Macrophage mitochondrial bioenergetics and tissue invasion are boosted by an Atossa‐Porthos axis in Drosophila

• DOI: 10.15252/embj.2021109049
• Publication Date: 2022-03-23T10:59:00Z
• Authors: Shamsi Emtenani; Elliot T Martin; Attila Gyoergy; Julia Bicher; Jakob‐Wendelin Ge...; Thomas Köcher; Maria Akhmanova; Mariana Guarda; Marko Roblek; Andreas Bergthaler; Thomas R Hurd; Prashanth Rangan; Daria E Siekhaus
• Keywords: Bioenergetics

Article23 March 2022Open Access Source DataTransparent process Macrophage mitochondrial bioenergetics and tissue invasion are boosted by an Atossa-Porthos axis in Drosophila Shamsi Emtenani orcid.org/0000-0001-6981-6938 Institute of Science Technology Austria, Klosterneuburg, Austria Contribution: Conceptualization, Formal analysis, ​Investigation, Methodology, Writing - original draft Search for more papers this author Elliot T Martin Department Biological Sciences, RNA Institute, University at Albany, NY, USA draft, review & editing Attila Gyoergy orcid.org/0000-0002-1819-198X Julia Bicher ​Investigation Jakob-Wendelin Genger orcid.org/0000-0003-4502-1094 CeMM Research Center Molecular Medicine the Austrian Academy Vienna, Thomas Köcher Vienna BioCenter Core Facilities, Maria Akhmanova orcid.org/0000-0003-1522-3162 Visualization, Mariana Guarda Marko Roblek orcid.org/0000-0001-9588-1389 Methodology Andreas Bergthaler orcid.org/0000-0003-0597-1976 Resources, Data curation, Funding acquisition, R Hurd Genetics, Toronto, ON, Canada Prashanth Rangan orcid.org/0000-0002-1452-8119 Daria E Siekhaus Corresponding Author [email protected] orcid.org/0000-0001-8323-8353 Supervision, Project administration, Information Emtenani1, Martin2, Gyoergy1, Bicher1, Genger3, Köcher4, Akhmanova1, Guarda1, Roblek1, Bergthaler3, Hurd5, Rangan2 *,1 1Institute 2Department 3CeMM 4Vienna 5Department *Corresponding author. Tel: [phone]; E-mail: The EMBO Journal (2022)41:e109049https://doi.org/10.15252/embj.2021109049 See also: P Latorre-Muro Puigserver (June 2022) PDFDownload PDF article text main figures. Peer ReviewDownload a summary editorial decision including letters, reviewer comments responses to feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures Info Abstract Cellular metabolism must adapt changing demands enable homeostasis. During immune or cancer metastasis, cells leading migration into challenging environments require energy boost, but what controls capacity is unclear. Here, we study previously uncharacterized nuclear protein, Atossa (encoded CG9005), which supports macrophage germband controlling cellular metabolism. First, increases mRNA levels Porthos, DEAD-box two metabolic enzymes, lysine-α-ketoglutarate reductase (LKR/SDH) NADPH glyoxylate (GR/HPR), thus enhancing bioenergetics. Then Porthos ribosome assembly thereby raises translational efficiency subset mRNAs, those affecting functions, electron transport chain, Mitochondrial respiration measurements, metabolomics, live imaging indicate that power up OxPhos production promote forging path tissues macrophages. Since many crucial physiological output, undescribed genetic program may modulate wide range behaviors. Synopsis How satisfy increased during infiltration remains This combines imaging, genetics metabolomic analysis fly macrophages identifies new pathway supporting pioneer cell via reprogramming. uncharacterised protein CG9005) promotes embryos. expression enzymes (GR/HPR) lysine α-ketoglutarate (LKR/SDH), Krebs cycle ATP levels. targets (Pths), increasing assembled 40S translation OXPHOS components. Mammalian orthologs can substitute flies increase Video Emtenani, colleagues Introduction Charged with protecting organism against continuously threats, system constantly adapt, altering location, number, differentiation status its different subtypes (Nicholson, 2016). Such continuous adjustment requires high energy. these requirements just beginning be understood (O'Neill et al, 2016; Guak Krawczyk, 2020). currency ATP, produced from carbohydrates cytoplasmic glycolysis TCA feeds donors oxidative phosphorylation (OxPhos) complexes I through IV, components chain (ETC). Anaerobic quick, respiratory extracts considerably single molecule glucose, albeit slowly (Berg 2002). most directly regulated activity amount V carry it out (Hüttemann 2007). Upregulation known required important such as B antibody (Price 2018), pathogenic T-cell autoimmunity (Shin 2020), CD8+ memory development expansion (van der Windt 2012), T-reg suppressive function (Weinberg 2019), activation (Kiritsy 2021), maturation anti-inflammatory (Vats 2006). However, changes initiate upregulate unclear how shifts could influence unexplored. Immune move within distribution (Kierdorf 2015) respond homeostatic challenges, injuries, tumors, infections (Luster 2005; Ratheesh 2015). To migrate across unimpeded environments, expend restructuring their actin cytoskeleton, activating myosin ATPase reorganizing membrane (Cuvelier Even greater exist when also remodel surroundings they ahead resistance flanking extracellular matrix (Zanotelli 2018, 2019; Kelley 2019). Most vitro vivo studies on enables diverse types have highlighted importance (Semba 2018; Kishore 2018). our knowledge, only one has demonstrated need functional ETC, speed neutrophil potentially enabling polarized secretion amplify guidance cues (Zhou Increases triggered PGC-1’s transcriptional upregulation proteins underlie enhanced metastasis some suppress others (LeBleu 2014; Torrano Davis been shown particularly first coordinated chains (Khalil Friedl, 2010; Commander 2020); leader higher create (Zhang Although ability invade tumors depends movement surrounding resistance, not if similarly do achieve boost. identify mechanisms governing migration, macrophages, called plasmatocytes. Macrophages primary innate share remarkable similarities vertebrate ontogeny, migratory behavior (Ratheesh 2015; Wood Martin, 2017). These resolve infections, homeostasis (Bunt Buck Caputa Riera-Domingo reach places where needed proper development, follow extended between closely apposed ectoderm mesodermal tissues, moving (Siekhaus Valoskova Belyaeva 2022). Importantly, rate-limiting step macrophage, affected both properties 2018) well themselves (Valoskova Here induces concerted reprogramming support enzyme expression. Our data lay foundation mammalian pathological conditions, cancer, independent migration. Results CG9005 early find molecular pathways mediating invasion, examined BDGP situ project identified gene whose enriched prior entry (BDGP mRNA) (Tomancak 2002, maternally deposited expressed mesoderm, region specified Stage 4–6. further upregulated starting 7 while decreases remaining mesoderm. Stages 9–12 ingression, dissemination, toward germband. After downregulated match lower found ubiquitously embryo. We P-element insertion allele, CG9005BG02278 (CG9005PBG), visualizing fluorescent marker. Quantification revealed 36% decrease CG9005PBG mutant embryos compared control (Fig 1A, D), similar placed over either Df(2R)ED2222 Df(2R)BSC259 remove entirely 1D), demonstrating allele null invasion. Expressing completely restored 1C D). Driving any three interference (RNAi) lines decreased 37–40% 1E) sitting yolk near site yet invaded EV1A) 24–27%, shift seen EV1B). counted migrating along ventral nerve cord (vnc) late 12 embryos, route guided same factors lead (Wood 2017) requiring There was no significant difference EV1C) RNAi-expressing EV1D–F) controls, arguing basic processes recognition chemotactic signals unperturbed. Moreover, detected change total number genotypes EV1G H). Taken together, results fixed specifically steps Figure 1. acts spur A–C. Confocal images control, P{GT1}CG9005BG02278 Macrophage: red. Phalloidin visualize embryo: green. Germband edge: dotted white line. D. penetrated (A-C) deficiencies (Df) gene. n = 35, 56, 25, 9, 18 respectively; < 0.0001 versus CG9005PBG, Df1, Df2; 0.98 mac>CG9005 rescue; 0.91, 0.90 Df1 Df2. E. Macrophage-specific knockdown UAS-RNAi lines. 22, 20, 21, 23, 28 all comparisons. F. Stills two-photon movies showing (nuclei, red) 10 head invading tissue. Elapsed time indicated minutes. edge (white line) autofluorescence. For quantification parameters see (G-L). G, H. (G) (H) sac edge. (G): 8 movies, 3; 360 tracks, 450, 0.65. (H): 46 19, 0.62. I. nucleus enter Control 5. Time entry: 23 min, 38 0.0001. J–L. (J) 1st, (K) 2nd, (L) 3rd-5th 25–30 µm mesoderm ectoderm. In schematics, analyzed macrophages—light blue, other macrophages—red, ectoderm—green, mesoderm—purple, yolk—beige. (J): 6 5, 0.012. (K): 5 0.03. (L): 4, 0.17. information: Scale bars: 50 (A–C), 30 (F). Throughout paper mac> indicates GAL4 driven UAS construct srpHemo-GAL4. N represents work, stomodeal invagination retraction away anterior 29% were defined 10, 29–31% 11, 35–40% 12. (D) One-way ANOVA Tukey. (E G-L) Unpaired t-tests. Graphs show mean ± SEM; ns > 0.05, *P ****P 1 2 Fig available online figure. [embj2021109049-sup-0008-SDataFig1.zip] Download figure PowerPoint Click here expand EV1. normally vnc A, B. shows (A) expressing each RNAis (mac>) (B) element control. (n 43 embryos) CG9005PB 50), mutant/Df1 28) mutant/Df2 9), rescue 20) 0.99. alone versus, 0.001. 21 RNAi 0.0002; 25) 19) 0.0001; 3 16) 15) C–F. segments reveals (C) (D-F) controls. 0.05. 13) 0.25; (E) 0.5; (F) 50) 0.69. 17) 0.9; 27) 0.84; 23) 0.16. (J-L). J. Speed: 2.2 µm/min, 0.78; 373 124. K, L. directionality directionality: 0.39, 0.37, 0.74; 3. 0.40, 0.86; (A-L) labeled srpHemo-H2A::3xmCherry nuclei. red light green, purple, beige. work srpHemo-GAL4 driving constructs Mean SEM, ns=P **P 0.01, ***P 0.001, Tukey unpaired t-test (B-H) bar: (I). (CG9005) efficient assess CG9005’s role conducted nuclei (Figs 1F EV1I, Movies EV1 EV2). observed 9 neighboring point beyond 1G H, EV1J–L) (Speed yolk: µm/min CG9005PBG; 0.65, 0.78, respectively. Directionality: 0.39 0.37 regions, 0.74 head, 0.86 yolk. Speed 2.6, 2.5 0.62). 65% than (time 0.0001) 1I). pioneering significantly slower 1J K) (1st cell: 2.5, 0.012; 2nd 2.9, 2.1 0.03). next few 1L) (3rd–5th cells: 2.4 0.17). conclude regulates initial facilitating subsequent stream becomes much reduced atossa (atos), powerful Persian queen name means trickling. conserved motifs TADs (Atos) contains domain unknown (DUF4210) chromosome segregation (Chr_Seg) 2A). Atos displays trans-activating domains (TADs) common among transcription factors, localization (NLS), export signal (NES). FLAG::HA-tagged mainly embryonic 2B) macrophage-like S2R+ partially cytoplasm EV2A). forms lacking present EV2A) unable 2C D, EV2B C). Consistent defect, atosPBG atos mutants had wild-type EV2D). 40% identical murine orthologs, mFAM214A-B, maintain Expression mFAM214A rescued defect efficiently itself 2E F) normal EV2E). clearly critical primarily Atos, facilitate 2. CG9005/Atossa linked enhance maintained A. Deduced structure highlighting domains. FAM214A-B 44–45% Atos. (red) 11/12 tagged terminus HA (HA-antibody, green) under direct macrophage-specific promoter. Nucleus stained DAPI (blue). C, atosPBG, variants particular Transgene promoter (mac-). 32 56) mac-atos 18) 0.99; mac-atosDUF- 0.0003; mac-atosChrSeg-(n 21) mac-atosDUF-/ChrSeg-(n 0.00014; mac-atosTAD1-/ TAD2- 0.0009, 0.0031. Atossa’s (mFAMA-B) 24 0.7; mac-mFAMA 22) 0.6; mac-mFAMB 0.086. 0.0006; 0.0043. Mac srpHemo (C, E) (red), staining (green). (D,F) (B), E). [embj2021109049-sup-0009-SDa