Monochromosome transfer and microarray analysis identify a critical tumor-suppressive region mapping to chromosome 13q14 and THSD1 in esophageal carcinoma.
Loss of chromosome 13q regions in esophageal squamous cell carcinoma (ESCC) is a frequent event. Monochromosome transfer approaches provide direct functional evidence for tumor suppression by chromosome 13 in SLMT-1, an ESCC cell line, and identify critical regions at 13q12.3, 13q14.11, and 13q14.3. Differential gene expression profiles of three tumor-suppressing microcell hybrids (MCH) and their tumorigenic parental SLMT-1 cell line were revealed by competitive hybridization using 19k cDNA oligonucleotide microarrays. Nine candidate 13q14 tumor-suppressor genes (TSG), including RB1, showed down-regulation in SLMT-1, compared with NE1, an immortalized normal esophageal epithelial cell line; their average gene expression was restored in MCHs compared with SLMT-1. Reverse transcription-PCR validated gene expression levels in MCHs and a panel of ESCC cell lines. Results suggest that the tumor-suppressing effect is not attributed to RB1, but instead likely involves thrombospondin type I domain-containing 1 (THSD1), a novel candidate TSG mapping to 13q14. Quantitative reverse transcription-PCR detected down-regulation of THSD1 expression in 100% of ESCC and other cancer cell lines. Mechanisms for THSD1 silencing in ESCC involved loss of heterozygosity and promoter hypermethylation, as analyzed by methylation-specific PCR and clonal bisulfite sequencing. Transfection of wild-type THSD1 into SLMT-1 resulted in significant reduction of colony-forming ability, hence providing functional evidence for its growth-suppressive activity. These findings suggest that THSD1 is a good candidate TSG.
Ko
Josephine M Y
JM
Department of Biology, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, People's Republic of China.
Chan
Pui Ling
PL
Yau
Wing Lung
WL
Chan
Ho Kin
HK
Chan
King Chi
KC
Yu
Zhuo You
ZY
Kwong
Fung Mei
FM
Miller
Lance D
LD
Liu
Edison T
ET
Yang
Li Chun
LC
Lo
Paulisally H Y
PH
Stanbridge
Eric J
EJ
Tang
Johnny C O
JC
Srivastava
Gopesh
G
Tsao
Sai Wah
SW
Law
Simon
S
Lung
Maria L
ML
D000483:Alleles; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002874:Chromosome Mapping; D020090:Chromosome Segregation; D002882:Chromosomes, Human, Pair 13 / Q000235:genetics*; D019175:DNA Methylation / Q000187:drug effects; D003841:Deoxycytidine / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology; D004938:Esophageal Neoplasms / Q000235:genetics*; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D016147:Genes, Tumor Suppressor; D015894:Genome, Human / Q000235:genetics; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D017404:In Situ Hybridization, Fluorescence; D046228:Microarray Analysis; D018895:Microsatellite Repeats / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019699:Thrombospondins / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014410:Tumor Stem Cell Assay
D006877:Hydroxamic Acids; C511241:THSD1 protein, human; D019699:Thrombospondins; D003841:Deoxycytidine; C012589:trichostatin A