1 Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158, USA
2 Radboud University, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, the Netherlands
3 Department of Physics and Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA
4 Gladstone Center for Cell Circuitry, Gladstone Institutes, San Francisco, CA 94158, USA
5 Departments of Pharmaceutical Chemistry and Biochemistry and Biophysics, University of California, San Francisco, CA 94158, USA
6 Huntsman Cancer Institute, University of Utah, Salt Lake City, UT 84112, USA
7 Department of Dermatology, University of Utah School of Medicine, Salt Lake City, UT 84103, USA
Phenotypic distribution of the stem-like melanoma initiating state depends on CDKN2A status
August 27, 2020 Länk till PDFAbstract
Many cancers contain distinct tumor-initiating cell populations.
However, the existence of distinct stem-like melanoma initiating cells and their contribution to tumorigenesis remains contested1–5.
To identify this cell population in melanoma, we used quantitative single cell approaches linking gene expression, genotype and phenotype in melanoma cells, and observed that bidirectional interconversion between tumor-initiating and differentiated non-tumorigenic states establishes distinct phenotypic equilibria dependent on genotype.
Genetic loss of the CDKN2A locus corresponds to a uniform adoption of a neural crest stem cell (NCSC) like tumorinitiating state.
Exposure to a putative chemopreventative α-melanocyte stimulating hormone (αMSH) analog can substitute for CDKN2A loss and shift phenotype distribution toward the tumor-initiating state. Alarmingly, in vivo application of the analog is sufficient to induce tumorigenesis in otherwise non-tumorigenic populations.
Our results demonstrate that dynamic stemness in melanoma is dependent on secondary mutation status, highlighting the need to incorporate genomic characterization when developing potential chemopreventative agents.
Methods (urval)
Measurement of morphology and motility:
100,000 cells were seeded in a standard tissue‐culture treated 6‐well polystyrene plate (Sarstedt, 83.3920.005) for digital holographic cytometry (DHC) using the M4 Holomonitor (Phase Holographic Imaging, Sweden).
Cells were monitored for 72 hours. Motility and morphology were analyzed using the HStudio Software as previously described 3,14.
Comparison of cell morphologies was conducted as previously described 15.
For quantitative phase images coupled with mCherry fluorescence, cells were imaged with the LiveCyte (Phasefocus, United Kingdom) and similarly analyzed using manufacture’s software.
I deras slutsatser saxar jag ett urval.
- Using quantitative phase cytometry, we conducted single cell phenotype profiling38,39 (Fig. 2e),and unexpectedly identified the POU3F2 reporter high population to be moderately less motile
than the reporter low counterparts (Fig. 2f) – a result confirmed via scratch assay (Fig. 2g).
To determine whether either population were enriched for TICs, FACS-isolated reporter low and high cells were implanted subcutaneously into NOD scid gamma mice and tumor growth was monitored for five weeks.
In conclusion, we demonstrate that cancer cell phenotypic equilibria depend on the mutational landscape.
Subtle shifts in phenotypic equilibria may promote tumor progression through induction of minor stem-like subpopulations, highlighting the importance of quantitative approaches with single-cell resolution.
We also suggest a broader role for the tumor suppressor, CDKN2A, in regulating global cell state plasticity in addition to its established role as a cell cycle inhibitor.
This study improves our understanding of how secondary mutation status alters cell state plasticity and, consequently, the effect of environmental cues on tumor progression.
Min kommentar
Robbie levererar återigen helt ny kunskap inom hudforskningen.Det med benägen assistans av omistliga HoloMonitor naturligtvis.Han kommer återses i Stockholm om x år i den årliga utdelningen av det finaste priset en forskare kan få.Det sätter jag 1 spänn på.
Bra jobbat Robbie m kollegor !
Mvh the99
Inga kommentarer:
Skicka en kommentar