Mutation analysis of the apoptotic "death-receptors" and the adaptors TRADD and FADD/MORT-1 in osteosarcoma tumor samples and osteosarcoma cell lines.
Apoptosis is a key mechanism of the organism that regulates embryogenesis and development, maintains homeostasis of the immune system and removes potentially hazardous cells. A dysregulation of apoptosis signaling may thus disturb the balance of cell survival and cell death, leading to the development of several diseases including cancer. In order to determine whether osteosarcomas display an increased frequency of genetic alterations that affect apoptosis signaling, we analyzed the death domains of the death receptor genes CD95/Fas/Apo1, TNFR1, DR3/Apo3/WSL-1/LARD/TRAMP, DR5/TRAIL-R2/TRICK2/KILLER, DR6 and the complete coding sequences of the death receptor gene DR4/TRAIL-R1 and the genes of the adaptors TRADD and FADD/MORT-1. The investigation included 15 osteosarcoma tumor samples, 3 osteosarcoma cell lines (SAOS-2, HOS and MG63) and peripheral blood from 20 donors as controls. We were able to identify 4 different sequence variations within the DR4 gene located on exons 3, 4, 5 and 10 (death-domain). No alterations have been detected in the other genes or exons investigated. Except the sequence variant affecting exon 4, the alterations were homozygous in 15% of the tumor samples and cell lines, whereas the same alterations found in the control group were heterozygous or even not detectable. Three out of 4 alterations are located in the receptor's extracellular cysteine rich domain, which contains the ligand binding area and 1 on exon 10 coding for the death-domain. They may thus exert influence on ligand-receptor interactions and subsequent apoptosis induction. Our findings suggest that homozygous genetic alterations within the DR4 gene may be implicated in the formation of osteosarcoma.
Department of Orthopaedic Research, University of Heidelberg, Schlierbacher Landstrasse 200a, 69118 Heidelberg, Germany.
Markus J
MJ
Dechant
Jörg
J
Fellenberg
Christian G
CG
Scheuerpflug
Volker
V
Ewerbeck
Klaus-Michael
KM
Debatin
D048868:Adaptor Proteins, Signal Transducing; D015703:Antigens, CD / Q000235:genetics; D017209:Apoptosis / Q000235:genetics*; D001859:Bone Neoplasms / Q000235:genetics* / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics*; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis; D017931:DNA Primers; D004273:DNA, Neoplasm / Q000032:analysis; D005091:Exons; D053200:Fas-Associated Death Domain Protein; D006720:Homozygote; D006801:Humans; D009154:Mutation; D020125:Mutation, Missense; D012516:Osteosarcoma / Q000235:genetics* / Q000378:metabolism; D012150:Polymorphism, Restriction Fragment Length; D011506:Proteins / Q000235:genetics*; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor / Q000235:genetics*; D053219:Receptors, Tumor Necrosis Factor, Member 25; D047888:Receptors, Tumor Necrosis Factor, Type I; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D047990:TNF Receptor-Associated Factor 1; D019014:fas Receptor / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; D015703:Antigens, CD; D002352:Carrier Proteins; D017931:DNA Primers; D004273:DNA, Neoplasm; C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; D011506:Proteins; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor; D053219:Receptors, Tumor Necrosis Factor, Member 25; D047888:Receptors, Tumor Necrosis Factor, Type I; D047990:TNF Receptor-Associated Factor 1; C105190:TNFRSF10A protein, human; C505639:TNFRSF10B protein, human; C114109:TNFRSF21 protein, human; C506033:TNFRSF25 protein, human; D019014:fas Receptor