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. 2020 Jun 23;11:3172. doi: 10.1038/s41467-020-16911-4

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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. 3 — a Immunofluorescence staining of HSPCs with F-actin (red) and α-tubulin. White arrows: F-actin protrusions. Yellow arrows: polarization axis to determine the staining intensity in b. The experiments were repeated three times with similar results. b F-actin and α-tubulin staining intensity across the yellow arrows from a. c Quantitative analyses of F-actin staining intensity from a. N = 17 cells in mDia2^fl/fl group. N = 13 cells in mDia2^fl/flVav-Cre group. d Quantitative analyses of the percentage of cells with F-actin protrusions in a. N = 5 fields in mDia2^fl/fl group. N = 6 fields in mDia2^fl/fl Vav-Cre group. e Quantitative analysis of the percentages of MAL nuclear localization in c-Kit+ HSPCs incubated with or without FBS for 15 min. mDia2^fl/fl no FBS: N = 71 cells from 17 random fields; mDia2^fl/flVav-Cre no FBS: N = 80 cells from 15 random fields; mDia2^fl/fl with 10% FBS: N = 56 cells from 12 random fields; mDia2^fl/flVav-Cre with 10% FBS: N = 108 cells from 19 random fields. Data are from three independent experiments. f Quantitative RT-PCR of the indicated SRF target genes in c-Kit+ HSPCs from the indicated mice. g Quantitative RT-PCR of the indicated integrins in c-Kit+ HSPCs from the indicated mice. The experiments were performed in triplicate from n = 5 mice per group in f, g. h Quantitative analyses of the surface expression level of beta2 integrins using flow cytometry in the indicated HSPC populations. n = 2 mice per group. i Representative flow cytometry plots showing the expression of beta2 integrins in ex vivo-cultured c-Kit+ HSPCs. j Quantitative analyses of i. n = 2 mice per group. k In vitro adhesion assay of the cultured c-kit+ HSPCs from the indicated mice on extracellular matrix protein coated coverslips. Data are presented as adhesion ability relative to the control cells. mDia2^fl/fl, Collagen: 25 fields, Fibronectin: 20 fields, poly-d-lysine: 15 fields. mDia2^fl/fl Vav-Cre, Collagen: 10 fields, Fibronectin: 14 fields, poly-d-lysine: 9 fields. l In vitro adhesion assay of indicated c-kit+ HSPCs cultured on ICAM-1-coated coverslips. Error bars represent the SEM of the mean. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Two-tailed unpaired Student’s t test was used to generate the p values.