Integrated stress response signaling acts as a metabolic sensor in fat tissues to regulate oocyte maturation and ovulation.

Reproduction is an energy-intensive process requiring systemic coordination. However, the inter-organ signaling mechanisms that relay nutrient status to modulate reproductive output are poorly understood. Here, we use Drosophila melanogaster as a model to establish the integrated stress response (ISR) transcription factor, Atf4, as a fat tissue metabolic sensor that instructs oogenesis. We demonstrate that Atf4 regulates lipase activity to mediate yolk lipoprotein synthesis in the fat body. Depletion of Atf4 in the fat body also blunts oogenesis recovery after amino acid deprivation and re-feeding, suggestive of a nutrient-sensing role for Atf4. We also discovered that Atf4 promotes secretion of a fat-body-derived neuropeptide, CNMamide, which modulates neural circuits that promote egg-laying behavior (ovulation). Thus, we posit that ISR signaling in fat tissue acts as a "metabolic sensor" that instructs female reproduction-directly by impacting yolk lipoprotein production and follicle maturation and systemically by regulating ovulation.

Integrated Stress Response signaling acts as a metabolic sensor in fat tissues to regulate oocyte maturation and ovulation.

Reproduction is an energy-intensive process requiring systemic coordination. However, the inter-organ signaling mechanisms that relay nutrient status to modulate reproductive output are poorly understood. Here, we use Drosophila melanogaster as a model to establish the Integrated Stress response (ISR) transcription factor, Atf4, as a fat tissue metabolic sensor which instructs oogenesis. We demonstrate that Atf4 regulates the lipase Brummer to mediate yolk lipoprotein synthesis in the fat body. Depletion of Atf4 in the fat body also blunts oogenesis recovery after amino acid deprivation and re-feeding, suggestive of a nutrient sensing role for Atf4. We also discovered that Atf4 promotes secretion of a fat body-derived neuropeptide, CNMamide, which modulates neural circuits that promote egg-laying behavior (ovulation). Thus, we posit that ISR signaling in fat tissue acts as a "metabolic sensor" that instructs female reproduction: directly, by impacting yolk lipoprotein production and follicle maturation, and systemically, by regulating ovulation.

On the illusion of auxotrophy: met15Δ yeast cells can grow on inorganic sulfur, thanks to the previously uncharacterized homocysteine synthase Yll058w.

Organisms must either synthesize or assimilate essential organic compounds to survive. The homocysteine synthase Met15 has been considered essential for inorganic sulfur assimilation in yeast since its discovery in the 1970s. As a result, MET15 has served as a genetic marker for hundreds of experiments that play a foundational role in eukaryote genetics and systems biology. Nevertheless, we demonstrate here through structural and evolutionary modeling, in vitro kinetic assays, and genetic complementation, that an alternative homocysteine synthase encoded by the previously uncharacterized gene YLL058W enables cells lacking Met15 to assimilate enough inorganic sulfur for survival and proliferation. These cells however fail to grow in patches or liquid cultures unless provided with exogenous methionine or other organosulfurs. We show that this growth failure, which has historically justified the status of MET15 as a classic auxotrophic marker, is largely explained by toxic accumulation of the gas hydrogen sulfide because of a metabolic bottleneck. When patched or cultured with a hydrogen sulfide chelator, and when propagated as colony grids, cells without Met15 assimilate inorganic sulfur and grow, and cells with Met15 achieve even higher yields. Thus, Met15 is not essential for inorganic sulfur assimilation in yeast. Instead, MET15 is the first example of a yeast gene whose loss conditionally prevents growth in a manner that depends on local gas exchange. Our results have broad implications for investigations of sulfur metabolism, including studies of stress response, methionine restriction, and aging. More generally, our findings illustrate how unappreciated experimental variables can obfuscate biological discovery.

A family of unusual immunoglobulin superfamily genes in an invertebrate histocompatibility complex.

Most colonial marine invertebrates are capable of allorecognition, the ability to distinguish between themselves and conspecifics. One long-standing question is whether invertebrate allorecognition genes are homologous to vertebrate histocompatibility genes. In the cnidarian Hydractinia symbiolongicarpus, allorecognition is controlled by at least two genes, Allorecognition 1 (Alr1) and Allorecognition 2 (Alr2), which encode highly polymorphic cell-surface proteins that serve as markers of self. Here, we show that Alr1 and Alr2 are part of a family of 41 Alr genes, all of which reside in a single genomic interval called the Allorecognition Complex (ARC). Using sensitive homology searches and highly accurate structural predictions, we demonstrate that the Alr proteins are members of the immunoglobulin superfamily (IgSF) with V-set and I-set Ig domains unlike any previously identified in animals. Specifically, their primary amino acid sequences lack many of the motifs considered diagnostic for V-set and I-set domains, yet they adopt secondary and tertiary structures nearly identical to canonical Ig domains. Thus, the V-set domain, which played a central role in the evolution of vertebrate adaptive immunity, was present in the last common ancestor of cnidarians and bilaterians. Unexpectedly, several Alr proteins also have immunoreceptor tyrosine-based activation motifs and immunoreceptor tyrosine-based inhibitory motifs in their cytoplasmic tails, suggesting they could participate in pathways homologous to those that regulate immunity in humans and flies. This work expands our definition of the IgSF with the addition of a family of unusual members, several of which play a role in invertebrate histocompatibility.

Genome-Wide Dynamic Evaluation of the UV-Induced DNA Damage Response.

Genetic screens in Saccharomyces cerevisiae have allowed for the identification of many genes as sensors or effectors of DNA damage, typically by comparing the fitness of genetic mutants in the presence or absence of DNA-damaging treatments. However, these static screens overlook the dynamic nature of DNA damage response pathways, missing time-dependent or transient effects. Here, we examine gene dependencies in the dynamic response to ultraviolet radiation-induced DNA damage by integrating ultra-high-density arrays of 6144 diploid gene deletion mutants with high-frequency time-lapse imaging. We identify 494 ultraviolet radiation response genes which, in addition to recovering molecular pathways and protein complexes previously annotated to DNA damage repair, include components of the CCR4-NOT complex, tRNA wobble modification, autophagy, and, most unexpectedly, 153 nuclear-encoded mitochondrial genes. Notably, mitochondria-deficient strains present time-dependent insensitivity to ultraviolet radiation, posing impaired mitochondrial function as a protective factor in the ultraviolet radiation response.

Ultrahigh-Density Screens for Genome-Wide Yeast EMAPs in a Single Plate.

Systematic measurements of genetic interactions have been used to classify gene functions and to categorize genes into protein complexes, functional pathways and biological processes. This protocol describes how to perform a high-throughput genetic interaction screen in S. cerevisiae using a variant of epistatic miniarray profiles (E-MAP) in which the fitnesses of 6144 colonies are measured simultaneously. We also describe the computational methods to analyze the resulting data.