Use of LiCl in phospholipase C assays masks the impaired functionality of a mutant angiotensin II receptor

Cell Signal. 1997 Aug;9(5):379-82. doi: 10.1016/s0898-6568(97)00032-6.

Abstract

We recently reported that replacement of Tyr302 for Ala in the human angiotensin II type 1 receptor (hAT1) severely impaired its ability to activate phospholipase C (PLC). Another study demonstrated that the same mutation in the rat AT1 receptor only slightly impaired its ability to activate PLC. The most striking difference between the two studies was the use of LiCl in the experimental conditions. Thus, in the present report we evaluated the effect of LiCl on the rate of accumulation of inositol trisphosphate (IP3) in transfected cells stimulated with angiotension II (Ang II). In the presence of LiCl, Ang II caused a significant accumulation of IP3 in COS-7 cells transfected with the hAT1Y302A mutant receptor. In stably expressing CHO cells, stimulation of hAT1Y302A did not induce any IP3 elevation even in the presence of LiCl whereas the hAT1 wild-type receptor increased the production of IP3 exclusively in the presence of LiCl. These results show that LiCl is a convenient tool to enhance the sensitivity of PLC assays. However, in structure-activity relationship studies, it may underestimate or mask the debilitating effect of some mutations.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Angiotensin II / metabolism
  • Angiotensin II / pharmacology
  • Animals
  • CHO Cells
  • COS Cells
  • Cricetinae
  • Enzyme Activation
  • Inositol 1,4,5-Trisphosphate / analogs & derivatives
  • Inositol 1,4,5-Trisphosphate / metabolism
  • Lithium Chloride / metabolism
  • Lithium Chloride / pharmacology*
  • Mutagenesis, Site-Directed
  • Organothiophosphorus Compounds / metabolism*
  • Receptor, Angiotensin, Type 1
  • Receptors, Angiotensin / metabolism*
  • Transfection
  • Type C Phospholipases / metabolism*

Substances

  • Organothiophosphorus Compounds
  • Receptor, Angiotensin, Type 1
  • Receptors, Angiotensin
  • Angiotensin II
  • inositol 1,4,5-triphosphorothioate
  • Inositol 1,4,5-Trisphosphate
  • Type C Phospholipases
  • Lithium Chloride