Two nuclear proteins in tracheal epithelial cells are recognized by antibodies specific to a squamous differentiation marker, sprI.
In cell-free translations of RNA from primary cultures of pig trachea surface epithelial cells we observed that a mRNA encoding a 20 kDa proline-rich protein (sPRP) was dramatically induced during culturing (Tesfaigzi et al., 1990, Biochem. Biophys. Res. Commun., 172:M1304-1309). This mRNA was not detected in tracheal tissue or in epithelial cells prior to culturing. Antisera were raised to synthetic peptide sequences corresponding to 23 amino acids on the C-terminus (C23-antiserum) and 29 amino acids on the N-terminus (N29 antiserum) of sPRP. On Western blot analysis, C23 antiserum reacted with a 20 kDa protein in cytosolic extracts from pig tracheal cells maintained in culture for 4 days. The reaction with the 20 kDa protein was inhibited by adding C23 peptide. Two nuclear proteins (66 and 70 kDa) obtained by micrococcal nuclease treatment of tracheal cell nuclei were detected on Western blots with C23 antiserum. These proteins were present in cells both before and after culturing. Sucrose gradient fractionation indicated that these nuclear proteins are associated with chromatin. Small amounts of the 66 and 70 kDa proteins were obtained from nuclear matrix fractions. These nuclear proteins also reacted with N29 antiserum. Since these proteins share similar epitopes with the N- and C-termini of sPRP, it is likely that the 20 kDa protein (sPRP) is part of these proteins. However, purification of the nuclear proteins followed by an amino acid sequence analysis is necessary to clarify whether sPRP is part of these proteins.
Tesfaigzi
J
J
Section of Molecular and Cellular Biology, University of California, Davis 95616, USA.
An
G
G
Wu
R
R
Carlson
D M
DM
D000595:Amino Acid Sequence; D000818:Animals; D002454:Cell Differentiation; D003600:Cytosol / Q000737:chemistry; D004848:Epithelium / Q000276:immunology; D005455:Fluorescent Antibody Technique; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015530:Nuclear Matrix / Q000737:chemistry; D009687:Nuclear Proteins / Q000276:immunology*; D010455:Peptides / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D055232:Proline-Rich Protein Domains; D013552:Swine; D014132:Trachea / Q000276:immunology*
D009687:Nuclear Proteins; D010455:Peptides