Apobec-1 transcription in rat colon cancer: decreased apobec-1 protein production through alterations in polysome distribution and mRNA translation associated with upstream AUGs.
Apobec-1 catalyzes C to U editing of apolipoprotein B (apoB) mRNA in the mammalian intestine. Rat apobec-1 is transcribed from three distinct promoters, which contain distinct 5' untranslated regions (5'UTRs) accompanied by variable numbers of in-frame upstream AUGs (uAUGs). We have observed a shift in apobec-1 promoter usage in an experimental model of colon carcinogenesis, resulting in transcripts loaded with 5'AUGs. In colon cancer, apobec-1 protein levels decreased by 90% in the cancer tissue as compared to normal tissue, suggesting an inhibitory effect of the 5'UTR on apobec-1 translation. We investigated the effects of these different 5'UTRs by site-directed mutagenesis coupled with in vitro translation studies. These studies established that the uAUGs within the 5'UTR of the alternative transcripts inhibit apobec-1 translation. This effect was independent of the length of the 5'UTR. Further analysis demonstrated that these uAUGs altered the polysome distribution, shifting the mRNA towards a denser, post-polyribosomal fraction. These findings were confirmed in transient transfection studies in vivo using HepG2 cells, where functional expression of apobec-1 was restored by mutagenesis of the uAUGs. Taken together, these data imply that rat apobec-1 gene expression is downregulated through alternative promoter usage. This dominant translational control of apobec-1 gene expression is most plausibly exerted through uAUGs.
Anant
Shrikant
S
Division of Gastroenterology, Department of Internal Medicine, Washington University School of Medicine, Campus Box 8124, 660 South Euclid Avenue, St. Louis, MO 63110, USA. sanant@im.wustl.edu
Mukhopadhyay
Debnath
D
Hirano
Ken-Ichi
K
Brasitus
Thomas A
TA
Davidson
Nicholas O
NO
D020121:5' Untranslated Regions / Q000235:genetics; D000071479:APOBEC-1 Deaminase; D017398:Alternative Splicing / Q000235:genetics; D000818:Animals; D018387:Codon, Initiator / Q000235:genetics* / Q000502:physiology; D003110:Colonic Neoplasms / Q000235:genetics* / Q000378:metabolism; D003564:Cytidine Deaminase / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics* / Q000502:physiology; D008297:Male; D011132:Polyribosomes / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D051381:Rats; D011916:Rats, Inbred F344; D014158:Transcription, Genetic
D020121:5' Untranslated Regions; D018387:Codon, Initiator; D000071479:APOBEC-1 Deaminase; C000605091:Apobec1 protein, rat; D003564:Cytidine Deaminase