Purification and characterization of novel heparin-binding growth factors in uterine secretory fluids. Identification as heparin-regulated Mr 10,000 forms of connective tissue growth factor.
Uterine growth factors are potential effector molecules in embryo growth signaling pathways. Pig uterine luminal flushings contained a heparin-binding growth factor (HBGF) that required 0.8 M NaCl for elution from heparin columns and was termed HBGF-0.8. This factor, which was heat- and acid-labile and of Mr 10,000 as assessed by gel filtration, stimulated DNA synthesis in fibroblasts and smooth muscle cells but not endothelial cells. Two forms of HBGF-0.8, termed HBGF-0.8-P1 and HBGF-0.8-P2, exhibited differential heparin-binding properties. SDS-polyacrylamide gel electrophoresis showed that each form of HBGF-0.8 migrated with an apparent Mr of 10, 000 under reducing conditions. Amino acid sequencing revealed the N-terminal sequence EENIKKGKKXIRTPKI for HBGF-0.8-P1 and ENIKKGKKXIRT for HBGF-0.8-P2. These sequences corresponded, respectively, to residues 247-262 and 248-259 of the 349-residue predicted primary translation product of porcine connective tissue growth factor (pCTGF). 10-kDa CTGF-mediated fibroblast DNA synthesis was modulated by exogenous heparin, and CTGF-immunoreactive proteins of 10, 16, and 20 kDa were present in unfractionated uterine luminal flushings. These data reveal the identity of a novel growth factor in uterine fluids as a highly truncated form of CTGF and show that the N-terminal two-thirds of the CTGF primary translation product is not required for mitogenic activity or heparin binding.
Brigstock
D R
DR
Department of Surgery, Cellular, and Developmental Biology Program, Ohio State University and Children's Hospital, Columbus, Ohio 43205, USA.
Steffen
C L
CL
Kim
G Y
GY
Vegunta
R K
RK
Diehl
J R
JR
Harding
P A
PA
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid; D055513:Connective Tissue Growth Factor; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006133:Growth Substances / Q000302:isolation & purification*; D006493:Heparin / Q000378:metabolism*; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D008934:Mitogens / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013552:Swine; D014599:Uterus / Q000378:metabolism*
D006133:Growth Substances; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D008934:Mitogens; D055513:Connective Tissue Growth Factor; D006493:Heparin