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Steroid hormones influence diverse biological processes throughout the animal life cycle, including metabolism, stress resistance, reproduction, and lifespan. In insects, the steroid hormone, 20-hydroxyecdysone (20E), is the central hormone regulator of molting and metamorphosis, and plays roles in tissue morphogenesis. For example, amnioserosa contraction, which is a major driving force in Drosophila dorsal closure (DC), is defective in embryos mutant for 20E biosynthesis. Here, we show that 20E signaling modulates the transcription of several DC participants in the amnioserosa and other dorsal tissues during late embryonic development, including zipper, which encodes for non-muscle myosin. Canonical ecdysone signaling typically involves the binding of Ecdysone receptor (EcR) and Ultraspiracle heterodimers to ecdysone-response elements (EcREs) within the promoters of responsive genes to drive expression. During DC, however, we provide evidence that 20E signaling instead acts in parallel to the JNK cascade via a direct interaction between EcR and the AP-1 transcription factor subunit, Jun, which together binds to genomic regions containing AP-1 binding sites but no EcREs to control gene expression. Our work demonstrates a novel mode of action for 20E signaling in Drosophila that likely functions beyond DC, and may provide further insights into mammalian steroid hormone receptor interactions with AP-1., Summary: During Drosophila dorsal closure, 20E signaling acts non-canonically through an interaction between EcR and the AP-1 subunit, Jun, to control gene expression at regions containing AP-1 motifs but no EcREs.

Homeodomain-interacting protein kinases (Hipks) have been previously associated with cell proliferation and cancer, however, their effects in the nervous system are less well understood. We have usedDrosophila melanogasterto evaluate the effects of altered Hipk expression on the nervous system and muscle. Using genetic manipulation of Hipk expression we demonstrate that knockdown and over-expression of Hipk produces early adult lethality, possibly due to the effects on the nervous system and muscle involvement. We find that optimal levels of Hipk are critical for the function of dopaminergic neurons and glial cells in the nervous system, as well as muscle. Furthermore, manipulation of Hipk affects the structure of the larval neuromuscular junction (NMJ) by promoting its growth. Hipk regulates the phosphorylation of the synapse-associated cytoskeletal protein Hu-li tai shao (Hts; adducin in mammals) and modulates the expression of two important protein kinases, Calcium-calmodulin protein kinase II (CaMKII) and Partitioning-defective 1 (PAR-1), all of which may alter neuromuscular structure/function and influence lethality. Hipk also modifies the levels of an important nuclear protein, TBPH, the fly orthologue of TAR DNA-binding protein 43 (TDP-43), which may have relevance for understanding motor neuron diseases.

Both functional and dysfunctional mitochondria are known to underlie tumor progression. Here, we establish use of the proto-oncogene Drosophila Homeodomain-interacting protein kinase (Hipk) as a new tool to address this paradox. We find that, in Hipk-overexpressing tumor-like cells, mitochondria accumulate and switch from fragmented to highly fused interconnected morphologies. Moreover, elevated Hipk promotes mitochondrial membrane hyperpolarization. These mitochondrial changes are at least in part driven by the upregulation of Myc. Furthermore, we show that the altered mitochondrial energetics, but not morphology, is required for Hipk-induced tumor-like growth, because knockdown of pdsw (also known as nd-pdsw; NDUFB10 in mammals; a Complex I subunit) abrogates the growth. Knockdown of ATPsynβ (a Complex V subunit), which produces higher levels of reactive oxygen species (ROS) than pdsw knockdown, instead synergizes with Hipk to potentiate JNK activation and the downstream induction of matrix metalloproteinases. Accordingly, ATPsynβ knockdown suppresses Hipk-induced tumor-like growth only when ROS scavengers are co-expressed. Together, our work presents an in vivo tumor model featuring the accumulation of hyperfused and hyperpolarized mitochondria, and reveals respiratory complex subunit-dependent opposing effects on tumorigenic outcomes. This article has an associated First Person interview with the first author of the paper., Summary: Altered mitochondrial energetics plays a more decisive role than morphology in determining tumor-like growth in a fly tumor model caused by elevated Homeodomain-interacting protein kinase.