The animal germline lineage needs to be maintained along generations. However, some Caenorhabditis elegans wild isolates display a mortal germline phenotype, leading to sterility after several generations at 25°C. Using a genome-wide association approach, we detect a significant peak on chromosome III around 5 Mb, confirmed by introgressions. Thus, a seemingly deleterious genotype is maintained at intermediate frequency in the species. Environmental rescue is a likely explanation, and indeed associated bacteria and microsporidia suppress the phenotype of wild isolates as well as mutants in small RNA inheritance (nrde-2) and histone modifications (set-2). Escherichia coli strains of the K-12 lineage suppress the phenotype compared to B strains. By shifting a wild strain from E. coli K-12 to E. coli B, we find that memory of the suppressing condition is maintained over several generations. Thus, the mortal germline phenotype of wild C. elegans is in part revealed by laboratory conditions and may represent variation in epigenetic inheritance and environmental interactions. This study also points to the importance of non-genetic memory in the face of environmental variation.

Caenorhabditis elegans strains with the heat-sensitive mortal germline phenotype become progressively sterile over the course of a few tens of generations when maintained at temperatures near the upper range of C. elegans\' tolerance. Mortal germline is transgenerationally heritable, and proximately under epigenetic control. Previous studies have suggested that mortal germline presents a relatively large mutational target and that mortal germline is not uncommon in natural populations of C. elegans. The mortal germline phenotype is not monolithic. Some strains exhibit a strong mortal germline phenotype, in which individuals invariably become sterile over a few generations, whereas other strains show a weaker (less penetrant) phenotype in which the onset of sterility is slower and more stochastic. We present results in which we (1) quantify the rate of mutation to the mortal germline phenotype and (2) quantify the frequency of mortal germline in a collection of 95 wild isolates. Over the course of ∼16,000 meioses, we detected one mutation to a strong mortal germline phenotype, resulting in a point estimate of the mutation rate UMrt≈ 6×10-5/genome/generation. We detected no mutations to a weak mortal germline phenotype. Six out of 95 wild isolates have a strong mortal germline phenotype, and although quantification of the weak mortal germline phenotype is inexact, the weak mortal germline phenotype is not rare in nature. We estimate a strength of selection against mutations conferring the strong mortal germline phenotype s¯≈0.1%, similar to selection against mutations affecting competitive fitness. The appreciable frequency of weak mortal germline variants in nature combined with the low mutation rate suggests that mortal germline may be maintained by balancing selection.

Chromatin modifications, including methylation of histone H3 at lysine 27 (H3K27me) by the Polycomb group proteins, play a broadly conserved role in the maintenance of cell fate. Diverse chromatin organization modifier (chromo) domain proteins act as \"readers\" of histone methylation states. However, understanding the functional relationships among chromo domains and their roles in the inheritance of gene expression patterns remains challenging. Here, we identify two chromo-domain proteins, CEC-1 and CEC-6, as potential readers of H3K27me in Caenorhabditis elegans, where they have divergent expression patterns and contribute to distinct phenotypes. Both cec-1 and cec-6 genetically interact with another chromo-domain gene, cec-3, a reader of H3K9 methylation. Combined loss of cec-1 and cec-3 leads to developmental defects in the adult that result in decreased fitness. Furthermore, loss of cec-6 and cec-3 surprisingly leads to a progressive loss of fertility across generations, a \"mortal germline\" phenotype. Our results provide evidence of functional compensation between H3K27me and H3K9me heterochromatin pathways, and show that histone methylation readers contribute to both somatic development and transgenerational fitness.

Although heredity mostly relies on the transmission of DNA sequence, additional molecular and cellular features are heritable across several generations. In the nematode Caenorhabditis elegans, insights into such unconventional inheritance result from two lines of work. First, the mortal germline (Mrt) phenotype was defined as a multigenerational phenotype whereby a selfing lineage becomes sterile after several generations, implying multigenerational memory [1, 2]. Second, certain RNAi effects are heritable over several generations in the absence of the initial trigger [3-5]. Both lines of work converged when the subset of Mrt mutants that are heat sensitive were found to closely correspond to mutants defective in the RNAi-inheritance machinery, including histone modifiers [6-9]. Here, we report the surprising finding that several C. elegans wild isolates display a heat-sensitive mortal germline phenotype in laboratory conditions: upon chronic exposure to higher temperatures, such as 25°C, lines reproducibly become sterile after several generations. This phenomenon is reversible, as it can be suppressed by temperature alternations at each generation, suggesting a non-genetic basis for the sterility. We tested whether natural variation in the temperature-induced Mrt phenotype was of genetic nature by building recombinant inbred lines between the isolates MY10 (Mrt) and JU1395 (non-Mrt). Using bulk segregant analysis, we detected two quantitative trait loci. After further recombinant mapping and genome editing, we identified the major causal locus as a polymorphism in the set-24 gene, encoding a SET- and SPK-domain protein. We conclude that C. elegans natural populations may harbor natural genetic variation in epigenetic inheritance phenomena.

In C. elegans, alterations to chromatin produce transgenerational effects, such as inherited increase in lifespan and gradual loss of fertility. Inheritance of histone modifications can be induced by double-stranded RNA-derived heritable small RNAs. Here, we show that the mortal germline phenotype, which is typical of met-2 mutants, defective in H3K9 methylation, depends on HRDE-1, an argonaute that carries small RNAs across generations, and is accompanied by accumulated transgenerational misexpression of heritable small RNAs. We discovered that MET-2 inhibits small RNA inheritance, and, as a consequence, induction of RNAi in met-2 mutants leads to permanent RNAi responses that do not terminate even after more than 30 generations. We found that potentiation of heritable RNAi in met-2 animals results from global hyperactivation of the small RNA inheritance machinery. Thus, changes in histone modifications can give rise to drastic transgenerational epigenetic effects, by controlling the overall potency of small RNA inheritance.