henn-1/HEN1 promotes germline immortality in Caenorhabditis elegans

Small RNA and mRNA high-throughput sequencing was done to explore the roles of henn-1 and small RNA methylation in regulating gene expression and small RNA stability in C. elegans. Our results point to a requirement for henn-1 in methylating piRNAs, primary siRNAs, and a subset of miRNAs. In the absence of henn-1, modest changes in gene expression were observed in animals grown under optimal condition. To explore the role of henn-1 on the various classes of small RNAs, RNA was isolated from various mutant animals and from Argonaute co-immunoprecipitates and subjected to small RNA and/or mRNA sequencing. Differential expression in mutants compared to wild type animals was done using a variety of methods, primary DESeq2 when doing genome-wide comparisons. Enrichment in Argonaute co-immunoprecipitates relative to cell lysates was used to assess Argonaute-small RNA interactions. RNA polyphosphatase treatment was used to assess 5' phosphorylation of small RNAs and to enable capture of 22G-RNAs. Sodium periodate treatment was used to asses 3'-2'-O-methylation of small RNAs.