On some level, biology is all about regulation, regulation, regulation. One way that our cells regulate their genes is through a process called alternative splicing, which entails the selective inclusion of discrete portions of a gene (called exons) into the final protein product—think of this like buying a new phone or car and deciding which gadgets or gizmos to add on to the base model. Just like the addition of gadgets to a car gives you the ability to make several slightly different cars from a single base model, alternative splicing vastly increases the diversity of proteins that can be produced from a more limited set of genes.
While the mechanisms involved in and consequences of alternative splicing remain active areas of research, there is one class of alternative splicing events that stands out as particularly puzzling. These splicing events are characterized by inclusion of a special type of exon called a poison exon—poison, because these exons encode a premature ‘stop’ codon which leads to the degradation of the RNA molecule. To return to our analogy, a poison exon would be akin to buying a 2024 Subaru Crosstrek with an automatic self-destruct feature. While their self-destruct function might seem counterintuitive at first, poison exons are thought to serve a regulatory role by allowing cells to precisely control the expression of certain genes via modulating their splicing. Too much protein X around? Just alter the splicing of gene X to include the poison exon, and protein X levels are now downregulated.
Poison exons aren’t exactly rare. There are hundreds, if not thousands of them, in most genomes. More importantly, they’re generally very well conserved among the genomes of animals and plants, implying that they serve a crucial function—crucial enough for Mother Nature to keep them around. Surprisingly, however, research into these enigmatic genetic elements is relatively scant. A recent study from Dr. Robert Bradley’s lab at Fred Hutch led by M.D. Ph.D. student Dr. Andrea Belleville addresses this knowledge gap and finds an important role for a poison exon that spans across kingdoms of life.
In their study, Belleville and Bradley focus on a single gene, SMNDC1, that encodes a component of cellular splicing machinery and was previously studied by the group for its role in controlling splicing dynamics in tumors. Taking a closer look at the poison exon of SMNDC1 across different species, they found that among similar splicing factors, SMNDC1 and its poison exon were particularly well-conserved—in fact, they even discovered evidence of a previously unknown poison exon in SPF30, the plant ortholog of SMNDC1!
Was this previously unannotated poison exon in SPF30 real? And if so, what was its function at the organismal level, as well as the function of SMNDC1’s poison exon? To answer these questions, Belleville and colleagues conducted a series of in vitro experiments which confirmed that indeed, SPF30 has a poison exon just like its human counterpart. Moreover, when the team overexpressed (mouse) SMNDC1 in mouse cells, they observed greater inclusion of its poison exon, suggesting that the poison exon functioned to regulate SMNDC1 abundance. Strikingly, they also found that this effect crossed species: artificially expressing plant SPF30 in mouse cells correlated with greater poison exon inclusion in the endogenous mouse SMNDC1, demonstrating functional conservation of the SMNCD1 poison exon that crosses kingdoms of life.
To really understand what role this poison exon plays in vivo, Belleville and colleagues used CRISPR-Cas9 technology to generate transgenic mice lacking this poison exon. First, they confirmed their in vitro findings with the observation that SMNDC1 protein levels were higher in animals lacking the SMNDC1 poison exon. Since SMNDC1 is itself a splicing factor, the team next examined splicing in their poison exon-null mice. Indeed, their analysis revealed global changes to gene splicing that appeared to impact specific pathways more than others, including those involved in central carbon metabolism. Despite these molecular changes, Belleville and colleagues were initially surprised that the poison exon-null mice appeared completely healthy—they grew as well as their wild-type littermates and showed no signs of organ damage or altered behavior, even though it was previously shown that complete deletion of SMNDC1 in mice is embryonically lethal.
After several more rounds of mouse breeding, however, they made a discovery: mice born to breeding pairs in which both parents were poison exon-null were significantly smaller than their wild-type counterparts. While Belleville and team were busy characterizing these mice, they were also collaborating with Dr. Christine Quietsch and her lab at UW Genome Sciences, who generated plants (Arabidopsis thaliana, for the officionados) that lacked the SPF30 poison exon. Surprisingly, these poison exon-null plants showed a very similar phenotype to poison exon-null mice: they also produced smaller progeny than the wild-type controls. Thus, it seems that despite being separated by millions of years of evolution, humans, mice, and plants all rely on SMNDC1 orthologs and their corresponding poison exons to maintain maximal organismal fitness.
“To our knowledge, this is one of the first times anyone has studied the consequences of specifically removing a poison exon in a living animal,” noted Dr. Belleville, “and our study really highlights a striking example of how functionally conserved a poison exon can be. Clearly, these genetic elements play crucial biological roles that should be better understood. While we still don’t know exactly why removing this poison exon leads to growth impediments in mice or plants, we hope that this study can serve as a starting point for further research on SMNDC1’s poison exon, as well as the hundreds of other poison exons out there.”
Dr. Bradley holds the McIlwain Family Endowed Chair in Data Science.
The spotlighted work was funded by the National Institutes of Health, Leukemia & Lymphoma Society, Mark Foundation for Cancer Research, and Paul G. Allen Frontiers Group.
Fred Hutch/University of Washington/Seattle Children’s Cancer Consortium member Dr. Robert Bradley contributed to this study.
Belleville, A. E., Thomas, J. D., Tonnies, J., Gabel, A. M., Borrero Rossi, A., Singh, P., Queitsch, C., & Bradley, R. K. (2024). An autoregulatory poison exon in Smndc1 is conserved across kingdoms and influences organism growth. PLOS Genetics, 20(8), e1011363.
Science Spotlight writer David Sokolov is a graduate student in the Sullivan Lab at the Fred Hutch. He studies how cancer cells modify their metabolism to facilitate rapid proliferation and accommodate tumor-driving mitochondrial defects. He's originally from the east coast and has bachelors' and masters' degrees from West Virginia University. Outside of the lab, you'll find him enjoying the outdoors, playing music, or raising composting worms in his front yard.
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