EMA2 is required for target-directed scnRNA degradation.

Piwi-associated small RNAs of the ciliated protozoan Tetrahymena uses such mechanism to induce heterochromatin formation followed by programmed DNA elimination. In this process, target-directed small RNA degradation (TDSD) of small RNAs complementary to the somatic genome enables small RNAs to specifically target the germline-limited sequences for DNA elimination. The data here indicate that the SUMO E3 ligase Ema2 is required for the accumulation of long non-coding RNAs (lncRNAs) from the somatic genome and thus for TDSD. Overall design: High-throughput sequencing and analyses of small RNAs were performed as described (Mutazono et al 2019). The 2020 version of the MAC genome assembly (Sheng et al. 2020) was obtained from Tetrahymena Genome Database and fragmented into 10 kb pieces. Each 10 kb fragment that contains longer than 3 kb mappable sequence was used as a MDS tile (total 10,235 tiles). Each Type-A IES (Noto et al. 2015) was used as an IES tile (total of 4,691 tiles). Normalized numbers (RPKM [reads per kilobase of unique sequences per million]) of 26- to 32-nt small RNAs that uniquely matched one of the MDS and IES tiles were counted.

纤毛原生动物四膜虫（Tetrahymena）中的Piwi结合小RNA（Piwi-associated small RNAs）借助此类机制诱导异染色质形成，继而引发程序性DNA消除。在该过程中，与体细胞基因组互补的小RNA会发生靶导向小RNA降解（target-directed small RNA degradation, TDSD），使得小RNA能够特异性靶向种系限制性序列以完成DNA消除。本数据表明，SUMO E3连接酶（SUMO E3 ligase）Ema2对于体细胞基因组来源的长链非编码RNA（long non-coding RNAs, lncRNAs）的积累是必需的，进而对TDSD过程不可或缺。

总体实验设计：参照已发表方法（Mutazono等，2019）开展小RNA的高通量测序与分析。从四膜虫基因组数据库获取2020版大核（MAC）基因组组装序列（Sheng等，2020），并将其打断为10 kb的片段。将其中包含大于3 kb可比对序列的10 kb片段作为MDS瓦片（总计10235个瓦片）；将每一类A型内部删除序列（IES，Noto等，2015）作为IES瓦片（总计4691个瓦片）。统计唯一匹配任一MDS瓦片或IES瓦片的26~32 nt小RNA的标准化计数（RPKM，即每百万reads中对应每千碱基唯一序列的reads数）。