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Distribution and biochemical characterization of the INO antigen during chick neural crest cell migration

Lallier, Thomas and Artinger, Michael and Matthew, William and Bronner-Fraser, Marianne (1990) Distribution and biochemical characterization of the INO antigen during chick neural crest cell migration. In: 15th Seiriken Conference on the Molecular and Cellular Mechanisms of Neural Development and Reorganization. Neuroscience Research Supplement. No.Suppl. 13. Elsevier Ireland Ltd , Ireland, S126-S140. http://resolver.caltech.edu/CaltechAUTHORS:20160308-095624786

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Abstract

The INO (inhibitor of neurite outgrowth) antibody recognizes a laminin-heparan sulfate proteoglycan complex and was isolated for its ability to functionally inhibit axonal outgrowth of peripheral neurons. Here, we examine the distribution and biochemical characteristics of INO in the early chick embryo. Because the INO antigen is sensitive to most classical fixation procedures and fixation leads to abundant nuclear staining, the antibody was directly injected into 1.5–2.5-day-old embryos prior to fixation. The distribution of the injected antibody was then observed in cryostat sections by indirect immunofluorescence. Particular attention was focussed upon regions of ongoing neural crest cell migration. The INO antigen was observed along both cranial and trunk neural crest cell migratory pathways. The antigen was seen around the basement membrane surrounding the neural tube and notochord, and underneath the ectoderm and endoderm. In addition, fibrillar staining was observed in the cranial mesenchyme and in both rostral and caudal halves of the somitic sclerotome in the trunk. The distribution pattern was identical to that previously observed for laminin or heparan sulfate proteoglycan. To confirm the nature of the INO antigen, we performed immunoprecipitations of chick embryos ranging from 1.5 to 9 days of incubation. Half of each sample was digested with heparinase prior to SDS-PAGE and silver staining. In material from young embryos, bands of 200 and 180 kD (probably corresponding to the B-chains of laminin) plus two broad smears of bands at 180−150 kD and 130−85 kD were observed without heparinase digestion. Following enzymatic digestion, the 200-kD and 180-kD bands remained, while the smears disappeared and were replaced by numerous low-molecular-weight bands. In contrast to preparations from young embryos, samples taken from embryos at day 3 or beyond did not enter the 8% gel without heparinase digestion, though the banding pattern appeared identical to younger samples after heparinase digestion in the presence or absence of Ca^(2+). This change in the INO antigen with age could result from an increase in the heparin-side-chains attached to similar core proteins, or from an increase in the stability of the laminin-heparan sulfate proteoglycan containing complex with time.


Item Type:Book Section
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/0921-8696(90)90041-ZDOIArticle
http://www.sciencedirect.com/science/article/pii/092186969090041ZPublisherArticle
ORCID:
AuthorORCID
Bronner-Fraser, Marianne0000-0003-4274-1862
Additional Information:© 1990 Elsevier Scientific Publishers Ireland Ltd. This work was supported by USPHS Grant HD15527 and a grant from the Muscular Dystrophy Association to MB-F. MB-F is a Sloan Foundation Fellow.
Funders:
Funding AgencyGrant Number
NIHHD15527
Muscular Dystrophy AssociationUNSPECIFIED
Alfred P. Sloan FoundationUNSPECIFIED
Subject Keywords:Laminin-heparan sulfate proteoglycan; Extracellular matrix; Neurite outgrowth
Record Number:CaltechAUTHORS:20160308-095624786
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20160308-095624786
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:65188
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:14 Mar 2016 21:49
Last Modified:14 Mar 2016 21:49

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