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Molecular cloning of human intestinal mucin cDNAs. Sequence analysis and evidence for genetic polymorphism
1Gastrointestinal Research Laboratory (151M2), Veterans Administration Medical Center, San Francisco, California 94121.
Abstract:
A human small intestine lambda gt11 cDNA library was screened using antisera prepared against the deglycosylated protein backbone of human colon cancer xenograft mucin. Three cDNAs were isolated from this screening, designated SMUC 40-42. These cDNAs were all found to contain tandem repeats of 69 nucleotides which encoded a threonine- and proline-rich protein consensus sequence of PTTTPITTTTTVTPTPTPTGTQT. RNA blots probed with one of these cDNAs, SMUC 41, exhibited large, polydisperse hybridization bands at approximately 7,600 bases. Band intensities were strongest when human small intestine, colon, and colon cancer poly(A)+ RNA was used. In vitro translation of poly(A)+ RNA from human small intestine, colon, and colon cancer cells produced a 162,000-dalton peptide that was immunoprecipitated with antibodies to deglycosylated mucin. SMUC 41 was also used to probe DNA blots, which indicated the presence of restriction fragment length polymorphisms in the intestinal mucin gene. These findings may be important in assessing the abnormal mucins found associated with several human diseases.
Insights
Researchers identified novel human small intestine mucin cDNAs (SMUC 40-42) encoding a proline- and threonine-rich protein. These findings are crucial for understanding abnormal mucins in various human diseases.
Area of Science:
- Molecular Biology
- Gastroenterology
- Biochemistry
Background:
- Mucins are high molecular weight glycoproteins crucial for epithelial barrier function.
- Abnormal mucin expression and structure are associated with various gastrointestinal diseases, including cancer.
- Understanding the genetic basis of mucin production is vital for disease diagnosis and treatment.
Purpose of the Study:
- To isolate and characterize cDNAs encoding human small intestine mucin.
- To investigate the expression patterns of intestinal mucin genes.
- To explore the potential role of mucin gene variations in human diseases.
Main Methods:
- Screening of a human small intestine lambda gt11 cDNA library using antisera against deglycosylated mucin.
- Isolation and characterization of three cDNAs (SMUC 40-42) containing tandem repeats.
- RNA and DNA blot analyses to assess gene expression and identify polymorphisms.
- In vitro translation to determine peptide size and immunoprecipitation.
Main Results:
- Three cDNAs (SMUC 40-42) encoding a threonine- and proline-rich protein (PTTTPITTTTTVTPTPTPTGTQT) were isolated.
- SMUC 41 hybridized to large, polydisperse RNA bands (~7,600 bases) in small intestine, colon, and colon cancer tissues.
- In vitro translation produced a 162,000-dalton peptide recognized by anti-mucin antibodies.
- DNA blot analysis revealed restriction fragment length polymorphisms in the intestinal mucin gene.
Conclusions:
- Novel cDNAs for human small intestine mucin have been identified.
- The identified mucin gene is expressed in the small intestine, colon, and colon cancer tissues.
- The presence of polymorphisms suggests genetic variations in intestinal mucin that may be relevant to human diseases.