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 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 non-repeat region of PGM-2A has five cysteine residues, the arrangement of which is identical with that reported for human intestinal mucin gene MUC2.
Turner
B S
BS
Evans Department of Clinical Research, Boston University Medical Center, MA 02118, USA.
Bhaskar
K R
KR
Hadzopoulou-Cladaras
M
M
Specian
R D
RD
LaMont
J T
JT
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005455:Fluorescent Antibody Technique; D005753:Gastric Mucosa / Q000737:chemistry; D015870:Gene Expression; D005796:Genes; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D009077:Mucins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D016415:Sequence Alignment; D013552:Swine
D018076:DNA, Complementary; D009077:Mucins; D012333:RNA, Messenger