Transcriptional states of retroelement-inserted regions and specific KRAB zinc finger protein association are correlated with DNA methylation of retroelements in human male germ cells
- RIKEN, Japan
- The University of Tokyo, Japan
- Tokyo Dental College Ichikawa General Hospital, Japan
- Nagoya University, Japan
Abstract
DNA methylation, repressive histone modifications, and PIWI-interacting RNAs are essential for controlling retroelement silencing in mammalian germ lines. Dysregulation of retroelement silencing is associated with male sterility. Although retroelement silencing mechanisms have been extensively studied in mouse germ cells, little progress has been made in humans. Here, we show that the Krüppel-associated box domain zinc finger proteins (KRAB-ZFPs) are associated with DNA methylation of retroelements in human primordial germ cells (hPGCs). Further, we show that the hominoid-specific retroelement SINE-VNTR-Alus (SVA) is subjected to transcription-directed de novo DNA methylation during human spermatogenesis. The degree of de novo DNA methylation in SVAs varies among human individuals, which confers significant inter-individual epigenetic variation in sperm. Collectively, our results highlight potential molecular mechanisms for the regulation of retroelements in human male germ cells.
Data availability
All reads from amplicon-seq in this study have been submitted to the Gene Expression Omnibus under accession number GSE174562.
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Amplicon-seq of SVA methylation in human spermNCBI Gene Expression Omnibus, GSE174562.
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ChIP-exo of human KRAB-ZNFs transduced in HEK 293T cells and KAP1 in hES H1 cellsNCBI Gene Expression OmnibusGSE78099.
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The Transcriptome and DNA Methylome Landscapes of Human Primordial Germ CellsNCBI Gene Expression OmnibusGSE63818.
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Genome-Wide Analysis of DNA Methylation Dynamics during Early Human Developmentthe Japanese Genotype-phenotype Archive, JGAS00000000006.
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Neonatal and adult human testis defined at the single-cell levelNCBI Gene Expression Omnibus,GSE124263.
Article and author information
Author details
Funding
Japan Society for the Promotion of Science (18H05530,18H03991)
- Yoichi Shinkai
RIKEN (SPDR)
- Kei Fukuda
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Human subjects: This study was approved by the ethics committees of RIKEN, Tokyo University, and Ichikawa General Hospital.All study participants were briefed about the aims of the study and the parameters to be measured, and consent was obtained.
Copyright
© 2022, Fukuda et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Transcriptional states of retroelement-inserted regions and specific KRAB zinc finger protein association are correlated with DNA methylation of retroelements in human male germ cells
eLife 11:e76822.
https://doi.org/10.7554/eLife.76822
Further reading
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Gene regulation is essential for life and controlled by regulatory DNA. Mutations can modify the activity of regulatory DNA, and also create new regulatory DNA, a process called regulatory emergence. Non-regulatory and regulatory DNA contain motifs to which transcription factors may bind. In prokaryotes, gene expression requires a stretch of DNA called a promoter, which contains two motifs called –10 and –35 boxes. However, these motifs may occur in both promoters and non-promoter DNA in multiple copies. They have been implicated in some studies to improve promoter activity, and in others to repress it. Here, we ask whether the presence of such motifs in different genetic sequences influences promoter evolution and emergence. To understand whether and how promoter motifs influence promoter emergence and evolution, we start from 50 ‘promoter islands’, DNA sequences enriched with –10 and –35 boxes. We mutagenize these starting ‘parent’ sequences, and measure gene expression driven by 240,000 of the resulting mutants. We find that the probability that mutations create an active promoter varies more than 200-fold, and is not correlated with the number of promoter motifs. For parent sequences without promoter activity, mutations created over 1500 new –10 and –35 boxes at unique positions in the library, but only ~0.3% of these resulted in de-novo promoter activity. Only ~13% of all –10 and –35 boxes contribute to de-novo promoter activity. For parent sequences with promoter activity, mutations created new –10 and –35 boxes in 11 specific positions that partially overlap with preexisting ones to modulate expression. We also find that –10 and –35 boxes do not repress promoter activity. Overall, our work demonstrates how promoter motifs influence promoter emergence and evolution. It has implications for predicting and understanding regulatory evolution, de novo genes, and phenotypic evolution.
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The male-specific lethal complex (MSL), which consists of five proteins and two non-coding roX RNAs, is involved in the transcriptional enhancement of X-linked genes to compensate for the sex chromosome monosomy in Drosophila XY males compared with XX females. The MSL1 and MSL2 proteins form the heterotetrameric core of the MSL complex and are critical for the specific recruitment of the complex to the high-affinity ‘entry’ sites (HAS) on the X chromosome. In this study, we demonstrated that the N-terminal region of MSL1 is critical for stability and functions of MSL1. Amino acid deletions and substitutions in the N-terminal region of MSL1 strongly affect both the interaction with roX2 RNA and the MSL complex binding to HAS on the X chromosome. In particular, substitution of the conserved N-terminal amino-acids 3–7 in MSL1 (MSL1GS) affects male viability similar to the inactivation of genes encoding roX RNAs. In addition, MSL1GS binds to promoters such as MSL1WT but does not co-bind with MSL2 and MSL3 to X chromosomal HAS. However, overexpression of MSL2 partially restores the dosage compensation. Thus, the interaction of MSL1 with roX RNA is critical for the efficient assembly of the MSL complex on HAS of the male X chromosome.
