Affiliations: 1) University of Pittsburgh; 2) New York University School of Medicine
Keywords: g. unfolded protein response; b. oogenesis
Metazoans have evolved various stress response mechanisms to cope with cellular stress inflicted by external and physiological conditions. The Integrated Stress Response (ISR) is an evolutionarily conserved pathway that mediates adaptation to cellular stress via the transcription factor, ATF4. Here we describe a previously unknown role for Drosophila ATF4, encoded by the gene cryptocephal (crc), in oocyte maturation. We found that crc mutants have decreased egg laying and increased frequency of dorsal appendage defects in their eggs, with no apparent effect on fertilization rates. We observed substantial arrest in mid-oogenesis in crc mutant ovaries accompanied by premature apoptosis in early-stage egg chambers. Careful examination of ISR reporters revealed no detectable expression of ATF4 in the ovary, so we propose a non-autonomous role for ATF4 in the regulation of oogenesis.
Using a GFP insertion in the crc locus, we observed high levels of ATF4 expression in fat tissues surrounding the ovary. RNAi-mediated depletion of crc in fat tissues impaired oocyte maturation and decreased fertility similar to crc mutants. High levels of ATF4 expression in fat tissues suggest that this tissue has elevated basal levels of cellular stress. ATF4 activation in Drosophila can be mediated by one of two upstream ISR kinases: PERK, which responds to endoplasmic reticulum stress, or GCN2, which responds to amino acid deprivation. We are currently examining which of these kinases drives ATF4 expression in adult fat tissues to regulate oogenesis. Recent work has demonstrated that ATF4 acts as a coactivator for the Ecdysone receptor isoform EcR-B2 to regulate molting behavior. Since ecdysone signaling is a prominent regulator of oogenesis, we are currently parsing the role of ATF4-EcR-B2 signaling in fat tissues and its non-autonomous effects on oocyte maturation.
The crc 5’ leader has multiple cis-regulatory elements called upstream open reading frames (uORFs) that typically influence translation of the main ATF4-encoding ORF. We previously demonstrated that crc translation is regulated by the translation reinitiation factors eIF2D and the DENR-MCTS1 heterodimer. Consistently, loss of these factors yields similar fertility defects, implicating translation reinitiation in fat tissues as a regulator of oogenesis. Together, our data indicate that stress signaling in fat tissues non-autonomously regulates oocyte maturation.