title: Differential gene expression in nuclear label-retaining cells in the developing mouse mammary gland.
abstract: The immortal strand theory postulates stem cells protect themselves from DNA replication-associated mutations and subsequent cancer risk through selective segregation of template DNA strands. Stem cells self-renew by asymmetric cellular division. During asymmetric division, stem cells maintain their template DNA strands, while the newly synthesized DNA strands segregate to newly formed daughter cells. Previous studies have demonstrated that self-renewing mammary stem cells originate in the expanding mammary ducts during puberty-associated allometric growth. In this study, we labeled newly forming mammary stem cells with the thymidine analog 5-ethynl-2'-deoxyuridine for 2 weeks during allometric ductal expansion. Cells that incorporate and retain the nuclear label following extended chase periods are termed label-retaining cells (LRCs). A second nuclear label, 5-bromodeoxyuridine, was administered before euthanasia to identify cells traversing the cell cycle. Mammary cells collected following euthanasia were sorted based on nuclear label retention. Members of the Notch and Wnt signaling pathways were found differentially expressed by mammary LRCs. These pathways are involved in the regulation of stem cells in the mouse mammary gland. Upon further analysis, we found that in contrast to non-LRCs, Notch1 and Notch2 are expressed and localized in the nuclei of the LRCs. Expression of Notch-inducible genes, Hes1 and Hey2, was elevated in LRCs. Inhibition of Notch1 by shRNA reduced colony forming potential and label retention by mammary epithelial cells in vitro. These results indicate that genes are differentially regulated in the LRC population of mammary glands and Notch1 mediates asymmetric cell division of mammary progenitor cells.
authors.affiliation: Institute for Biological Interfaces of Engineering, Clemson University, Clemson, South Carolina 29634, USA.
authors.forename: Jang Pyo
authors.initials: JP
authors.lastname: Park
authors.forename: Ahmed
authors.initials: A
authors.lastname: Raafat
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Feltracco
authors.forename: Walker M
authors.initials: WM
authors.lastname: Blanding
authors.forename: Brian W
authors.initials: BW
authors.lastname: Booth
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000008:administration & dosage / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002478:Cells, Cultured; D003857:Deoxyuridine / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D034622:RNA Interference; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism; D051883:Receptor, Notch2 / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics; D013194:Staining and Labeling / Q000379:methods; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D000072056:Transcription Factor HES-1; D060449:Wnt Signaling Pathway / Q000235:genetics
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D001973:Bromodeoxyuridine; D003857:Deoxyuridine; D018398:Homeodomain Proteins; D051881:Receptor, Notch1; D051883:Receptor, Notch2; D012097:Repressor Proteins; D000072056:Transcription Factor HES-1; C489734:Hes1 protein, mouse; C485778:Hey2 protein, mouse; C031086:5-ethynyl-2'-deoxyuridine