Suppression of anchorage-independent growth and matrigel invasion and delayed tumor formation by elevated expression of fibulin-1D in human fibrosarcoma-derived cell lines.
Using differential display, we identified an mRNA that is markedly down-regulated in cell line 6A/SB1, derived from a fibrosarcoma formed in an athymic mouse following injection of carcinogen-transformed MSU-1.1 cells. The nontumorigenic parental cell strain, MSU-1.1, expresses high levels of this mRNA. Sequencing of the corresponding cDNA fragment revealed that it corresponded to an expressed sequence tag, which ultimately led to its identification as the fibulin-1D gene. Fibulin-1 is a cysteine-rich, calcium-binding extracellular matrix and plasma protein, which has four isoforms, A-D, derived from alternative splicing. Northern and Western blotting analysis of 16 cell lines established from tumors formed in athymic mice by MSU-1.1-derived cell strains independently transformed in culture showed that 44% exhibited low level or lack of expression of fibulin-1D mRNA and protein. In a similar analysis of 15 malignant cell lines derived from patients, 80% showed low level or no expression. To study the role of fibulin-1D in transformation, we transfected 6A/SB1 cells and a human fibrosarcoma-derived cell line (SHAC) with a fibulin-1D cDNA expression construct. Transfectants displaying high levels of fibulin-1D were isolated and characterized. Elevated expression of fibulin-1D led to reduced ability to form colonies in soft agar and reduced invasive potential as tested in a matrigel in vitro invasion assay. Furthermore, expression of fibulin-1D resulted in a markedly extended latency in tumor formation in athymic mice. These results indicate that low expression of fibulin-1D plays a role in tumor formation and invasion.
Qing
J
J
Department of Biochemistry, The Cancer Center, Michigan State University, East Lansing 48824, USA.
Maher
V M
VM
Tran
H
H
Argraves
W S
WS
Dunstan
R W
RW
McCormick
J J
JJ
D000818:Animals; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D002448:Cell Adhesion / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002471:Cell Transformation, Neoplastic; D003094:Collagen; D004338:Drug Combinations; D005354:Fibrosarcoma / Q000378:metabolism* / Q000473:pathology*; D006801:Humans; D007797:Laminin; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009361:Neoplasm Invasiveness; D011509:Proteoglycans; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D002135:Calcium-Binding Proteins; D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; D012333:RNA, Messenger; C060820:fibulin; C059485:matrigel; D003094:Collagen