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Molecular cloning and sequence analysis of the human ribosomal protein S16
S K Batra1, R S Metzgar, M A Hollingsworth
1Department of Microbiology and Immunology, Duke University Medical Center, Durham, North Carolina 27710.
Abstract:
A cDNA library from a poorly differentiated human pancreatic tumor cell line was screened for differentially expressed mRNAs using single-stranded cDNA probes synthesized from poly(A+) RNA of the poorly differentiated cell line Panc 1 and a very well differentiated cell line CD11. One of the cDNA clones isolated hybridized to a transcript size of 650 base pairs on Northern blot analysis and showed 30-fold higher expression in the poorly differentiated cell line as compared with the well differentiated cell line. Sequence analysis of this cDNA clone and its deduced amino acid sequence showed an open reading frame of 441 nucleotides with 100 and 98.6% homology to ribosomal protein S16 (rpS16) from rat and mouse, respectively. Northern blot analyses with a panel of 14 pancreatic cell lines, 2 breast cell lines, 2 colon cell lines, and several other tissues showed higher expression of rpS16 only in the poorly differentiated pancreatic tumor cell line Panc 1. The expression of mRNA for two other ribosomal proteins, rpL30 and rpL32, were not elevated in Panc 1. Southern blot analysis of genomic DNA showed a 20-fold amplification of a single band among the rpS16 family only in the Panc 1 cell line.
Insights
This study identified ribosomal protein S16 (rpS16) as highly expressed in poorly differentiated pancreatic cancer cells. Gene amplification in rpS16 may drive pancreatic tumor progression.
Area of Science:
- Molecular Biology
- Oncology
- Genetics
Background:
- Pancreatic cancer is often diagnosed at late stages, necessitating research into its molecular underpinnings.
- Understanding gene expression differences between well-differentiated and poorly differentiated tumors is crucial for identifying therapeutic targets.
Purpose of the Study:
- To identify differentially expressed genes in poorly differentiated human pancreatic cancer cells.
- To investigate the role of ribosomal protein S16 (rpS16) in pancreatic tumor progression.
Main Methods:
- Differential screening of a cDNA library from pancreatic tumor cell lines (Panc 1 and CD11).
- Northern blot analysis to assess mRNA expression levels.
- Sequence analysis to identify homologous genes.
- Southern blot analysis to detect gene amplification.
Main Results:
- A 650-base pair cDNA clone showed 30-fold higher expression in poorly differentiated Panc 1 cells compared to well-differentiated CD11 cells.
- Sequence analysis revealed the clone to be ribosomal protein S16 (rpS16), with high homology to rat and mouse rpS16.
- Elevated rpS16 mRNA expression was specific to the poorly differentiated Panc 1 cell line among various tested cell lines and tissues.
- Southern blot analysis indicated a 20-fold amplification of rpS16 in the Panc 1 cell line, unlike other ribosomal proteins (rpL30, rpL32).
Conclusions:
- Ribosomal protein S16 (rpS16) is significantly overexpressed in poorly differentiated human pancreatic cancer cells.
- The observed rpS16 gene amplification in Panc 1 cells suggests its potential role in the progression of pancreatic cancer.