Jk. Xu et al., Cloning, sequencing, and functional characterization of the rat homologue of receptor activator of NF-kappa B ligand, J BONE MIN, 15(11), 2000, pp. 2178-2186
A complementary DNA (cDNA) encoding the rat homologue of receptor activator
of NF-kappaB ligand/osteoprotegerin ligand/osteoclast differentiation fact
or/tumor necrosis factor (TNF)-related activation-induced cytokine (RANKL/O
PGL/ODF/TRANCE) was cloned and sequenced from tibias of ovariectomized (OVX
) rats. The predicted amino acid sequence of rat RANKL (rRANKL) has 84% and
96% identity to that of human and mouse RANKL, respectively, and 35% and 3
7% similarity to that of human and mouse TNF-related apoptosis-inducing lig
and (TRAIL), respectively. RANKL transcripts were expressed abundantly in t
he thymus and bone tissues of OVX rats. rRANKL has a single hydrophobic reg
ion between residues 53 and 69, which is most likely to serve as a transmem
brane domain, The long C-terminal region containing beta -sheet-forming seq
uences of the TNF-like core is considered the extracellular region. Three t
runcated domains within the TNF-like core region were expressed as glutathi
one S-transferase (GST) fusion proteins and investigated for their ability
to induce osteoclastogenesis, The results showed that GST-rRANKL, (aa160-31
8) containing the full TNF-like core region had the highest capability to i
nduce the formation of osteoclast-like cells from RAW(264.7) cells, GST-rRA
NKL (aa239-318 and aa160-268) had lesser degrees of osteoclast inductivity,
Furthermore, the GST-rRANKL (aa160-318) is capable of (1) inducing osteocl
ast formation from rat spleen cells in the presence of macrophage colony-st
imulating factor (M-CSF), (2) stimulating mature rat osteoclast polarizatio
n and bone resorption ex vivo, and (3) inducing systemic hypercalcemia in v
ivo; thus the full TNF-like core region of rRANKL is an important regulator
of calcium homeostasis and osteoclastic function.