Flura-seq identifies organ-specific metabolic adaptations during early metastatic colonization
Abstract
Metastasis-initiating cells dynamically adapt to the distinct microenvironments of different organs, but these early adaptations are poorly understood due to the limited sensitivity of in situ transcriptomics. We developed fluorouracil-labeled RNA sequencing (Flura-seq) for in situ analysis with high sensitivity. Flura-seq utilizes cytosine deaminase (CD) to convert fluorocytosine to fluorouracil, metabolically labeling nascent RNA in rare cell populations in situ for purification and sequencing. Flura-seq revealed hundreds of unique, dynamic organ-specific gene signatures depending on the microenvironment in mouse xenograft breast cancer micrometastases. Specifically, the mitochondrial electron transport Complex I, oxidative stress and counteracting antioxidant programs were induced in pulmonary micrometastases, compared to mammary tumors or brain micrometastases. We confirmed lung metastasis-specific increase in oxidative stress and upregulation of antioxidants in clinical samples, thus validating Flura-seq's utility in identifying clinically actionable microenvironmental adaptations in early metastasis. The sensitivity, robustness and economy of Flura-seq are broadly applicable beyond cancer research.
Data availability
Sequencing data have been deposited in GEO under accession codes GSE93605 and GSE118937.
-
Organ-specific in situ transcriptomics of MDA231 cells identified by Flura-seqNCBI Gene Expression Omnibus, GSE118937.
-
Flura-seq of TGFB treated MDA231 cellsNCBI Gene Expression Omnibus, GSE93605.
-
Integrated RNA and DNA sequencing reveals early drivers of metastatic breast cancerNCBI Gene Expression Omnibus, GSE110590.
-
ubpopulations of MDA-MB-231 and Primary Breast CancersNCBI Gene Expression Omnibus, GSE2603.
-
Breast cancer relapse free survival and lung metastasis free survivalNCBI Gene Expression Omnibus, GSE5327.
-
Breast cancer relapse free survivalNCBI Gene Expression Omnibus, GSE2034.
-
Expression data from primary breast tumorsNCBI Gene Expression Omnibus, GSE12276.
Article and author information
Author details
Funding
National Institutes of Health (P01-CA094060)
- Joan Massagué
Damon Runyon Cancer Research Foundation (DR-12998)
- Harihar Basnet
Department of Defense (W81XWH-12-0074)
- Joan Massagué
National Institutes of Health (T32-CA009207)
- Karuna Ganesh
National Institutes of Health (T32-GM07739)
- Yun-Han Huang
National Institutes of Health (K08-CA230213)
- Karuna Ganesh
National Institutes of Health (F30-CA203238)
- Yun-Han Huang
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Matthew G Vander Heiden, Massachusetts Institute of Technology, United States
Ethics
Animal experimentation: Mouse experiments were performed following the protocols approved by the MSKCC Institutional Animal Care and Use Committee (IACUC) (#99-09-032).
Version history
- Received: November 14, 2018
- Accepted: March 6, 2019
- Accepted Manuscript published: March 26, 2019 (version 1)
- Version of Record published: March 29, 2019 (version 2)
Copyright
© 2019, Basnet et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 5,511
- views
-
- 799
- downloads
-
- 44
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Cancer Biology
- Genetics and Genomics
Relapse of acute myeloid leukemia (AML) is highly aggressive and often treatment refractory. We analyzed previously published AML relapse cohorts and found that 40% of relapses occur without changes in driver mutations, suggesting that non-genetic mechanisms drive relapse in a large proportion of cases. We therefore characterized epigenetic patterns of AML relapse using 26 matched diagnosis-relapse samples with ATAC-seq. This analysis identified a relapse-specific chromatin accessibility signature for mutationally stable AML, suggesting that AML undergoes epigenetic evolution at relapse independent of mutational changes. Analysis of leukemia stem cell (LSC) chromatin changes at relapse indicated that this leukemic compartment underwent significantly less epigenetic evolution than non-LSCs, while epigenetic changes in non-LSCs reflected overall evolution of the bulk leukemia. Finally, we used single-cell ATAC-seq paired with mitochondrial sequencing (mtscATAC) to map clones from diagnosis into relapse along with their epigenetic features. We found that distinct mitochondrially-defined clones exhibit more similar chromatin accessibility at relapse relative to diagnosis, demonstrating convergent epigenetic evolution in relapsed AML. These results demonstrate that epigenetic evolution is a feature of relapsed AML and that convergent epigenetic evolution can occur following treatment with induction chemotherapy.
-
- Cancer Biology
- Cell Biology
Rapid recovery of proteasome activity may contribute to intrinsic and acquired resistance to FDA-approved proteasome inhibitors. Previous studies have demonstrated that the expression of proteasome genes in cells treated with sub-lethal concentrations of proteasome inhibitors is upregulated by the transcription factor Nrf1 (NFE2L1), which is activated by a DDI2 protease. Here, we demonstrate that the recovery of proteasome activity is DDI2-independent and occurs before transcription of proteasomal genes is upregulated but requires protein translation. Thus, mammalian cells possess an additional DDI2 and transcription-independent pathway for the rapid recovery of proteasome activity after proteasome inhibition.