SMAD5 gene expression, rearrangements, copy number, and amplification at fragile site FRA5C in human hepatocellular carcinoma.
Signaling by the transforming growth factor (TGF)-family members is transduced from the cell surface to the nucleus by the Smad group of intracellular proteins. Because we detected alterations on the long arm of chromosome 5, we examined the status of the SMAD5 gene in human hepatocellular carcinoma (HCC) cell lines and primary HCC. In 16 cell lines, chromosome alterations of chromosome 5 were observed in nine cell lines by fluorescence in situ hybridization (FISH), and an increase in SMAD5 gene copy number relative to the ploidy level was found in eight lines. The breakpoints in unbalanced translocations and deletions frequently occurred near the SMAD5 locus, but apparently did not cause loss of SMAD5. In one cell line, where comparative genomic hybridization showed DNA copy number gain confined to the region 5q31, we detected by FISH high-level amplification of the SMAD5 gene located within the fragile site FRA5C. Semiquantitative polymerase chain reaction did not reveal changes in SMAD5 DNA levels in 15 of 17 primary HCC specimens. In 17 HCC cell lines, SMAD5 mRNA levels were either maintained or upregulated by an increase in gene dosage or another mechanism. Collectively, our results show that SMAD5 undergoes copy number gain and increased expression, rather than loss of expression, and therefore suggest that this gene does not act as a tumor-suppressor gene in HCC. The Hep-40 HCC cell line with high-level amplification and significant overexpression of SMAD5 may be useful in studying the interaction of SMAD5 with other genes.
Zimonjic
Drazen B
DB
Laboratory of Experimental Carcinogenesis, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
Durkin
Marian E
ME
Keck-Waggoner
Catherine L
CL
Park
Sang-Won
SW
Thorgeirsson
Snorri S
SS
Popescu
Nicholas C
NC
D015152:Blotting, Northern; D015139:Blotting, Southern; D006528:Carcinoma, Hepatocellular / Q000235:genetics*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002875:Chromosomes / Q000648:ultrastructure; D004247:DNA / Q000648:ultrastructure; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics*; D017353:Gene Deletion; D018628:Gene Dosage; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008113:Liver Neoplasms / Q000235:genetics*; D010750:Phosphoproteins / Q000096:biosynthesis* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D051902:Smad5 Protein; D015534:Trans-Activators / Q000096:biosynthesis* / Q000235:genetics*; D014178:Translocation, Genetic
D004268:DNA-Binding Proteins; D010750:Phosphoproteins; D012333:RNA, Messenger; C497797:SMAD5 protein, human; D051902:Smad5 Protein; D015534:Trans-Activators; D004247:DNA