oocytes and sperm in the proximal region. This developmental program depends on the spatio-temporal restriction of RBP activity, which is mediated largely by translational control of their own mRNAs through their 3′UTRs [14, 16, 17]. For example GLD-1, which is involved in the transition from mitosis to meiosis and oocyte differentiation [18,19,20], binds multiple targets [21] to both repress translation [22] and stabilize transcripts [17, 18, 23]. Translation of GLD-1 mRNA is negatively regulated by another RBP, FBF-1, which promotes mitotic proliferation and represses transcripts required for
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