Role of p53 in UVB-induced apoptosis in human HaCaT keratinocytes

被引:83
作者
Henseleit, U [1 ]
Zhang, J [1 ]
Wanner, R [1 ]
Haase, I [1 ]
Kolde, G [1 ]
Rosenbach, T [1 ]
机构
[1] HUMBOLDT UNIV BERLIN,VIRCHOW CLIN,DEPT DERMATOL,D-13344 BERLIN,GERMANY
关键词
cell death; p53; transfection; protein synthesis inhibition;
D O I
10.1111/1523-1747.ep12340708
中图分类号
R75 [皮肤病学与性病学];
学科分类号
100206 ;
摘要
Apoptosis represents an active form of cell death that is involved in the control of tissue homeostasis and in the deletion of DNA-damaged cells. Because the product of the tumor suppressor gene p53 has been demonstrated to be crucial for the induction of apoptosis in certain cell types, the present study was aimed at elucidating its role in ultraviolet-induced apoptosis in HaCaT keratinocytes. After in vitro ultraviolet B irradiation, p53 protein levels were noted to increase prior to the induction of apoptosis in a time-and concentration-dependent fashion. This increase could not be inhibited by the protein synthesis inhibitor cycloheximide, Because HaCaT keratinocytes are known to bear two p53 point mutations and because it is unclear whether p53 in HaCaT cells is still functional regarding induction of apoptosis, HaCaT cells were stably transfected with wild-type p53 cDNA inserted into the expression vector pCMV-Neo-Bam iu. sense (pC53-SN3) and anti-sense (pC53-ASN) direction. After selection With geniticin, growing colonies were screened for the presence of the transfected cDNA constructs by polymerase chain reaction. Cell clones bearing the anti-sense product were further analyzed for p53 expression by western blotting. Clones showing reduced p53 protein levels were irradiated with ultraviolet B light, and there was a clear reduction of apoptosis in the pC53-ASN bearing cell clones compared with the parental HaCaT cells. These studies demonstrate that blocking mutated p53 can partially block apoptosis in HaCaT keratinocytes and furthermore can confirm the key role for p53 in ultraviolet-induced apoptosis in human keratinocytes. Moreover, HaCaT keratinocytes and their p53-transfectants provide a convenient model that allows for further detailed analyses of apoptosis-associated biochemical sand molecular events in human keratinocytes.
引用
收藏
页码:722 / 727
页数:6
相关论文
共 51 条
  • [11] ELDEIRY WS, 1994, CANCER RES, V54, P1169
  • [12] GONG JP, 1993, J CELL PHYSIOL, V157, P263
  • [13] Hall PA, 1996, J PATHOL, V180, P1
  • [14] HALL PA, 1993, ONCOGENE, V8, P203
  • [15] INDUCTION OF APOPTOSIS IN HELA-CELLS BY TRANS-ACTIVATION-DEFICIENT P53
    HAUPT, Y
    ROWAN, S
    SHAULIAN, E
    VOUSDEN, KH
    OREN, M
    [J]. GENES & DEVELOPMENT, 1995, 9 (17) : 2170 - 2183
  • [16] DISSOCIATION OF ERYTHEMA AND P53 PROTEIN EXPRESSION IN HUMAN SKIN FOLLOWING UVB IRRADIATION, AND INDUCTION OF P53 PROTEIN AND MESSENGER-RNA FOLLOWING APPLICATION OF SKIN IRRITANTS
    HEALY, E
    REYNOLDS, NJ
    SMITH, MD
    CAMPBELL, C
    FARR, PM
    REES, JL
    [J]. JOURNAL OF INVESTIGATIVE DERMATOLOGY, 1994, 103 (04) : 493 - 499
  • [17] Induction of apoptosis in human HaCaT keratinocytes
    Henseleit, U
    Rosenbach, T
    Kolde, G
    [J]. ARCHIVES OF DERMATOLOGICAL RESEARCH, 1996, 288 (11) : 676 - 683
  • [18] HERZINGER T, 1995, ONCOGENE, V11, P2151
  • [19] Hockenbery D M, 1992, Semin Immunol, V4, P413
  • [20] P53 MUTATIONS IN HUMAN CANCERS
    HOLLSTEIN, M
    SIDRANSKY, D
    VOGELSTEIN, B
    HARRIS, CC
    [J]. SCIENCE, 1991, 253 (5015) : 49 - 53