5015

OTX2

CPHD6|MCOPS5;orthodenticle homeobox 2;OTX2;orthodenticle homeobox 2

homeobox protein OTX2|orthodenticle homolog 2

14q22.3

protein-coding

Serial analysis of gene expression (SAGE) was used to identify genes that might be involved in the development or growth of medulloblastoma, a childhood brain tumor. Sequence tags from medulloblastoma (10229) and fetal brain (10692) were determined. The distributions of sequence tags in each population were compared,and for each sequence tag, pairwise chi2 test statistics were calculated. Northern blot was used to confirm some of the results obtained by SAGE. For 16 tags, the chi2 test statistic was associated with a P value < 10(-4). Among those transcripts with a higher expression in medulloblastoma were the genes for ZIC1 protein and the OTX2 gene, both of which are expressed in the cerebellar germinal layers. The high expression of these two genes strongly supports the hypothesis that medulloblastoma arises from the germinal layer of the cerebellum. This analysis shows that SAGE can be used as a rapid differential screening procedure.

Both OTX2 and MYC are important oncogenes in medulloblastoma, the most common malignant brain tumor in childhood. Much is known about MYC binding to promoter regions, but OTX2 binding is hardly investigated. We used ChIP-on-chip data to analyze the binding patterns of both transcription factors in D425 medulloblastoma cells. When combining the data for ALL promoter regions in the genome, OTX2 binding showed a remarkable bi-modal distribution pattern with peaks around -250 bp upstream and +650 bp downstream of the transcription start sites (TSSs). Indeed, 40.2% of ALL OTX2-bound TSSs had more than one significant OTX2-binding peak. This OTX2-binding pattern was very different from the TSS-centered single peak binding pattern observed for MYC and other known transcription factors. However, in individual promoter regions, OTX2 and MYC have a strong tendency to bind in proximity of each other. OTX2-binding sequences aredepleted near TSSs in the genome, providing an explanation for the observed bi-modal distribution of OTX2 binding. This contrasts to the enrichment of E-boxsequences at TSSs. Both OTX2 and MYC binding independently correlated with higher gene expression. Interestingly, genes of promoter regions with multiple OTX2 binding as well as MYC binding showed the highest expression levels in D425 cells and in primary medulloblastomas. Genes within this class of promoter regions were enriched for medulloblastoma and stem cell specific genes. Our data suggest an important functional interaction between OTX2 and MYC in regulating gene expression in medulloblastoma.

The OTX2 gene encodes a key transcription factor in developmental processes. In particular, it is required for the early specification of the brain and the embryonic development of sensory organs, including the pituitary gland, pineal gland, inner part of the ear, eyes, and optic nerve. In later stages, it is important for maintaining intact retina and brain function. Overexpression of OTX2 was found in medulloblastoma, a malignant brain tumor that occurs mainly inchildhood. OTX2 mutations are associated with severe ocular phenotypes and central nervous system abnormalities such as seizures, short stature and developmental delay, combined pituitary hormone deficiency (CPHD), structural abnormalities of the pituitary gland and early onset retinal dystrophy. There isno genotype-phenotype correlation and the severity of the disease varies not only by the specific OTX2 mutations but also among individuals harboring the same mutation, suggesting the involvement of multiple levels of regulation.