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. 2020 Sep 8;11:4483. doi: 10.1038/s41467-020-18135-y

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a STAT92E^Act (green) and Tep4⁺ (magenta) or Antp⁺ (white) cells are mutually exclusive (Tep4-gal4 UAS-mCherry; STAT92E::edGFP). Magnified images: STAT92E^Act and Tep4⁺ cells near Antp⁺ PSC. b STAT92E^Act (green) do not co-localize with cells expressing high (PSC) or low (PHs) levels of collier (magenta). Magnified images: STAT92E^Act and col⁺ cells near the PSC. c. The number of STAT92E^Act cells (green) increases during lymph gland development (72 (n = 24), 96 (n = 17), and 120 h (n = 20) AEL). Antp⁺ (magenta) and STAT92E^Act are separable at all timepoints. Graphs represent quantitation of the number (left) or the proportion (right) of STAT92E^Act cells in one lymph gland lobe. d. Genetic ablation of the PSC (pCol85-gal4; STAT92E::edGFP UAS-Hid, Rpr) attenuates STAT92E^Act (green) expression (top). Graph indicates quantitations of the number of STAT92E^Act cells in one lymph gland lobe (bottom, Mann–Whitney test, ***P < 0.0001). pCol85-gal4/+ (n = 13) or pCol85-gal4 UAS-Hid, Rpr (n = 19). e Dl⁺ cells (magenta) are localized adjacent to the PSC (Antp, green). Magnified images: Dl⁺ and Antp⁺ cells. Cyan dotted lines demarcate Dl⁺ cells. f Dl⁺ cells (magenta) co-localize with STAT92E^Act (green). Magnified view: Dl⁺ and STAT92E^Act cells (right). A few Dl⁺ cells that do not express STAT92E::edGFP are indicated (arrowhead). g Lineage tracing of Dl⁺ cells (green, traced; magenta, real-time; blue, Antp). Delta^BL45136-gal4 UAS-GTRACE covers the entire lymph gland. Arrowheads represent Dl⁺ cells next to the Antp⁺ Dl⁺ PSC. h Model of the lymph gland hematopoiesis. PH1 cells are adjacent to the PSC. PH1 and PSC or PH2 are mutually exclusive. There are multiple states of prohemocytes including GST-rich and intermediary PHs/PMs. Plasmatocytes represent a heterogeneous cell population including adipohemocytes and late plasmatocytes, PM3-4. Colors indicate subclusters. Putative subclusters in the medullary zone (MZ), the cortical zone (CZ), or the intermediate zone (IZ) are described (right). Subcluster-specific markers verified in this study are listed (right). Red asterisks indicate 120 h AEL-specific subclusters. White scale bar, 30 μm; yellow scale bar, 3 μm. White dotted line demarcates the lymph gland. The dotted box (a–b, e–g) indicates a magnified region. Each panel at the right displays magnified views. Median value is represented in graphs (c, d).