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Post-translational processing of chicken bone phosphoproteins. Identification of the bone phosphoproteins of embryonic tibia.

ABSTRACT: In order to understand the mechanism of the post-translational processing of bone phosphoproteins in embryonic bone, periosteal bone strips isolated from 12-day-embryonic-chick tibiae were cultured and the bone proteins labelled with Na2H32PO4. Of the total radiolabelled proteins recovered from the medium and bone extracts in the absence of SDS ('medium', 'EDTA extract' and 'EDTA/guanidinium chloride extract'), nearly 80% of the radioactivity was found in the EDTA extract. The three major radiolabelled phosphoproteins in the EDTA extract of apparent Mr 68,000, 63,000 and 58,000 reacted with polyclonal as well as monoclonal antibodies raised against '32-kDa' and '150-kDa' bone phosphoproteins which were derived from 14-week-old chicken. Therefore these phosphorylated embryonic proteins are identified as chicken bone phosphoproteins. Judging from their common N-terminal sequences, differences in the patterns obtained by labelling them with several radioisotopes, and slightly different amino acid compositions, these components seem to have been derived from the same original protein by sequential proteolytic cleavage and other processing such as glycosylation and phosphorylation.

SUBMITTER: Mikuni-Takagaki Y

PROVIDER: S-EPMC1131478 | biostudies-other | 1990 Jun

REPOSITORIES: biostudies-other

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Project description:Although it is known that mechanical forces are needed for normal bone development, the current understanding of how biophysical stimuli are interpreted by and integrated with genetic regulatory mechanisms is limited. Mechanical forces are thought to be mediated in cells by "mechanosensitive" genes, but it is a challenge to demonstrate that the genetic regulation of the biological system is dependant on particular mechanical forces in vivo. We propose a new means of selecting candidate mechanosensitive genes by comparing in vivo gene expression patterns with patterns of biophysical stimuli, computed using finite element analysis. In this study, finite element analyses of the avian embryonic limb were performed using anatomically realistic rudiment and muscle morphologies, and patterns of biophysical stimuli were compared with the expression patterns of four candidate mechanosensitive genes integral to bone development. The expression patterns of two genes, Collagen X (ColX) and Indian hedgehog (Ihh), were shown to colocalise with biophysical stimuli induced by embryonic muscle contractions, identifying them as potentially being involved in the mechanoregulation of bone formation. An altered mechanical environment was induced in the embryonic chick, where a neuromuscular blocking agent was administered in ovo to modify skeletal muscle contractions. Finite element analyses predicted dramatic changes in levels and patterns of biophysical stimuli, and a number of immobilised specimens exhibited differences in ColX and Ihh expression. The results obtained indicate that computationally derived patterns of biophysical stimuli can be used to inform a directed search for genes that may play a mechanoregulatory role in particular in vivo events or processes. Furthermore, the experimental data demonstrate that ColX and Ihh are involved in mechanoregulatory pathways and may be key mediators in translating information from the mechanical environment to the molecular regulation of bone formation in the embryo.

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Post-translational processing of chicken bone phosphoproteins. Identification of the bone phosphoproteins of embryonic tibia.

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