Preproinsulin mRNA engineering and its application to the regulation of insulin secretion from human hepatomas

FEBS Lett. 2003 Feb 27;537(1-3):193-7. doi: 10.1016/s0014-5793(03)00121-2.

Abstract

Cell-based therapies for treating insulin-dependent diabetes (IDD) can provide a more physiologic regulation of blood glucose levels in a less invasive fashion than daily insulin injections. Promising cells include non-beta cells genetically engineered to secrete insulin in response to physiologic cues; responsiveness can be introduced at the transcriptional level to regulate preproinsulin (PPI) mRNA biosynthesis. However, these cells exhibit sluggish secretion dynamics, which is not appropriate for achieving euglycemia in higher animals and, eventually, humans. In this work, we have engineered the PPI mRNA so as to destabilize it through nonsense-mediated mRNA decay (NMD). When expressed under transcriptional regulation in HepG2 hepatomas, the engineered PPI mRNA level and of the insulin secretion rate declined faster upon switching off transcription, compared to the one-copy non-engineered control. Our work provides a simple and straightforward method to improve the dynamics of transcriptionally regulated insulin secretion, which can be a useful tool in developing cell-based therapies for IDD.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Base Sequence
  • Carcinoma, Hepatocellular
  • DNA Primers
  • Gene Expression Regulation, Neoplastic*
  • Genes, Reporter
  • Genetic Engineering
  • Genetic Vectors
  • Humans
  • Insulin / genetics*
  • Insulin / metabolism*
  • Insulin Secretion
  • Liver Neoplasms
  • Luciferases / genetics
  • Plasmids
  • Polymerase Chain Reaction
  • Proinsulin / genetics*
  • Protein Precursors / genetics*
  • RNA, Messenger / genetics*
  • Recombinant Fusion Proteins / metabolism
  • Transfection
  • Tumor Cells, Cultured

Substances

  • DNA Primers
  • Insulin
  • Protein Precursors
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • preproinsulin
  • Proinsulin
  • Luciferases