Mt. Hincke et al., Molecular cloning and ultrastructural localization of the core protein of an eggshell matrix proteoglycan, ovocleidin-116, J BIOL CHEM, 274(46), 1999, pp. 32915-32923
The role of avian eggshell matrix proteins in shell formation is poorly und
erstood. This calcitic biomaterial forms in a uterine fluid where the prote
in composition varies during the initial, calcification, and terminal phase
s of eggshell deposition, A specific antibody was raised to a 116-kDa prote
in, which is most abundant in uterine fluid during active eggshell calcific
ation. This antiserum was used to expression screen a bacteriophage cDNA li
brary prepared using mRNA extracted from pooled uterine tissue harvested at
the midpoint of eggshell calcification. Plasmids containing inserts of dif
fering 5'-lengths were isolated with a maximum cDNA sequence of 2.4 kilobas
es, Northern blotting and reverse transcriptase-polymerase chain reaction d
emonstrated that the 2,35-kilobase message was expressed in a uterine-speci
fic manner,. The hypothetical translational product from the open reading f
rame corresponded to a novel 80-kDa protein, which we have named ovocleidin
-116, After removal of the predicted signal peptide, its N-terminal sequenc
e corresponded almost exactly with that determined from direct microsequenc
ing of the 116-kDa uterine protein (this work) and with that previously det
ermined for the core protein of a 120-kDa egg shell dermatan sulfate proteo
glycan (Corrino, D. A., Rodriguez, J, P,, and Caplan, A. I. (1997) Connect.
Tissue Res. 36, 175-193), Ultrastructural colloidal gold immunocytochemist
ry of ovocleidin-116 demonstrated its presence in the organic matrix, in sm
all vesicles found throughout the mineralized palisade layer, and the calci
um reserve assembly of the mammillary layer. Ovocleidin-116 thus is a candi
date molecule for the regulation of calcite growth during eggshell calcific
ation.