Characterization of an ATP-stimulatable Ca(2+)-independent phospholipase A2 from clonal insulin-secreting HIT cells and rat pancreatic islets: a possible molecular component of the beta-cell fuel sensor

Biochemistry. 1994 Jun 14;33(23):7442-52. doi: 10.1021/bi00189a052.


Isolated pancreatic islets from rats and humans express a plasmalogen-preferring ATP-stimulatable, Ca(2+)-independent phospholipase A2 (ASCI-PLA2) enzyme which participates in the glucose-stimulated hydrolysis of arachidonate from membrane phospholipids and in insulin secretion. Here we report that clonal insulin-secreting HIT beta-cells contain substantial amounts of endogenous plasmalogens and express a similar ASCI-PLA2 activity with the following properties: (1) Enzymatic activity as well as glucose-induced eicosanoid release and insulin secretion are inhibited by a mechanism-based suicide substrate directed towards ASCI-PLA2. (2) HIT cell ASCI-PLA2 is selectively activated and protected against thermal denaturation by ATP. (3) The magnitude of ASCI-PLA2 activation by the nonhydrolyzable ATP analog AMP-PCP is similar to that by ATP. (4) The ATP concentrations required to activate ASCI-PLA2 fall within physiologic ranges in the presence of Mg2+. (5) ADP induces a concentration-dependent attenuation of the activation of ASCI-PLA2 by ATP. HIT cell ASCI-PLA2 exhibited an apparent isoelectric point of 7.5 on chromatofocusing analysis and was quantitatively adsorbed to an ATP-agarose matrix and selectively desorbed from this column by ATP. Mono-Q anion-exchange analysis of the active ATP-agarose eluant yielded a peak of ASCI-PLA2 activity associated with a single protein band with an apparent molecular mass of 40 kDa. Similar chromatographic behavior of the rat pancreatic islet ASCI-PLA2 activity was observed during sequential ATP-agarose and Mono-Q anion-exchange steps. These results indicate that HIT cells express an ASCI-PLA2 similar to the analogous islet enzyme and suggest that expression of this enzyme and of its preferred plasmalogen substrates may be a general property of insulin-secreting beta-cells.

Publication types

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

MeSH terms

  • Adenosine Triphosphate / pharmacology*
  • Animals
  • Autoradiography
  • Calcium / metabolism*
  • Cells, Cultured
  • Chromatography, Affinity
  • Chromatography, Ion Exchange
  • Cricetinae
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme Activation
  • Humans
  • Islets of Langerhans / cytology
  • Islets of Langerhans / enzymology*
  • Islets of Langerhans / metabolism
  • Mesocricetus
  • Phospholipases A / antagonists & inhibitors
  • Phospholipases A / isolation & purification
  • Phospholipases A / metabolism*
  • Phospholipases A2
  • Plasmalogens / metabolism
  • Rats
  • Substrate Specificity


  • Plasmalogens
  • Adenosine Triphosphate
  • Phospholipases A
  • Phospholipases A2
  • Calcium