Long Non-coding RNAs Facilitate Germline Stem-Cells Differentiation Through Phase Condensation

The transition from proliferative stem-cells into differentiating meiocytes has been extensively studied, yet our knowledge of the factors which control this critical process is limited. In the nematode C. elegans the cells are organized in a developmental-spatial manner, from the stem-cells to the mature oocyte, providing the perfect setup to answer this question. Two evolutionary-conserved PUF proteins reduce the stability of hundreds of transcripts to maintain stem-cells proliferation in the worm`s gonad. However, how differentiation initiates is still unknown since the PUFs are present at high-levels within differentiating meiocytes.

We identified three long-intergenic-non-coding RNAs (lincRNAs) which bind the PUFs. We engineered lincRNA deletion strains and found that individual deletions lead to additive reduction in fertility. Immunofluorescence showed that in gonads of the triple lincRNA mutant there are less mitotic and proliferative cells. RNA-Seq analyses showed that genes which are downregulated in the triple lincRNA deletion worms are highly enriched with genes which are bound by the PUF proteins. This suggests that the lincRNAs inhibit the PUFs action of reducing the stability of meiosis promoting transcripts. The lincRNAs are exclusively localized to the germ-granules, an evolutionary-conserved phase separated aggregates. In WT worms most of the PUF proteins are also localized in the germ granules, but in the triple strain there is a significant reduction in the association of the PUF proteins to these condensates. We propose that three lincRNAs promote oogenesis by sequestering the PUF proteins into phase separated granules thus increasing the expression of meiosis-promoting genes, and initiate oogenesis.