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MicroRNA In situ Hybridization for Formalin Fixed Kidney Tissues
Published on: November 30, 2013
Androgen-regulated ornithine decarboxylase mRNAs of mouse kidney
Abstract:
Ornithine decarboxylase, the first enzyme of the polyamine biosynthetic pathway, is induced by androgens in the mouse kidney. We have isolated from a kidney cDNA clone bank two plasmids, pODC1440 and pODC934 , that contain different cDNA inserts corresponding to ornithine decarboxylase mRNA. Identification was based upon the ability of plasmid-specific mRNAs to encode a 53,000-dalton polypeptide that reacts with antibody to purified mouse kidney ornithine decarboxylase. Plasmid pODC1440 hybridizes predominantly to a mRNA that is 2.1 kilobases (kb) long and is induced about 20-fold in the kidneys of female mice treated with testosterone. Plasmid pODC934 hybridizes to mRNAs of lengths 2.2 and 2.6 kb, which are induced about 8-fold by testosterone. There are probably no more than 1-2 copies of the pODC1440 -specific sequence in the mouse genome, while there may be as many as 12 copies of the pODC934 -specific sequence. That the two plasmids correspond to different ornithine decarboxylase mRNAs is suggested by two observations. First, several mouse strains express normal levels of pODC934 -specific RNA and little or no pODC1440 -specific RNA; furthermore, pODC934 -specific RNA is expressed in several tissues while pODC1440 -specific RNA is kidney-specific. Thus, androgen-mediated stimulation of kidney ornithine decarboxylase activity levels involves alterations in the concentrations of at least two distinct mRNAs.
Insights
Androgens stimulate kidney ornithine decarboxylase (ODC) activity by altering at least two distinct ODC messenger RNA (mRNA) concentrations. Researchers identified two ODC mRNA types, differing in size and gene copy number.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Ornithine decarboxylase (ODC) is the initial enzyme in polyamine synthesis.
- Androgens induce ODC in the mouse kidney, suggesting hormonal regulation of polyamine metabolism.
Purpose of the Study:
- To isolate and characterize cDNA clones for mouse kidney ornithine decarboxylase mRNA.
- To investigate the specific ornithine decarboxylase mRNAs regulated by androgens.
Main Methods:
- Isolation of cDNA clones (pODC1440 and pODC934) from a mouse kidney library.
- Northern blot analysis to detect mRNA sizes and androgen-induced changes.
- Quantification of gene copy numbers and tissue-specific expression.
Main Results:
- Two distinct ODC mRNAs (2.1 kb and 2.2/2.6 kb) were identified, corresponding to pODC1440 and pODC934, respectively.
- Testosterone treatment significantly increased the 2.1 kb mRNA (20-fold) and moderately increased the 2.2/2.6 kb mRNAs (8-fold).
- pODC1440 sequence is kidney-specific with low gene copy number, while pODC934 sequence is found in multiple tissues and has a higher gene copy number.
Conclusions:
- Androgen stimulation of kidney ODC activity involves at least two different ODC mRNA species.
- Differential regulation and expression patterns suggest distinct roles for these ODC mRNA variants.
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