# Regulation of endogenous genes by sexually dimorphic piRNA expression during germline development in C. elegans

> **NIH NIH F31** · JOHNS HOPKINS UNIVERSITY · 2022 · $46,752

## Abstract

PROJECT ABSTRACT
Small RNAs – short, noncoding RNAs – are critical regulators in animal physiology and disease that silence
gene expression by complementary base pairing interactions to control mRNA stability/translation and epigenetic
modifications. Small RNA-mediated silencing pathways are evolutionarily conserved, and the largest class of
small RNAs comprise Piwi-interacting RNAs (piRNAs). Extensive studies in fly established that piRNAs silence
transposons and are expressed in a sex-specific manner; piRNAs are essential for genome integrity and
germline development. However, most piRNAs in mammals and worm do not map to transposons, and instead
are predicted to regulate germline-expressed genes. Although novel associations of individual piRNAs with
endogenous genes have been reported, endogenous gene targets of piRNAs remain largely unknown.
 In C. elegans, each of the ~15,000 piRNAs is autonomously transcribed and contains an upstream cis-
regulatory element, the Ruby motif. We found piRNAs are differentially expressed during spermatogenesis and
oogenesis, and male piRNAs have a strong bias for the 5’C nucleotide in the Ruby motif. Furthermore, we
identified SNPC-1.3 as a sex-specific transcription factor critical for male fertility. SNPC-1.3 depends on a known
core piRNA biogenesis factor, SNPC-4, to drive male piRNA expression during spermatogenesis. Loss of snpc-
1.3 during spermatogenesis results in global depletion of male piRNAs and sperm maturation defects. Other
piRNA biogenesis factors have been identified, but how these trans-acting factors interact with each other and
the Ruby motif is poorly understood.
 In this proposed research, I hypothesize that sex-specific regulatory mechanisms underlie piRNA
expression to regulate endogenous genes critical for proper germline development. To test my hypothesis in the
context of spermatogenesis, in Aim 1 I will use a recently developed strategy Cleavage Under Targets and
Release Using Nuclease (CUT&RUN) to characterize protein-DNA binding profiles of 5 piRNA biogenesis trans-
acting factors at the Ruby motif cis-regulatory element. Furthermore, I will determine if the 5’ C nucleotide in the
Ruby motif acts as a male specific element for SNPC-1.3. I will also use computational tools to identify putative,
novel cis-regulatory elements for male and female piRNAs. In Aim 2, I will computationally identify endogenous
gene targets of male piRNAs from small RNA-seq and mRNA-seq in wild-type and snpc-1.3(-) animals as well
as piRTarBase, a database of computationally predicted and experimentally identified piRNA targeting sites. I
will experimentally validate male piRNAs and their predicted endogenous gene targets using reporter assays for
endogenous and synthetic piRNAs and applying CRISPR/Cas9-mediated mutagenesis of piRNAs or their
targets. Collectively, this research will strengthen our understanding of how sex-specific piRNA expression is
transcriptionally regulated and provide new insights in...

## Key facts

- **NIH application ID:** 10389801
- **Project number:** 1F31HD107960-01
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Margaret Starostik
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $46,752
- **Award type:** 1
- **Project period:** 2022-06-01 → 2025-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10389801

## Citation

> US National Institutes of Health, RePORTER application 10389801, Regulation of endogenous genes by sexually dimorphic piRNA expression during germline development in C. elegans (1F31HD107960-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10389801. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*