The concentration of P-enolpyruvate carboxykinase protein in murine tissues in diabetes of chemical and genetic origin.

P-enolpyruvate carboxykinase protein was measured by radioimmunoassay in liver, kidney, and adipose tissue extracts from alloxan- and streptozotocin-diabetic rats, in liver extracts from C57BL/KsJ-db+/db+ "diabetic" mice and in liver extracts from normal mice subjected to different dietary or hormonal states. The radioimmunoassay method measured tissue enzyme concentration (nanomoles/g) and total organ enzyme content (nanomoles/liver) independently of assayable activity (units/g). The "apparent" specific activity (units/nmol) was calculated from the maximum velocity and enzyme concentration data. Extracts of rat liver mitochondria and of skeletal muscle cytosol were also analyzed for P-enolpyruvate carboxykinase by the radioimmunoassay. In "chemical" diabetes, P-enolpyruvate carboxykinase by the radioimmunoassay. In "chemical" diabetes, P-enolpyruvate carboxykinase concentration increased approximately 3-fold over the fed, normal liver value of 0.89 microM, approximately 1.6-fold over the normal kidney value of 1.9 microM and approximately 2.9-fold over the normal adipose tissue value of 0.030 microM. Chemical diabetes caused the specific activity to decrease from 0.38 to approximately 0.27 units/nmol in liver and from 0.48 to approximately 0.32 units/nmol in kidney. Insulin replacement by in vivo injection not only promptly lowered the abnormally high enzyme concentration in both tissues but, paradoxically, decreased the apparent specific activities further to approximately 0.16 in liver and to 0.23 in kidney. In the db+/db+ diabetic mouse the liver enzyme increased from 0.22 microM at 5 weeks of age to 0.44 microM at 18 weeks when serum glucagon concentration is known to be highest and the pancreatic beta cells to be depleted of insulin. While the enzyme protein concentration increased 2-fold in the 18-week-old db+/db+ mouse, total liver enzyme content had actually increased 5-fold due to liver enlargement. In fasted normal mice, glucagon-treated normal mice, and alloxan/streptozotocin-treated mice, the concentration of liver enzyme increased significantly compared to the values in fed control mice. Pharmacological doses of dexamethasone did not induce the mouse enzyme. Rat liver mitochondria contained only trace quantities of immunoassayable enzyme which can be explained by contamination with cytosolic proteins. Rat skeletal muscle also contained only insignificant quantities of the enzyme or perhaps another cross-reacting, immunoassayable protein. The data obtained in diabetes before and after treatment show that complex mechanisms of control by insulin and glucagon do operate to regulate P-enolpyruvate carboxykinase in liver, kidney, and adipose tissue.