Abstract:
:This project examines the transcellular membrane protein polarity of bovine aortic endothelial cell (BAEC) monolayers in vitro with respect to the roles that intercellular junctions (as defined by comparing confluent and subconfluent monolayers) and the submembranous cytoskeleton play in controlling this phenomenon. Plasma membrane (PM) proteins obtained from apical (AP) and basolateral (BL) PM domains of confluent BAEC monolayers were isolated using the cationic colloidal silica technique and resolved by two-dimensional gel electrophoresis (2-D PAGE). To facilitate the identification of domain-specific PM proteins, an isoelectric point/molecular weight database of the proteins from AP and BL PM domains was constructed. Domain-specific PM proteins were assessed for their interaction with the cytoskeleton by determining whether they co-isolated with a Triton X-100 detergent-resistant cytoskeletal/extracellular matrix fraction. The maintenance of polarized PM protein segregation by intercellular junctional complexes was determined by comparing AP and BL protein patterns of confluent monolayers with patterns generated by subconfluent monolayers, which lack such junctional structures. Proteins isolated from AP and BL PM domains from both confluent states were immunoblotted with antibodies to angiotensin-converting enzyme (ACE) and collagen receptors (CR). ACE was restricted exclusively to the AP PM domain in the subconfluent condition, even though no apparent cytoskeletal interaction was observed. CRs, found to interact with the cytoskeleton in either confluence state, were predominantly segregated to the BL PM domain regardless of the presence or absence of cell-cell contact. Membrane proteins found by 2-D PAGE to be asymmetrically distributed in the absence of intercellular junctions were assessed for cytoskeletal interaction by their inability to be extracted by Triton X-100 from monolayers in the subconfluent state. Computer cross-referencing of 2-D PAGE peak lists and immunodetection generated from the above fractionation protocols identifies a set of four proteins associated with the cytoskeleton that remain segregated in the proper domain, and five proteins associated with the cytoskeleton that become equally distributed between AP and BL PM domains in the absence of intercellular junctions. Additionally, six proteins not associated with the cytoskeleton remain asymmetrically distributed to the AP domain in the subconfluent state. The data suggest that BAEC monolayers have unknown mechanisms, apart from intercellular junctions expressed at confluency or cytoskeletal binding, for maintaining transcellular PM protein polarity.
journal_name
J Cell Scijournal_title
Journal of cell scienceauthors
Stolz DB,Bannish G,Jacobson BSkeywords:
subject
Has Abstractpub_date
1992-09-01 00:00:00pages
53-68eissn
0021-9533issn
1477-9137journal_volume
103 ( Pt 1)pub_type
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journal_title:Journal of cell science
pub_type: 杂志文章
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journal_title:Journal of cell science
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journal_title:Journal of cell science
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journal_title:Journal of cell science
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:10.1242/jcs.223974
更新日期:2019-05-31 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1996-10-01 00:00:00
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pub_type: 杂志文章
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更新日期:2005-06-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:10.1242/jcs.03035
更新日期:2006-07-15 00:00:00
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journal_title:Journal of cell science
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更新日期:2001-05-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:10.1242/jcs.01075
更新日期:2004-05-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:10.1242/jcs.195552
更新日期:2017-03-15 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1993-05-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1975-08-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1981-10-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1982-08-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章,评审
doi:10.1242/jcs.093575
更新日期:2014-05-15 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1997-03-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1992-04-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1994-10-01 00:00:00
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journal_title:Journal of cell science
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doi:
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1999-06-01 00:00:00
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journal_title:Journal of cell science
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更新日期:2013-03-15 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
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更新日期:2012-09-01 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
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更新日期:2003-08-15 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
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更新日期:2003-10-15 00:00:00
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journal_title:Journal of cell science
pub_type: 杂志文章
doi:
更新日期:1994-03-01 00:00:00
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journal_title:Journal of cell science
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更新日期:2003-02-15 00:00:00