Identification of two novel phenotypically distinct breast cancer cell subsets based on Sox2 transcription activity.
Sox2 (sex-determining region Y-box protein 2) is a transcription factor regulating pluripotency in embryonic stem cells. Sox2 is aberrantly expressed in breast and other cancers, though its biological significance remains widely unexplored. To understand the significance of this aberrancy, we assessed the transcription activity of Sox2 in two Sox2-expressing breast cancer cell lines, MCF7 and ZR751, using a lentiviral Sox2 GFP reporter vector. Surprisingly, Sox2 transcription activity, as measured by GFP expression encoded in a Sox2 reporter construct, was detectable only in a small subset of cells in both cell lines. Purification of GFP+ cells (cells with Sox2 activity) and GFP- cells (cells without Sox2 activity) was enriched for two phenotypically distinct cell populations in both MCF7 and ZR751 cell lines. Specifically, GFP+ cells formed significantly more colonies in methylcellulose and more mammospheres in vitro compared to GFP- cells. These phenotypic differences are directly linked to Sox2 as siRNA knockdown of Sox2 in GFP+ cells abolished these abilities. To provide a mechanistic explanation to our observations, we performed gel shift and chromatin immunoprecipitation studies; Sox2 was found to bind to its DNA binding consensus sequence and the promoters of Cyclin D1 and Nanog (two known Sox2 downstream targets) only in GFP+ cells. GFP+ cells also up-regulated CD49f, phospho-GSK3β, and β-catenin. In summary, we have identified two novel phenotypically distinct cell subsets in two breast cancer cell lines based on their differential Sox2 transcription activity. We demonstrate that Sox2 transcription activity, and not its protein expression alone, underlies the tumorigenicity and cancer stem cell-like phenotypes in breast cancers.
Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Alberta, Canada.
Fang
F
Wu
Jingdong
J
Zhang
Peng
P
Wang
Xiaoxia
X
Ye
Karen
K
Jung
Kathleen M
KM
Bone
Joel D
JD
Pearson
Robert J
RJ
Ingham
Todd P
TP
McMullen
Yupo
Y
Ma
Raymond
R
Lai
D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019938:Cyclin D1 / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D005260:Female; D017930:Genes, Reporter; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D039503:Integrin alpha6 / Q000378:metabolism; D061986:MCF-7 Cells; D000071317:Nanog Homeobox Protein; D010641:Phenotype; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D051176:beta Catenin / Q000378:metabolism
D018398:Homeodomain Proteins; D039503:Integrin alpha6; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D034741:RNA, Small Interfering; D055748:SOXB1 Transcription Factors; D051176:beta Catenin; D019938:Cyclin D1; D004247:DNA; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D038362:Glycogen Synthase Kinase 3