Effect of fasting on insulin-like growth factor (IGF)-IA and IGF-IB messenger ribonucleic acids and prehormones in rat liver.
The insulin-like growth factor I (IGF-I) gene generates by alternative splicing two IGF-I messenger RNAs (mRNAs) coding for IGF-I prehormones with different E domain sequences. In rats, these two mRNAs differ by the presence (IGF-IB) or absence (IGF-IA) of a 52-bp insert in the E domain coding region. The purpose of this study was to investigate the effect of nutritional perturbation on IGF-IA and -IB expression in rat liver. Northern blot analysis of liver mRNA revealed that the 1.5-1.9 kb and 0.9-1.2 kb IGF-I mRNA species were decreased in rats fasted for 48 h compared with either fasted-refed (48 h of each) or control-fed rats (each, P < 0.01), whereas the 7.5 kb IGF-I mRNA was decreased only when compared with the fasted-refed animals. Using semiquantitative RT-PCR, the IGF-IA transcript (114 bp amplicon) was not altered, whereas the IGF-IB transcript (166 bp amplicon) was decreased in fasted rats compared with the other two groups (both P < 0.01). We confirmed the RT-PCR results by RNase protection assay (RPA), observing that the IGF-IA (224 and 100 bases protected) was not decreased and that the IGF-IB transcript (376 bases protected), accounting for only 23% of the total IGF-I transcripts of control fed rats, was decreased by fasting. Because the results from RT-PCR and RPA do not necessarily predict full-length translatable mRNA, we subjected hepatic IGF-I transcripts to in vitro translation, and we immunoprecipitated IGF-IA and -IB prehormones. Both prehormones were translated principally from exon 1-containing mRNAs, with molecular weights of about 17K and 18K, representing 80% and 20% of the total IGF-I prehormones observed in control fed rats, respectively. Both peptides were reduced in fasted rats compared with controls (P < 0.01), and refeeding restored both. By immunoblotting of the protein extract from liver of fasted rats, IGF-IA was decreased by 77% compared with control-fed animals. Refeeding returned IGF-IA to normal. The lack of reduction of IGF-IA transcript at the alternative splice site suggests that posttranscriptional mechanisms are responsible for the reduction in steady-state IGF-I mRNAs that occurs during fasting. Additionally, we present evidence that biosynthesis of IGF-IA and -IB prehormones by liver is impaired at a posttranscriptional level.
Zhang
J
J
Department of Pediatrics, The University of North Carolina at Chapel Hill, 27599, USA.
Whitehead
R E
RE
Underwood
L E
LE
D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D017931:DNA Primers / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D005091:Exons; D005215:Fasting / Q000502:physiology*; D006128:Growth / Q000502:physiology; D007334:Insulin-Like Growth Factor I / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D008099:Liver / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D008297:Male; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011498:Protein Precursors / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014158:Transcription, Genetic
D017931:DNA Primers; D011498:Protein Precursors; D012333:RNA, Messenger; D007334:Insulin-Like Growth Factor I