Alternative fates of keratinocytes transduced by human papillomavirus type 18 E7 during squamous differentiation.
The human papillomavirus type 18 (HPV-18) E7 protein promotes S-phase reentry in postmitotic, differentiated keratinocytes in squamous epithelium to facilitate vegetative viral DNA amplification. To examine the nature and fate of the differentiated cells that reenter S phase, organotypic cultures of primary human keratinocytes transduced with HPV-18 E7 were pulse-chase-pulse-labeled with (3)H-thymidine ((3)H-TdR) and bromodeoxyuridine (BrdU). The kinetics of the appearance of doubly labeled suprabasal cells demonstrate that E7 expression did not promote prolonged S phase. Rather, there was a considerable lag before a small percentage of the cells reentered another round of S phase. Fluorescence in situ hybridization analysis, indeed, revealed a small fraction of the cells with more than 4n chromosomes in the differentiated strata. Differentiated cells positive for (3)H-TdR, BrdU, or both often had enlarged nuclei or were binucleated. These results suggest that S phase is not followed by cell division, although nuclear division may occur. Interestingly, a significant fraction of differentiated cells that entered S phase subsequently accumulated p27kip1 protein with a kinetics preceding the accumulation of cyclin E. We conclude that E7-transduced, differentiated keratinocytes that enter S phase have two alternative fates: (i) a low percentage of cells undergoes endoreduplication, achieving higher than 4n ploidy, and (ii) a high percentage of cells accumulates the p27kip1, cyclin E, and p21cip1 proteins, resulting in arrest and preventing further S-phase reentry.
Chien
Wei-Ming
WM
Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, Alabama 35294-0005, USA.
Noya
Francisco
F
Benedict-Hamilton
Heather M
HM
Broker
Thomas R
TR
Chow
Louise T
LT
D001345:Autoradiography; D001973:Bromodeoxyuridine / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis*; D004268:DNA-Binding Proteins; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D009856:Oncogene Proteins, Viral / Q000235:genetics* / Q000378:metabolism*; D027383:Papillomaviridae / Q000235:genetics / Q000378:metabolism; D030361:Papillomavirus Infections / Q000821:virology; D011123:Polyploidy; D016196:S Phase; D013936:Thymidine / Q000378:metabolism; D014161:Transduction, Genetic; D014316:Tritium / Q000378:metabolism; D014412:Tumor Virus Infections / Q000821:virology
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C052633:E7 protein, Human papillomavirus type 18; D009856:Oncogene Proteins, Viral; D014316:Tritium; D004247:DNA; D001973:Bromodeoxyuridine; D013936:Thymidine