Micropeptide ASAP encoded by LINC00467 promotes colorectal cancer progression by directly modulating ATP synthase activity
Qiwei Ge, … , Shujie Chen, Liangjing Wang
Qiwei Ge, … , Shujie Chen, Liangjing Wang
Published September 30, 2021
Citation Information: J Clin Invest. 2021;131(22):e152911. https://doi.org/10.1172/JCI152911.
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Research Article Gastroenterology Oncology

Micropeptide ASAP encoded by LINC00467 promotes colorectal cancer progression by directly modulating ATP synthase activity

Abstract

Emerging evidence has shown that open reading frames inside long noncoding RNAs (lncRNAs) could encode micropeptides. However, their roles in cellular energy metabolism and tumor progression remain largely unknown. Here, we identified a 94 amino acid–length micropeptide encoded by lncRNA LINC00467 in colorectal cancer. We also characterized its conservation across higher mammals, localization to mitochondria, and the concerted local functions. This peptide enhanced the ATP synthase construction by interacting with the subunits α and γ (ATP5A and ATP5C), increased ATP synthase activity and mitochondrial oxygen consumption rate, and thereby promoted colorectal cancer cell proliferation. Hence, this micropeptide was termed ATP synthase–associated peptide (ASAP). Furthermore, loss of ASAP suppressed patient-derived xenograft growth with attenuated ATP synthase activity and mitochondrial ATP production. Clinically, high expression of ASAP and LINC00467 predicted poor prognosis of colorectal cancer patients. Taken together, our findings revealed a colorectal cancer–associated micropeptide as a vital player in mitochondrial metabolism and provided a therapeutic target for colorectal cancer.

Authors

Qiwei Ge, Dingjiacheng Jia, Dong Cen, Yadong Qi, Chengyu Shi, Junhong Li, Lingjie Sang, Luo-jia Yang, Jiamin He, Aifu Lin, Shujie Chen, Liangjing Wang

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Figure 2

ASAP is localized to mitochondria and related to mitochondrial ATP production.

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ASAP is localized to mitochondria and related to mitochondrial ATP produ...
(A) GO analyses of genes coexpressed with LINC00467 in TCGA COAD database. (B) Immunofluorescence of ASAP (green) and MitoTracker (red) in HCT116 cells. Nuclei were stained with DAPI (blue). Scale bar: 10 μm. (C) Western blot detection of ASAP in the purified mitochondria from HCT116 cells with the indicated protein markers (SDHA and TOM20 for mitochondria and vinculin and α-tubulin for the cytosol). Cyto, cytosol; Mito, mitochondria. (D) Mitochondria were isolated from HCT116 cells and subjected to PK digestion at the indicated concentrations at 37°C. Mitochondria lysed in RIPA buffer served as a control for the total mitochondrial fraction. Western blots were performed for markers of OMM (TOM20), IMM (ATP5A), and ASAP. (E) Mitochondria were isolated from HCT116 cells and subjected to PK (103 ng/mL) proteolysis in the presence or absence of detergent (1% Triton). Western blots were performed for markers of OMM (TOM20), IMM (ATP5A), and ASAP. (F) Immunofluorescence staining was performed to mark ASAP (green) and IMM marker COX IV (red). The section was visualized using Leica TCS SP8 microscope with Lightning mode. Scale bar: 10 μm. (G) Relative mitochondrial ATP production was detected. Indicated HCT116 cells were treated with recording buffer (with 5 mM 2-DG and 5 mM pyruvate) to determine ATP generation under mitochondrial ATP synthesis. Data are presented as mean values ± SD from n = 6 biologically independent experiments. One-way ANOVA followed by Tukey’s test. ***P < 0.001. (H) OCR profile was monitored in indicated HCT116 cells with a Seahorse XF24 analyzer. The metabolic inhibitors were injected at different time points, as indicated. Data are presented as mean values ± SD. n = 3 biologically independent experiments. Data are representative of 3 independent experiments (B–F).