PAX6 suppresses the invasiveness of glioblastoma cells and the expression of the matrix metalloproteinase-2 gene.
Glioblastoma multiforme (GBM) is the most invasive brain tumor. We have previously reported that the transcription factor PAX6 suppresses the tumorigenecity of GBM cells. By an in vitro Matrigel invasion assay on two GBM cell lines stably transfected with wild-type and/or two mutant forms of PAX6, this study displays the first evidence that PAX6 inhibits the invasiveness of GBM cells and that the DNA-binding domain of PAX6 is required for this function. Using real-time quantitative reverse transcription-PCR (RT-PCR), gelatin zymography, and immunohistochemistry assays, the expression of the gene encoding matrix metalloproteinase-2 (MMP2) in GBM cell lines grown in vitro or in intracranial xenografts in nude mice was shown to be repressed by either stable or adenoviral-mediated overexpression of PAX6. Luciferase promoter assays revealed PAX6-mediated suppression of MMP2 promoter activity. Electrophoretic mobility shift assays showed direct binding of PAX6 to the MMP2 promoter. A significant reverse correlation (P < 0.05) occurred between PAX6 and MMP2 expression quantified by real-time quantitative RT-PCR in 41 GBMs, 43 anaplastic astrocytomas, and 7 adjacent normal tissues. Interestingly, the degree and significance of the reverse correlation increased after excluding astrocytomas, whereas it became insignificant after excluding GBMs. In GBM cells stably transfected with a dominant negative mutant PAX6 showing increased MMP2 expression and invasiveness, knock-down of MMP2 revealed that MMP2 is one of the PAX6 target genes mediating its suppression of invasion. Overall data delineated a mechanism for the suppressive function of PAX6 in GBM: suppression of cell invasion by repressing the expression of proinvasive genes such as MMP2.
Mayes
Debra A
DA
Department of Neurobiology and Developmental Sciences, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA.
Hu
Yuanjie
Y
Teng
Yue
Y
Siegel
Eric
E
Wu
Xiaosong
X
Panda
Kishori
K
Tan
Fang
F
Yung
W K Alfred
WK
Zhou
Yi-Hong
YH
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001254:Astrocytoma / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D001932:Brain Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D045744:Cell Line, Tumor; D005136:Eye Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D018398:Homeodomain Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D009361:Neoplasm Invasiveness; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D014183:Transplantation, Heterologous
D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D012097:Repressor Proteins; D020778:Matrix Metalloproteinase 2