Differential regulation of trypsinogen mRNA translation: full-length mRNA sequences encoding two oppositely charged trypsinogen isoenzymes in the dog pancreas.
In the absence of changes in functional mRNA levels, stimulation of the pancreas with caerulein, a peptide analog of cholecystokinin, has been previously shown to increase the synthesis of anionic but not cationic trypsinogen. To look for structure-function correlations, a high-yield, full-length cDNA library has been constructed from canine pancreatic poly(A)+ mRNA. Full-length clones coding for the two major trypsinogen isoenzyme forms have been identified by colony hybridization and verified by in vitro translation of hybrid-selected mRNA in the presence of microsomal membranes and an optimal redox potential. Disulfide-bonded translation products were separated and identified by two-dimensional isoelectric focusing-sodium dodecyl sulfate-gel electrophoresis. Nucleotide sequence analysis allowed us to deduce the amino acid sequences for the anionic and cationic forms of canine trypsinogen, which contain 232 and 231 residues, respectively (77% amino acid identity), and the 15-residue amino terminal signal sequences (53% amino acid identity) associated with the two presecretory forms. Measurements of relative and absolute mRNA levels, when related to relative protein synthesis values, indicated that the translational efficiency of anionic trypsinogen mRNA exceeded that of cationic trypsinogen mRNA by 1.5- to 2.9-fold under basal conditions. Analysis of the 5' noncoding regions of trypsinogen mRNAs revealed a striking conservation of sequence (10 of 12 bases) between dog and rat anionic trypsinogen forms. This contrasted markedly with the divergence of the 5' noncoding regions observed between dog anionic and cationic trypsinogen mRNAs.
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D004285:Dogs; D005786:Gene Expression Regulation; D007526:Isoelectric Point; D007527:Isoenzymes / Q000235:genetics*; D009690:Nucleic Acid Conformation; D010179:Pancreas / Q000201:enzymology / Q000502:physiology; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D014362:Trypsinogen / Q000235:genetics*
D007527:Isoenzymes; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D014362:Trypsinogen; D004247:DNA