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Sequence of rat liver alpha 2-macroglobulin and acute phase control of its messenger RNA
Abstract:
Six alpha 2-macroglobulin cDNA clones were isolated from two liver cDNA libraries produced from rats undergoing acute inflammation. The coding sequence for rat alpha 2-macroglobulin including its 27-residue signal peptide and the 3' - and part of the 5' nontranslated regions were determined. The mature protein consisting of 1445 amino acids is coded for by a 4790 +/- 40 nucleotide messenger RNA. It contains a typical internal thiol ester region and 25 cysteine residues which are conserved between rat and human alpha 2-macroglobulin. Although the amino acid sequences of rat and human alpha 2-macroglobulin share 73% identity, two small divergent areas of 17 and 38 residues were found, corresponding to 29 and 11% identity, respectively. These areas are located in the bait region and, therefore, may confer specific proteinase recognition capabilities on rat alpha 2-macroglobulin. Following an inflammatory stimulation, rat alpha 2-macroglobulin mRNA levels increased 214-fold over control values and reached a maximum at 18 h. By 24 h the levels had decreased to less than 30% of the maximum value. Transcription rates from the alpha 2-macroglobulin gene as measured in nuclear run-on experiments showed a less than 3-fold increase in nuclei from acutely inflamed rats as compared to controls. These results suggest that the accummulation of alpha 2M mRNA is due to the combined effects of increased transcription rates and post-transcriptional processing.
Insights
Researchers characterized rat alpha 2-macroglobulin (a2M) mRNA during inflammation. Findings suggest increased transcription and post-transcriptional processing drive a2M mRNA accumulation, impacting inflammatory responses.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Alpha 2-macroglobulin (a2M) is a key plasma protein involved in regulating proteinase activity.
- Understanding a2M gene regulation during inflammation is crucial for comprehending its physiological role.
- Previous studies have not fully elucidated the mechanisms controlling a2M mRNA levels in inflammatory states.
Purpose of the Study:
- To determine the complete coding sequence of rat alpha 2-macroglobulin (a2M).
- To investigate the regulation of a2M gene expression and mRNA accumulation during acute inflammation in rats.
- To compare structural features of rat a2M with its human counterpart.
Main Methods:
- Isolation and sequencing of alpha 2-macroglobulin cDNA clones from rat liver cDNA libraries.
- Determination of the full-length coding sequence, signal peptide, and untranslated regions of rat a2M.
- Quantification of a2M mRNA levels and gene transcription rates using Northern blot analysis and nuclear run-on assays.
Main Results:
- The complete nucleotide sequence of rat a2M mRNA (4790 nucleotides) and its 1445-amino acid mature protein were determined.
- Rat and human a2M share 73% amino acid identity, with distinct divergent regions in the bait region potentially conferring specific proteinase recognition.
- Following acute inflammation, rat a2M mRNA levels increased 214-fold, while transcription rates showed a modest increase, indicating significant post-transcriptional regulation.
Conclusions:
- The study provides a comprehensive characterization of rat alpha 2-macroglobulin and its gene.
- The significant induction of a2M mRNA during inflammation is primarily mediated by post-transcriptional mechanisms, alongside increased transcription.
- These findings highlight the complex regulatory network governing a2M expression in response to inflammatory stimuli.