Nonsense-mediated mRNA decay (NMD) is essential for removing premature termination codon-containing transcripts from cells. Studying the NMD pathway in model organisms can help to elucidate the NMD mechanism in humans and improve our understanding of how this biologically important process has evolved. Ciliates are among the earliest branching eukaryotes; their NMD mechanism is poorly understood and may be primordial. We demonstrate that highly conserved Upf proteins (Upf1a, Upf2 and Upf3) are involved in the NMD pathway of the ciliate, Tetrahymena thermophila. We further show that a novel protozoa-specific nuclease, Smg6L, is responsible for destroying many NMD-targeted transcripts. Transcriptome-wide identification and characterization of NMD-targeted transcripts in vegetative Tetrahymena cells showed that many have exon-exon junctions downstream of the termination codon. However, Tetrahymena may lack a functional exon junction complex (EJC), and the Tetrahymena ortholog of an EJC core component, Mago nashi (Mag1), is dispensable for NMD. Therefore, NMD is EJC independent in this early branching eukaryote.

Programmed DNA double-strand breaks (DSBs) are required for meiotic recombination, but the number is strictly controlled because they are potentially harmful. Here we report a novel protein, Pars11, which is required for Spo11-dependent DSB formation in the protist Tetrahymena. Pars11 localizes to chromatin early in meiotic prophase in a Spo11-independent manner and is removed before the end of prophase. Pars11 removal depends on DSB formation and ATR-dependent phosphorylation. In the absence of the DNA damage sensor kinase ATR, Pars11 is retained on chromatin and excess DSBs are generated. Similar levels of Pars11 persistence and DSB overproduction occur in a non-phosphorylatable pars11 mutant. We conclude that Pars11 supports DSB formation by Spo11 until enough DSBs are formed; thereafter, DSB production stops in response to ATR-dependent degradation of Pars11 or its removal from chromatin. A similar DSB control mechanism involving a Rec114-Tel1/ATM-dependent negative feedback loop regulates DSB formation in budding yeast. However, there is no detectable sequence homology between Pars11 and Rec114, and DSB numbers are more tightly controlled by Pars11 than by Rec114. The discovery of this mechanism for DSB regulation in the evolutionarily distant protist and fungal lineages suggests that it is conserved across eukaryotes.