Protein phosphorylation in isolated mitochondria and the effects of protein kinase C.

When isolated intact rat liver mitochondria are incubated with [gamma-32P]ATP the major phosphorylated proteins are those of 47 and 36 kDa. Phosphorylation of the 47 kDa protein, but not of the 36 kDa protein, is inhibited by carboxyatractyloside, an inhibitor of mitochondrial ATP uptake, while phosphorylation of the 36 kDa protein is inhibited by various uncouplers and an inhibitor of mitochondrial respiration. Addition of purified protein kinase C to the isolated mitochondria leads to the phosphorylation of 69, 37 and 17 kDa proteins.

Cellular uptake and localization of fluorescent derivatives of phorbol ester tumor promoters.

The synthetic fluorescent derivatives of 12-O-tetradecanoylphorbol-13-acetate (TPA), dansyl-TPA, dansyl-TPA-20-acetate and dansyl-TPA-13-desacetate, have ID50 values in the [3H]PDBu binding assay of 2nM, 30nM and 1000nM respectively; the ID50 value of TPA is 4nM. Dansyl-TPA is also equipotent with TPA as an activator of protein kinase C(PKC) producing half maximum stimulation at 2nM. Dansyl-TPA-13-desacetate is almost as potent as dansyl-TPA, while dansyl-TPA-20-acetate is completely inactive as an activator of PKC.

Activation of protein kinase C by tumor promoting phorbol esters, teleocidin and aplysiatoxin in the absence of added calcium.

We have found that maximum stimulation (greater than 10-fold) of kinase activity of a bovine brain preparation of calcium- and phospholipid-dependent protein kinase (PKC) by 12-O-tetradecanoylphorbol-13-acetate (TPA) occurs in the presence of phospholipid, but in the absence of added Ca2+. In effect, nM concentrations of TPA substitute for mM concentrations of added Ca2+, and the two agents are not synergistic. Biologically active analogs of TPA such as phorbol-12,13-dibutyrate (PDBu), 12-O-hexadecanoyl-16-hydroxyphorbol-13-acetate (HHPA) or mezerein were also effective activators of PKC, as were the chemically unrelated tumor promoters teleocidin and aplysiatoxin, when tested at nM concentrations in the absence of added Ca2+.

Mechanisms of multistage chemical carcinogenesis and their relevance to respiratory tract cancer.

The evolution of a fully malignant tumor is a multistep process resulting from the action of multiple factors, both environmental and endogenous, and involves alterations in the function of multiple cellular genes. Chemical carcinogens that initiate this process appear to do so by damaging cellular DNA. In addition to producing simple point mutations, this damage appears to induce the synthesis of a transacting factor that can induce asynchronous DNA replication.

Preparation and properties of nickel hemoglobin.

Hemoglobin A reconstituted with nickel protoporphyrin IX (NiHbA) has been prepared and characterized. Kinetics of its reaction with p-mercuribenzoate and with haptoglobin, absorption and circular dichroism spectra, and x-ray crystallographic properties have been investigated as probes of its structural conformation. The results suggest that NiHbA exists in a structure that is similar to the deoxy, or T-state of HbA.