Isolation and characterization of cDNA clones encoding pig gastric mucin

Polyclonal antibodies raised to deglycosylated pig gastric mucin were used to screen a cDNA library constructed with pig stomach mucosal mRNA. Immunocytochemistry indicated that the antibody recognizes intracellular and secreted mucin in surface mucous cells of pig gastric epithelium. A total of 70...

Full description

Saved in:
Bibliographic Details
Published in:Biochemical journal 1995-05, Vol.308 (1), p.89-96
Main Authors: Turner, B.S, Bhaskar, K.R, Hadzopoulou-Cladaras, M, Specian, R.D, Lamont, J.T
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
antibodies
Base Sequence
cloning
Cloning, Molecular
complementary DNA
DNA, Complementary - genetics
Fluorescent Antibody Technique
Gastric Mucosa - chemistry
genbank/u10281
genbank/u12768
Gene Expression
Genes
In Situ Hybridization
isolation
Molecular Sequence Data
mucins
Mucins - genetics
nucleotide sequences
Repetitive Sequences, Nucleic Acid
RNA, Messenger - genetics
Sequence Alignment
stomach
Swine
Citations: Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Polyclonal antibodies raised to deglycosylated pig gastric mucin were used to screen a cDNA library constructed with pig stomach mucosal mRNA. Immunocytochemistry indicated that the antibody recognizes intracellular and secreted mucin in surface mucous cells of pig gastric epithelium. A total of 70 clones producing proteins immunoreactive to this antibody were identified, two of which (PGM-2A,9B) were fully sequenced from both ends. Clone PGM-9B hybridized to a polydisperse mRNA (3-9 kb) from pig stomach, but not liver, intestine or spleen, nor to mRNA from human, mouse, rabbit or rat stomach. Sequence analysis indicated that PGM-9B encodes 33 tandem repeats of a 16-amino-acid consensus sequence rich in serine (46%) and threonine (17%). Using the restriction enzyme MwoI, which has a single target site in the repeat, it was demonstrated that PGM-9B consists entirely of this tandem repeat. Southern-blot analysis indicated that the repeat region is contained in a 20 kb HindIII-EcoRI fragment, and BamHI digestion suggested that most of the repeats are contained in a 10 kb fragment. In situ hybridization with an antisense probe to PGM-9B showed an intense signal in the entire gastric gland. Clone PGM-2A also contains the same repeat sequence as 9B, but, in addition, has a 64-amino-acid-long non-repeat region at its 5' end. Interestingly the nonrepeat region of PGM-2A has five cysteine residues, the arrangement of which is identical with that reported for human intestinal mucin gene MUC2.
ISSN:0264-6021
1470-8728
DOI:10.1042/bj3080089