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Combinatorial tagging generates a multi-purpose knock-in mouse model revealing phase separation-dependent germ granules in RNA homeostasis and germline development

ABSTRACT: A large resource of epitope-tagged or Cre/CreERT2-expressing mouse models are available for study of germ granules or germline development. The germ granules are proteinaceous, membraneless organelles implicated in germ cell differentiation and maturation; however, their protein and RNA transcript constituents, and their mechanistic insight remain not fully understood. Herein, we generated a versatile germline mouse model through combinatorial tagging of DDX4 for simultaneous expression of three cistronic coding products (C-terminal tagged DDX4 - DDX45HA, EGFP, and CreERT2) under the driving control of endogenous Ddx4 promoter. By leveraging the high-affinity HA tag, we optimized an efficient workflow for purification of germ granules (Chromatoid body, CB) from spermatids, and characterized the landscape of protein constituents and RNA transcripts in CB. Moreover, we explored and ascertained that DDX4-mediated, phase-separation dependent CB integrity is functionally important for recruiting distinctive long RNA transcripts and biogenesis of pachytene- and TE-derived piRNAs. Together, our study generated a versatile germline mouse model with a multiplicity of applications for germline study, and provided mechanistic insight into germline development as dictated by germ granules.

ORGANISM(S): Mus musculus

PROVIDER: GSE273344 | GEO | 2024/08/05

REPOSITORIES: GEO

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Germline-specific RNA helicase DDX4 forms cytoplasmic granules in cancer cells and promotes tumor growth

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2024-06-10 | PXD044856 | Pride

Germline-specific RNA helicase DDX4 forms cytoplasmic granules in cancer cells and promotes tumor growth

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C. elegans germ granules regulate both germline and somatic RNAome [miRNA-Seq]

2023-07-20 | GSE228857 | GEO

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