Numbers correspond to mean fluorescence intensity (MFI mean ± SD, n = 3). ( F) PC3 cells as in E were analyzed for mitochondrial outer membrane permeability by calcein staining and flow cytometry. ( E) PC3 cells transfected with control nontargeting siRNA (siCtrl) or Mic60-directed siRNA (siMic60) were analyzed by transmission electron microscopy, and cristae length (mean ± SD) was quantified ( n = 25). IgG condition, number of detected peptides, and molecular weight (MW) are indicated. ( D) Mic60 interactome identified in PC3 cells by mass spectrometry. Top Left, arrow, perinuclear expression in normal acinar cells (×200) Top Right, perinuclear to diffuse cytoplasmic staining in nerves, smooth muscle, and ganglion cells (arrow, ×200) Middle Left, apical cytoplasmic staining of high-grade pancreatic intraepithelial neoplasia (×400) Middle Right, faint perinuclear as well as bright apical staining (arrow) of well-differentiated PDAC (×400) Bottom Left, asterisk, absent stain in high-grade basaloid PDAC, arrow (×200) Bottom Right, transition between Mic60-positive well differentiated tumor (double arrows) and Mic60-negative high grade basaloid regions within the same tumor gland (single arrow) (×400). ( C) Intratumoral heterogeneity of Mic60 expression in PDAC patients. ( B) Patient-derived tissue samples (#1 to #5) of GBM (G) or disease-free margin (M) were analyzed by Western blotting. Scale bars, low magnification, 6 mm high magnification, 60 μm (×40). Cases of COREAD ( n = 6N, 8T), GBM ( n = 5N, 8T), BRCA ( n = 5N, 8T), PRAD ( n = 5N, 5T), and LUAD ( n = 6N, 8T) were examined. ( A) A universal TMA was analyzed for differential expression of Mic60 in tumor vs. Mechanistically, differentiation of patient-derived GBM neurospheres, a process associated with the modulation of stemness and proliferative potential ( 22), lowered Mic60 as well as HIF1α mRNA levels ( SI Appendix, Fig. When analyzed in patient samples of pancreatic ductal adenocarcinoma (PDAC), Mic60 expression ranged from focal perinuclear distribution in normal pancreatic acinar cells to disordered, submembranous or linear staining in in situ and invasive neoplastic epithelium to absence in poorly differentiated (basaloid) carcinomas by IHC ( Fig. Heterogeneous Mic60 expression was also observed intratumorally. S1 C) in public databases, our TMA analysis did not reach statistical significance ( SI Appendix, Fig. Although breast cancer showed increased Mic60 protein ( SI Appendix, Fig. Other potential Mic60-low tumors in this analysis included malignancies of kidney, thyroid, head and neck, and soft tissue, whereas uterine and cervix cancer had higher Mic60 mRNA levels compared to normal tissues ( SI Appendix, Fig. Consistent with these results, Mic60 mRNA levels in The Cancer Genome Atlas (TCGA) database were reduced in GBM and COREAD but prominently upregulated in LUAD ( SI Appendix, Fig. 1 A), where Mic60 expression was reduced in colorectal adenocarcinoma (COREAD) and glioblastoma (GBM), unchanged in breast (BRCA) and prostate adenocarcinoma (PRAD), or increased in lung adenocarcinoma (LUAD) compared to adjacent normal tissue ( SI Appendix, Fig. Immunohistochemical (IHC) staining of a universal tumor microarray (TMA n = 5 to 8 cases per tumor type) gave similar results ( Fig. Inspection of the Human Protein Atlas database showed that Mic60 expression was highly heterogeneous in cancer, as several tumor types had reduced, increased, or unchanged levels of Mic60 compared to normal tissues ( SI Appendix, Fig. To study the role of mitochondrial fitness in cancer, we focused on Mic60 as an essential scaffold of organelle integrity and function ( 18). These data demonstrate that acutely damaged, “ghost” mitochondria drive tumor progression and expose an actionable therapeutic target in metastasis-prone cancers. In a small-molecule drug screen, compensatory activation of stress response (GCN2) and survival (Akt) signaling maintains the viability of Mic60-low tumors and provides a selective therapeutic vulnerability.
In turn, this induces epithelial-mesenchymal transition (EMT), activates tumor cell movements through exaggerated mitochondrial dynamics, and promotes metastatic dissemination in vivo. Despite a catastrophic collapse of mitochondrial integrity, loss of bioenergetics, and oxidative damage, tumors with Mic60 depletion slow down cell proliferation, evade cell death, and activate a nuclear gene expression program of innate immunity and cytokine/chemokine signaling. Here, we show that many human tumors have heterogeneous and often reduced levels of Mic60, or Mitofilin, an essential scaffold of mitochondrial structure. Cancer metabolism, including in mitochondria, is a disease hallmark and therapeutic target, but its regulation is poorly understood.
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