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. 2019 Dec 9;10(12):940. doi: 10.1038/s41419-019-2134-8

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© The Author(s) 2019

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. 6 — a Representative immunoblottings (n = 5) of tyrosine-phosphorylation (pY) and the mitochondrial protein SDHa among subcellular fractions derived from MDA-MB-231 cells expressing empty vector (pcDNA) or MLS-Src-HA. Note the increased tyrosine-phosphorylation among mitochondrial proteins upon overexpression of mtSrc. b Representative immunoblottings of mitochondrial single-stranded DNA-binding protein (mtSSB) and tubulin in total cell lysates (TCL) derived from MDA-MB-231 cells expressing empty vector (pcDNA) or MLS-Src-HA (n = 3), indicating that overexpression of mtSrc does not change levels of mtSSB. c, d Representative immunoblottings (n = 3) of mtSSB, SDHa and tubulin among subcellular fractions of MDA-MB-231 cells expressing pcDNA or MLS-Src-HA (c), and representative micrographs (n = 3) of MDA-MB-231 cells co-expressing a mitochondria-targeted DsRed and pcDNA or MLS-Src-HA (D), indicating that subcellular localization of mtSSB does not change upon overexpression of mtSrc. TCL: total cell lysate; Cyto: cytosolic fraction; Mito: mitochondrial fraction. Scale bar: 25 μm. e Relative levels of mtDNA copies in MDA-MB-231 cells expressing empty vector, Src-HA or MLS-Src-HA (n = 4–5), indicating that overexpression of mtSrc reduces the amount of mtDNA. f Relative levels of mtDNA copies in MDA-MB-231 cells expressing pcDNA, MLS-Src-HA or co-expressing MLS-Src-HA with the phospho-deficient mtSSB-Y73F (n = 4–5), indicating that overexpression of mtSrc reduces the amount of mtDNA via phosphorylation of mtSSB-Y73. g Quantification of flow cytometry analyses of MDA-MB-231 cells expressing empty vector, MLS-Src-HA, or co-expressing mtSrc and mtSSB-Y73F stained with PI and Annexin V-FITC (n = 6), showing that mtSrc impacts on viability via phosphorylation of mtSSB-Y73 (see also Supplementary Fig. 6e). Data are presented as mean ± s.e.m. For E and F, *p ≤ 0.05 as compared to pcDNA, according to one-way ANOVA followed by Tukey’s multiple comparison test. Data presented in G were analyzed by two-way ANOVA followed by Tukey’s multiple comparison test (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001).