• Cellular retinaldehyde-binding protein (CRALBP) is a direct downstream target of transcription factor Pax6.

      Boppana, Sridhar; Scheglov, Alexander; Geffers, Robert; Tarabykin, Victor; Max-Planck-Institute for Experimental Medicine, Hermann-Rein Strasse 3, 37075 Göttingen, Germany. boppansr@umdnj.edu (2012-02)
      Transcription factor Pax6 plays an essential role in the expression of other transcription factors, cell adhesion molecules and is crucial for neurogenesis in the developing forebrain. Analysis of gene expression profiles through microarray experiments in Pax6 mutants allowed us to focus on CRALBP, one of the many genes that were downregulated.
    • Genome-Wide Sequencing Reveals MicroRNAs Downregulated in Cerebral Cavernous Malformations.

      Kar, Souvik; Bali, Kiran Kumar; Baisantry, Arpita; Geffers, Robert; Samii, Amir; Bertalanffy, Helmut; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017-02)
      Cerebral cavernous malformations (CCM) are vascular lesions associated with loss-of-function mutations in one of the three genes encoding KRIT1 (CCM1), CCM2, and PDCD10. Recent understanding of the molecular mechanisms that lead to CCM development is limited. The role of microRNAs (miRNAs) has been demonstrated in vascular pathologies resulting in loss of tight junction proteins, increased vascular permeability and endothelial cell dysfunction. Since the relevance of miRNAs in CCM pathophysiology has not been elucidated, the primary aim of the study was to identify the miRNA-mRNA expression network associated with CCM. Using small RNA sequencing, we identified a total of 764 matured miRNAs expressed in CCM patients compared to the healthy brains. The expression of the selected miRNAs was validated by qRT-PCR, and the results were found to be consistent with the sequencing data. Upon application of additional statistical stringency, five miRNAs (let-7b-5p, miR-361-5p, miR-370-3p, miR-181a-2-3p, and miR-95-3p) were prioritized to be top CCM-relevant miRNAs. Further in silico analyses revealed that the prioritized miRNAs have a direct functional relation with mRNAs, such as MIB1, HIF1A, PDCD10, TJP1, OCLN, HES1, MAPK1, VEGFA, EGFL7, NF1, and ENG, which are previously characterized as key regulators of CCM pathology. To date, this is the first study to investigate the role of miRNAs in CCM pathology. By employing cutting edge molecular and in silico analyses on clinical samples, the current study reports global miRNA expression changes in CCM patients and provides a rich source of data set to understand detailed molecular machinery involved in CCM pathophysiology.
    • Methylome analysis and integrative profiling of human HCCs identify novel protumorigenic factors.

      Neumann, Olaf; Kesselmeier, Miriam; Geffers, Robert; Pellegrino, Rossella; Radlwimmer, Bernhard; Hoffmann, Katrin; Ehemann, Volker; Schemmer, Peter; Schirmacher, Peter; Lorenzo Bermejo, Justo; et al. (2012-11)
      To identify new tumor-suppressor gene candidates relevant for human hepatocarcinogenesis, we performed genome-wide methylation profiling and vertical integration with array-based comparative genomic hybridization (aCGH), as well as expression data from a cohort of well-characterized human hepatocellular carcinomas (HCCs). Bisulfite-converted DNAs from 63 HCCs and 10 healthy control livers were analyzed for the methylation status of more than 14,000 genes. After defining the differentially methylated genes in HCCs, we integrated their DNA copy-number alterations as determined by aCGH data and correlated them with gene expression to identify genes potentially silenced by promoter hypermethylation. Aberrant methylation of candidates was further confirmed by pyrosequencing, and methylation dependency of silencing was determined by 5-aza-2'-deoxycytidine (5-aza-dC) treatment. Methylation profiling revealed 2,226 CpG sites that showed methylation differences between healthy control livers and HCCs. Of these, 537 CpG sites were hypermethylated in the tumor DNA, whereas 1,689 sites showed promoter hypomethylation. The hypermethylated set was enriched for genes known to be inactivated by the polycomb repressive complex 2, whereas the group of hypomethylated genes was enriched for imprinted genes. We identified three genes matching all of our selection criteria for a tumor-suppressor gene (period homolog 3 [PER3], insulin-like growth-factor-binding protein, acid labile subunit [IGFALS], and protein Z). PER3 was down-regulated in human HCCs, compared to peritumorous and healthy liver tissues. 5-aza-dC treatment restored PER3 expression in HCC cell lines, indicating that promoter hypermethylation was indeed responsible for gene silencing. Additionally, functional analysis supported a tumor-suppressive function for PER3 and IGFALS in vitro. CONCLUSION: The present study illustrates that vertical integration of methylation data with high-resolution genomic and transcriptomic data facilitates the identification of new tumor-suppressor gene candidates in human HCC.