Pnrc2 regulates 3’UTR-mediated decay of segmentation clock-associated transcripts during zebrafish segmentation

Abstract

Vertebrate segmentation is controlled by the segmentation clock, a molecular oscillator that regulates gene expression and cycles rapidly. The expression of many genes oscillates during segmentation, including hairy/Enhancer of split-related (her or Hes) genes, which encode transcriptional repressors that auto-inhibit their own expression, and deltaC (dlc), which encodes a Notch ligand. We previously identified the tortuga (tor) locus in a zebrafish forward genetic screen for genes involved in cyclic transcript regulation and showed that cyclic transcripts accumulate post-splicing in tor mutants. Here we show that cyclic mRNA accumulation in tor mutants is due to loss of pnrc2, which encodes a proline-rich nuclear receptor co-activator implicated in mRNA decay. Using an inducible in vivo reporter system to analyze transcript stability, we find that the her1 3′UTR confers Pnrc2-dependent instability to a heterologous transcript. her1 mRNA decay is Dicer-independent and likely employs a Pnrc2-Upf1-containing mRNA decay complex. Surprisingly, despite accumulation of cyclic transcripts in pnrc2-deficient embryos, we find that cyclic protein is expressed normally. Overall, we show that Pnrc2 promotes 3′UTR-mediated decay of developmentally-regulated segmentation clock transcripts and we uncover an additional post-transcriptional regulatory layer that ensures oscillatory protein expression in the absence of cyclic mRNA decay.

Deadenylation by the CCR4‐NOT complex contributes to the turnover of hairy‐related mRNAs in the zebrafish segmentation clock

“…The functional importance of the 3 0 UTR in oscillatory gene expression in the segmentation clock has been described in several species [11,[16][17][18][19][20]. We currently do not know how the 3 0 UTR of these hairyrelated genes mediates distinct turnover rates in the zebrafish PSM.…”

“…The 3′UTR frequently possesses a variety of cis ‐acting regulatory elements that are specifically recognized by trans ‐acting factors such as RNA‐binding proteins and microRNAs . Recently, zebrafish pnrc2 , which is involved in nonsense mRNA decay, was shown to mediate 3′UTR‐dependent mRNA clearance of cyclic genes , suggesting that Pnrc2 may participate in the control of different turnover rates of her1 , her7 , and hes6 mRNAs with the CCR4‐NOT complex. It should also be considered that other mechanisms may be involved.…”

“…Thus, the turnover of these mRNAs should be properly regulated to erase the transcribed genetic information from the cells. Several previous studies demonstrated important roles of the 5′UTR and 3′UTR of cyclic genes in the mRNA half‐life in the mouse, chicken, Xenopus , and zebrafish . However, the molecular mechanisms underlying the rapid clearance of cyclic mRNAs still remain largely unknown in zebrafish.…”

The vertebrate Embryo Clock: Common players dancing to a different beat

“…Similar findings were reported for zebrafish EC genes ( Fujino et al, 2018 ), evidencing that 3′UTR-mediated regulation of EC gene expression oscillations is a conserved feature in vertebrates. The Amacher lab went on to specify that mRNA decay of both zebrafish her1 and deltaC relies on the Pumilio response- and AU-rich-elements present in their distal 3′UTRs, in a Pnrc2-dependent manner ( Gallagher et al, 2017 ; Tietz et al, 2020 ).…”

Keeping development on time: Insights into post‐transcriptional mechanisms driving oscillatory gene expression during vertebrate segmentation

“…Rather than typical, striped expression, tortuga mutant embryos exhibit uniform her1 and dlc mRNA expression throughout the PSM. This misexpression phenotype arises due to a defect in the clearance of segmentation clock transcripts that occurs when the function of proline-rich nuclear receptor coactivator 2 (pnrc2), a gene deleted in the tortuga deficiency, is lost (Gallagher et al, 2017). In human cultured cells, PNRC2 has been described as a mediator between the nonsense-mediated mRNA decay (NMD) and mRNA decapping complexes, specifically via its interactions with the NMD factor UPF1 and decapping complex protein DCP1A (Cho et al, 2009(Cho et al, , 2013(Cho et al, , 2015Lai et al, 2012;Mugridge et al, 2016).…”