Involvement of protein kinase C- in DNA damage-induced apoptosis

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Caspases are essential for the execution of cell death by apoptotic stimuli.1,2,3,4 The pathway of cell death varies depending on the cell type as well as the apoptotic stimuli. It is generally believed that binding of Fas ligand or tumor necrosis factor- (TNF) to their receptors causes activation of the initiator caspase-8 followed by the activation of a caspase cascade to execute cell death.2,5 In contrast, DNA damaging agents are known to induce release of mitochondrial Cytochrome c, which facilitates the interaction of apoptotic protease activating factor (Apaf-1) with procaspase-9 to initiate the activation of downstream effector caspases, such as caspases-3 or -7 to cause cell death.3 Both receptor-mediated and anticancer drug-induced apoptosis may, however, involve more than one pathway and there may be cross-talk between these two pathways.6,7,8

cis-Diamminedichloroplatinum(II) (cDDP or cisplatin) is one of the most important anticancer agents used for the treatment of solid tumors.9 Although the antitumor activity of cDDP is believed to be due to its interaction with chromosomal DNA, only a small fraction of cDDP actually interacts with DNA and inhibition of DNA replication cannot solely account for its biological activity.10 The efficacy of chemotherapeutic drugs not only depends on their ability to induce DNA damage but also on the cell's ability to detect and respond to DNA damage.11 cDDP, like other chemotherapeutic drugs, causes activation of caspases although the sequence of events that follow cDDP-induced DNA damage and lead to apoptosis remains to be unraveled.

We and others have shown that the PKC signal transduction pathway regulates cell death by cDDP.12,13,14,15,16,17 PKC is a family of 11 isozymes that are classified as the conventional PKCs (, I, II and ), novel PKCs (, , , and ), atypical PKCs ( and / ) and novel/atypical PKC.18 PKC is a substrate for caspase-3 and the catalytic fragment of PKC has been directly associated with apoptotic cell death.19,20 We have, however, demonstrated that downregulation of PKC that decreased the abundance of PKC catalytic fragment was associated with increased cellular sensitivity to cDDP.17 These results raise the possibility that PKC acts upstream of caspases to regulate cell death by cDDP. It is not known which PKC isozyme regulates activation of caspases and which step(s) of the cisplatin-induced cell death pathway is regulated by PKC.

Mitochondria play a pivotal role in the decision making process of a cell's life and death.21 It is believed that once Cytochrome c is released from mitochondria, cells are committed to die.22 An inability to induce release of Cytochrome c from mitochondria has been associated with cellular resistance to anticancer agents, including cDDP.23 In the present study, we have investigated how PKC regulates release of Cytochrome c and activation of caspases that emanate from mitochondria. Our results show that in HeLa cells, PKC was localized not only in the cytosol but also in the mitochondrial fraction and cDDP induced processing of both cytosolic and membrane-associated PKC. Rottlerin, a specific inhibitor of PKC, blocked cDDP-induced activation of caspases and proteolytic cleavage of PKC in both cytosolic and HM fractions but inhibited only late but not early release of Cytochrome c. Furthermore, inhibition of nPKC, but not of cPKCs, protected cells against cDDP-induced cell death. Taken together, these results demonstrate that PKC influences cDDP-induced cell death by acting at an early step of the mitochondrial cell death pathway that precedes activation of caspases.

Camphor

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Description
The camphor tree is a dense broadleaved evergreen that is capable of growing 50-150 ft (15.2-45.7 m) tall and spreading twice that wide with a trunk up to 15 ft (4.6 m) in diameter, though the largest U.S. specimens are only half that size and those in the Caribbean are even smaller. The shiny foliage is made up of alternate 1-4 in (2.5-10.2 cm) oval leaves dangling from long petioles. Each leaf has three distinct yellowish veins. The outer margins of the leaves tend to be somewhat wavy and turn upward. The new foliage starts out a rusty burgundy color, but the leaves soon turn dark green on the upper sides and paler green underneath. New branches emerging from the shallowly fissured grayish brown trunk are smooth and green. Twigs are usually green, but may be tinged with red when young. The inconspicuous tiny cream colored flowers are borne in the spring on branching 3 in (7.6 cm) flower stalks. They are followed by large crops of fruit, comprised of round pea sized berries attached to the branchlets by cuplike little green cones. The berries first turn reddish, then ripen to black. Camphor tree can be readily identified by the distinctive odor of a crushed leaf.


This camphor tree shades a house in North Florida - a landscape use no longer considered appropriate for this invasive species.
Location
Cinnamomum camphora, the camphor tree, comes from China, Japan, Korea, Taiwan, and adjacent parts of East Asia, where it grows in mesic forests and on well-drained sites along streambanks. Camphor has become widely naturalized in Australia. In the United States, it is grown along the Gulf Coast and in California, and has escaped cultivation and become naturalized in many areas.

Culture
Camphor prefers fertile sandy soil. It will tolerate a pH anywhere in the range of 4.3 to 8. The roots are very sensitive to disturbance. They may extend far from the trunk of the tree, and can readily be identified by their characteristic odor.
Light: Camphor will grow in full sun or partial shade.
Moisture: Camphor tree does not do well in wet soils. Established trees are tolerant of drought.
Hardiness: USDA Zones 8 - 10. Hardened off camphor trees can survive freezes down to 10-15ºF (-12 - -9ºC), but new growth will suffer freeze burn when the temperature drops below 32ºF (0ºC) and branches will die back from temperatures in the low twenties.
Propagation: Camphor seed does not remain viable for long and should be planted in the greenhouse as soon as it ripens. Remove the fruit pulp first. At 68ºF (20ºC), germination will take 1-6 months. Cuttings of semiripe side shoots can be rooted in a warm humid place in midsummer. Pieces 2-3 in (5.1-7.6 cm) long with a heel work best.

Usage
Camphor is widely planted as a shade tree, screen, or windbreak. In China and Japan, it is grown commercially for its medicinal oil. Camphor oil has a strong penetrating fragrance, a pungent bitter flavor, and feels cool on the skin like menthol, though it also has irritating qualities as well as a numbing effect. Camphor has been used to treat ailments ranging from parasitic infections to toothaches. Scientific evidence has confirmed that chemicals in the plant have value in antiseptics and medications for treating diarrhea, inflammation, itching, and nervous conditions. Camphor wood is prized for its attractive red and yellow striping, amenability to woodworking, and insect repelling properties. It is light to medium in weight and soft to medium in hardness. Wood from the camphor tree is not especially strong, but it takes polishing well. It is commonly used for chests, closets, coffins, instruments, and sculptures. Camphor veneer is used in fine cabinetry. Camphor is also used in perfumes.


camphor tree trunk and bark
Features
This is a sturdy storm resistant tree which makes a good windbreak. Since it is hard to burn, it should also be valuable as a shade tree in areas that are prone to wildfires. Unfortunately, these desirable traits are offset by the tree's invasiveness and damaging effects on wildlife and natural communities. This fine tree should be grown and appreciated in its native range, but not planted in other regions where species and ecosystems have not adapted to its aggressiveness and toxicity. Camphor tree should not be grown in the United States.

WARNING
Camphor in large doses is toxic to humans. It stimulates the central nervous system and may affect respiration or cause convulsions. In Chinese medicine, camphor is forbidden for pregnant women and those with a deficiency of vital energy or yin. Camphor is a prolific seed producer that apparently does not have serious predators or diseases outside its native range. Seedlings and root sprouts are abundant near mature trees, but individual trees pop up far from seed sources. In Florida, camphor trees appear in undisturbed mesic hardwood forests, upland pine woods, and scrubs, as well as in the vacant lots and fencerows where it is more commonly observed. The Plant Conservation Alliance lists this species as an Alien Invader and it is listed as a Category I invasive exotic species by the Florida Exotic Pest Plant Council, which means that it is known to be "invading and disrupting native plant communities in Florida