Small interfering RNA-mediated knockdown of PRL phosphatases results in altered Akt phosphorylation and reduced clonogenicity of pancreatic cancer cells.
The PRL phosphatases have been implicated in cancer cell growth and metastasis in a variety of tumor types. Using cDNA microarray, we previously identified and reported PRL-1 as being highly up-regulated in pancreatic cancer cell lines. In this study, we sought to further evaluate the expression of all three PRL phosphatases in pancreatic cancer cell lines and extend our findings to in situ analysis of primary pancreatic tumors taken directly from patients. Additionally, we determine if small interfering RNA-mediated knockdown of relevant PRLs confers antitumor effects in pancreatic cancer cells. Using oligonucleotide expression arrays, mRNA levels of PRL-1 and PRL-2 but not PRL-3 were identified as up-regulated in pancreatic cancer cell lines and tumor samples taken directly from patients compared with those of normal pancreas. Focusing on PRL-1 and PRL-2, high levels of both proteins were detected in a subset of pancreatic cancer cell lines and tumor samples using Western blotting and immunohistochemistry, respectively. Small interfering RNA-mediated knockdown of PRL-1 and PRL-2 in combination resulted in a moderate reduction of cellular growth and migration in MIA PaCa-2 and PANC-1 cells. More importantly, knockdown of both PRL-1 and PRL-2 significantly inhibited colony formation of these cells in soft agar as well as serum-induced Akt phosphorylation. These data support the hypothesis that PRL phosphatases regulate key pathways involved in tumorigenesis and metastasis and that knockdown of both PRL-1 and PRL-2 is required to disrupt PRL phosphatase function in pancreatic cancer cells.
Stephens
Bret
B
Department of Molecular and Cellular Biology, University of Arizona, Tucson, USA. bstephens@tgen.org
Han
Haiyong
H
Hostetter
Galen
G
Demeure
Michael J
MJ
Von Hoff
Daniel D
DD
D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D020935:MAP Kinase Signaling System; D010190:Pancreatic Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics*; D012680:Sensitivity and Specificity; D014158:Transcription, Genetic / Q000235:genetics; D014410:Tumor Stem Cell Assay
D034741:RNA, Small Interfering; D051057:Proto-Oncogene Proteins c-akt; D017027:Protein Tyrosine Phosphatases