Quantification of SLIT-ROBO transcripts in hepatocellular carcinoma reveals two groups of genes with coordinate expression.
BACKGROUND
SLIT-ROBO families of proteins mediate axon pathfinding and their expression is not solely confined to nervous system. Aberrant expression of SLIT-ROBO genes was repeatedly shown in a wide variety of cancers, yet data about their collective behavior in hepatocellular carcinoma (HCC) is missing. Hence, we quantified SLIT-ROBO transcripts in HCC cell lines, and in normal and tumor tissues from liver.


METHODS
Expression of SLIT-ROBO family members was quantified by real-time qRT-PCR in 14 HCC cell lines, 8 normal and 35 tumor tissues from the liver. ANOVA and Pearson's correlation analyses were performed in R environment, and different clinicopathological subgroups were pairwise compared in Minitab. Gene expression matrices of cell lines and tissues were analyzed by Mantel's association test.


RESULTS
Genewise hierarchical clustering revealed two subgroups with coordinate expression pattern in both the HCC cell lines and tissues: ROBO1, ROBO2, SLIT1 in one cluster, and ROBO4, SLIT2, SLIT3 in the other, respectively. Moreover, SLIT-ROBO expression predicted AFP-dependent subgrouping of HCC cell lines, but not that of liver tissues. ROBO1 and ROBO2 were significantly up-regulated, whereas SLIT3 was significantly down-regulated in cell lines with high-AFP background. When compared to normal liver tissue, ROBO1 was found to be significantly overexpressed, while ROBO4 was down-regulated in HCC. We also observed that ROBO1 and SLIT2 differentiated histopathological subgroups of liver tissues depending on both tumor staging and differentiation status. However, ROBO4 could discriminate poorly differentiated HCC from other subgroups.


CONCLUSIONS
The present study is the first in comprehensive and quantitative evaluation of SLIT-ROBO family gene expression in HCC, and suggests that the expression of SLIT-ROBO genes is regulated in hepatocarcinogenesis. Our results implicate that SLIT-ROBO transcription profile is bi-modular in nature, and that each module shows intrinsic variability. We also provide quantitative evidence for potential use of ROBO1, ROBO4 and SLIT2 for prediction of tumor stage and differentiation status.
Avci
Mehmet Ender
ME
Department of Molecular Biology and Genetics, Bilkent University, Ankara, Turkey. aender@bilkent.edu.tr
Konu
Ozlen
O
Yagci
Tamer
T
D000704:Analysis of Variance; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D016000:Cluster Analysis; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D046888:Tissue Array Analysis; D000509:alpha-Fetoproteins / Q000235:genetics / Q000378:metabolism; D000097905:Roundabout Proteins; D000098552:Slit Homolog 2 Protein
D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D000509:alpha-Fetoproteins; D000097905:Roundabout Proteins; D000098552:Slit Homolog 2 Protein; C462951:ROBO4 protein, human; C492289:SLIT1 protein, human; C498990:SLIT3 protein, human