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. 2019 Apr 8:10:14.
doi: 10.1186/s13100-019-0156-5. eCollection 2019.

Subtype classification and functional annotation of L1Md retrotransposon promoters

Meng Zhou  1 , Andrew D Smith  1
Affiliations

Subtype classification and functional annotation of L1Md retrotransposon promoters

Meng Zhou et al. Mob DNA. .

Abstract

Background: L1Md retrotransposons are the most abundant and active transposable elements in the mouse genome. The promoters of many L1Md retrotransposons are composed of tandem repeats called monomers. The number of monomers varies between retrotransposon copies, thus making it difficult to annotate L1Md promoters. Duplication of monomers contributes to the maintenance of L1Md promoters during truncation-prone retrotranspositions, but the associated mechanism remains unclear. Since the current classification of monomers is based on limited data, a comprehensive monomer annotation is needed for supporting functional studies of L1Md promoters genome-wide.

Results: We developed a pipeline for de novo monomer detection and classification. Identified monomers are further classified into subtypes based on their sequence profiles. We applied this pipeline to genome assemblies of various rodent species. A major monomer subtype of the lab mouse was also found in other Mus species, implying that such subtype has emerged in the common ancestor of involved species. We also characterized the positioning pattern of monomer subtypes within individual promoters. Our analyses indicate that the subtype composition of an L1Md promoter can be used to infer its transcriptional activity during male germ cell development.

Conclusions: We identified subtypes for all monomer types using comprehensive data, greatly expanding the spectrum of monomer variants. The analysis of monomer subtype positioning provides evidence supporting both previously proposed models of L1Md promoter expansion. The transcription silencing of L1Md promoters differs between promoter types, which supports a model involving distinct suppressive pathways rather than a universal mechanism for retrotransposon repression in gametogenesis.

Keywords: Annotation; Classification; L1Md monomer; Retrotransposon.

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Conflict of interest statement

Not applicable.Not applicable.The authors declare that they have no competing interests.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
Application of monomer detection pipeline on various mouse genome assemblies reveals potential species-specific monomer subtypes. In this figure, genome names are abbreviated using the initials of their corresponding species name, except for mm10. They are: rn (Rattus norvegicus), mc (M. caroli), ms (M. spretus), mmc (M. m. castaneus), mmm (M. m. musculus), and mmd (M. m. domesticus). a Workflow of the monomer detection starting from a single consensus sequence. b Count of identified monomers in six mouse genomes. For each monomer type in all genomes, its corresponding consensus sequence reported in previous literature was used as the starting heuristic. c Length distribution of detected Type A monomers within the range of 180 – 220 bp. d Percentage of reads detected to share sequence similarity with L1 ORF2, Type A monomer and the A monomer subtype with a specific 7-bp insertion, respectively. Error bars indicate ±SD
Fig. 2
Fig. 2
Application of profile-HMM helps identify monomer subtypes and common truncation sites. a Visualization by tSNE of subtype identification based on the first 100 principal components using the HDBSCAN algorithm followed by k-NN clustering. b Heatmap showing sequence variation measured by edit distance for both intra- and inter-group subtypes of the Type A monomers. c – e Truncation profile indicated by the posterior expected number of states (top), and weighted read pileup of CAGE-seq data (bottom). Antisense read pileups were visualized as negative values
Fig. 3
Fig. 3
Analysis of monomer subtype composition in L1 promoters shows three position preference modes. a Distribution of monomer count per promoter of all three types. Promoters were generated by merging monomers within 20bp of each other. b Schematic diagram of ordering monomers based on their placement with respect to the orientation and ORF. c Three modes of monomer position preference. The distribution of monomer count for Type A promoters was used (left panel). Three A subtypes were chosen to represent distinct position preference modes, respectively. The three modes are: (1) Subtype 1 corresponding to the intermediate region mode, (2) Subtype 2 corresponding to the terminus region mode, and (3) Subtype 9 corresponding to the no preference mode
Fig. 4
Fig. 4
Differential response after piRNA pathway related KO experiments implies distinct L1Md promoter activity. a Distribution of methylation loss after KO experiments. b O/E ratios of A subtypes grouped by methylation loss after the Mili KO experiment. The groups were defined by the extent of methylation loss, including Positive (loss ≥0.5) and Negative (loss <0.5). The subtypes were arranged based on their position preference modes, which are intermediate region (top), terminus region (middle), and no preference (bottom), while Subtype 30 is listed as the outlier group
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