Nonsense-mediated mRNA decay

To defend against genetic nonsense mutations, our cells have evolved an mRNA surveillance pathway--nonsense-mediated mRNA decay (NMD)--to selectively degrade mRNAs containing premature stop codons. NMD is also part of the posttranscriptional quality control mechanisms to eliminate errors of transcription or mRNA processing and ensure fidelity of gene expression. However, to what extent NMD is involved to drive brain development and brain diseases is unclear.

We showed that NMD, more than just a passive quality control mechanism, is an essential regulatory mechanism to quantitatively determine the abundance of physiological transcripts (Zheng et al. 2012, Zheng 2016). For example, inhibition of Psd-95 exon 18 leads to a shift in the reading frame and a PTC in the downstream exon, resulting in NMD of the Psd-95 transcript without productive translation. Interestingly, non-neuronal cells express only the exon 18-skipping isoform due to high PTBP expression, whereas mature neurons express only the exon 18-plus isoform thanks to the loss of PTBPs. Therefore, alternative splicing coupled to NMD (AS-NMD) can act as an on/off switch of gene expression during development and enforce tissue specific expression (Zheng et al. 2013). Many RNA binding proteins use the similar mechanism for their homeostatic expression. Our study AS-NMD can effectively control non-RBP genes and developmental processes.

We harnessed this regulatory mechanism to devise a new method to measure cellular NMD efficiency. Briefly, NMD-sensitive isoforms and NMD-resistant counterparts of the same gene are assessed separately; comparing the responses of the two transcript isoforms distinguishes NMD regulation from transcriptional and alternative splicing regulations. Our method assays endogenous NMD targets rather than conventional exogenous plasmid reporters, therefore providing superior reproducibility, sensitivity, specificity, and ease of application. The method can be broadly applied to cells of low transfection efficiency and tissues. We are currently applying this method for large scale analysis of NMD efficiency across biological conditions.

We applied this method and screened for chemical modulators of NMD activity. After comprehensive characterization of one screening hit, we found that NMD is inhibited by endoplasmic reticulum (ER) stress through PERK kinase activation (Li et al. 2017). Interestingly, previous studies showed that NMD controls mediators of unfolded protein response (UPR) pathway to inhibit ER stress. These data together show that NMD is integrated into the cellular response to ER stress via a negative feedback loop. We further showed that ER stress compounded TDP-43 depletion in the upregulation of NMD isoforms that had been implicated in the pathogenic mechanisms of amyotrophic lateral sclerosis and frontotemporal demonetia. Our study also unexpectedly reveals that NMD substrates exhibit varied NMD-targeting efficiency, which we are now actively investigating.