p53-dependent up-regulation of CDKN1A and down-regulation of CCNE2 in response to beryllium.
OBJECTIVE
Beryllium salts (here, beryllium sulphate) can produce a cytostatic effect in some cell types. The basis for this effect may include increased expression of proliferation inhibitors, reduced expression of proliferation promoters, or both. This study sought to determine the role of p53, the tumour-suppressing transcription factor, in mediating beryllium-induced cytostasis.


METHODS
Human A172 glioma cells express wild-type TP53 gene. Activity of p53 was experimentally manipulated using siRNA and related approaches. Key elements of the beryllium-response were compared in normal and p53-knockdown A172 cells using RT-PCR and Western blotting.


RESULTS
In A172 cells, 10 μm BeSO4caused 300% increase in CDKN1A (cyclin-dependent kinase inhibitor p21) mRNA and 90% reduction of CCNE2 (cyclin E2) mRNA. The increased p21 mRNA and reduced cyclin E2 mRNA were each dependent on presence of functional p53. For p21, increased mRNA led to commensurately increased protein levels. In contrast, reduction in cyclin E2 mRNA levels did not lead to corresponding reductions in cyclin E2 protein. The proteasomal inhibitor MG-132 caused p53 protein to increase, but it had no effect on cyclin E2 protein levels. Cycloheximide time course studies indicated that the cyclin E2 protein half-life was more than 12 hours in these cells.


CONCLUSIONS
Beryllium elicited p53-dependent changes in mRNA levels of key determinants of cell proliferation such as p21 and cyclin E2. However, cyclin E2 protein appeared to be aberrantly regulated in this cell type, as its turnover was unexpectedly slow.
Department of Chemistry and Biochemistry, University of Nevada Las Vegas, Las Vegas, NV, USA.
P
P
Gorjala
Department of Clinical Neurosciences, University of Calgary, Calgary, AB, Canada.
J G
JG
Cairncross
Department of Chemistry and Biochemistry, University of Nevada Las Vegas, Las Vegas, NV, USA. ronald.gary@unlv.edu.
R K
http://orcid.org/0000-0001-5079-1953
RK
Gary
D000970:Antineoplastic Agents / Q000494:pharmacology*; D001608:Beryllium / Q000494:pharmacology*; D001921:Brain / Q000187:drug effects / Q000378:metabolism; D001932:Brain Neoplasms / Q000188:drug therapy* / Q000235:genetics; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics*; D016213:Cyclins / Q000235:genetics*; D054697:Cytostatic Agents / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D005909:Glioblastoma / Q000188:drug therapy* / Q000235:genetics; D006801:Humans; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*; D015854:Up-Regulation / Q000187:drug effects
D000970:Antineoplastic Agents; C503688:CCNE2 protein, human; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D054697:Cytostatic Agents; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; C020711:beryllium sulfate; D001608:Beryllium