A whole organism, such as a fly or a human, originates from a mass of uniform cells that then differentiates into various organs, skin, teeth, hair, etc. This devel­opment program also entails the intentional death of specific cells in order to sculpt the structures that make the organism, which istermed "apoptosis" or programmed cell death. For example, the loss of a tadpole's tail as it develops into a frog is due to apoptosis, as is the formation of fingers and toes, which entails the removal of tissue between them. Apoptosis also occurs when cells are damagedor infected by pathogens; the celldeath program is activated for the benefit of the entire organism. It is obvious that apoptosis is a vital mechanism for the function of a multi-cellular organism.

What happens when apoptosis doesn't occur? Cancer is a disease state caused by the uncontrolled division of cells due to genetic or environmentally caused mutations, which deprive the normal cells of nutrients and resources resulting in great discomfort and often death for the organism. Apoptosis is generally impaired in these cells, despitetheir mutated state, indicating that apoptosis is one of the body's primary natural defenses against cancer by preventing propagation of cancerous cells. In fact, the current chemotherapy and radiation treatments available todayare known to induce apoptosis in cancer cells, but they do not kill the cells selectively and do harmthe healthy normal cells. It is alsoknown that cancer cells can adapt to the therapeutic treatments and escape the cell death program, leading to highly resistant and persistent cancerous cells.

At the same time, inappropriate activation of the cell death program can lead to other diseasestates, such as stroke, spinal cord injury, Alzheimer's disease, Parkinson's disease, and other neurodegenerative diseases. Therefore, it is not surprising that a great deal of research is focused on how to understand and thus control apoptosis in order to develop improved therapies for the treatment of various diseases.

Apoptosis is characterized byseveral cellular events: the cells shrink, the membranes start to bubble up, the genetic material

breaks down and the cell is rapidly engulfed and cleared

The developing Drosophila (fruit fly) eye as a model system for studying the regulation of apoptosis. Left, an eye from a fly mutant that overproduces the Reaper protein resulting in an overacti­vation of apoptosis and thus a smaller eye. Right, an eye from a fly mutant that cannot effectively ubiqutinate proteins to target them for destruction and Reaper cannot function pro perly resulting in a larger eye.

by phagocytes. It is currently known that a multitude of externalsignals such as DNA damage and developmental errors culminate in the activation of a group of enzymes called cysteine aspartases, or caspases. These enzymes are the proteins that actually execute the functions that result in

apoptosis by destroying key cellular components.

Dr. Hermann Steller and his colleagues at the Rockefeller University are interested in understanding what molecules activate and inhibit caspases, and how they do so. Their primary model system is that of the fruit fly, Drosophila melanogaster, which is easy to study and manipulate genetically. In addition,they are also using reverse genetics in the mouse to test whether concepts originally established in Drosophila can be applied to understanding mammalian apoptosis. They began by looking for fly mutants that are globally blocked for apoptosis to identifygenes that are required for all cell death. The genes Reaper, Hid,

Dr. Hyung-Don

Dr. Hermann Stellar (right) Rockefeller University’s Strang Laboratory of Apoptosis and Cancer Biology. The work described in this article was published in the journal Nature Cell Biology in 2002 (vol. 4, p. 546).

and Grim, were identified as themessengers of death—they were able to induce apoptosis when introduced into healthy cells, theywere only active in cells doomedto die, and many pathways con­trol their activity. Reaper, Hid,and Grim link all the signaling pathways to the cell death program, and they do so by inhibiting a family of proteins named Inhibitors of Apoptosis Proteins, or IAPs for short.

A closer look at the interaction between IAPs and the Reaper protein showed that a conservedN-terminal sequence in Reapernamed the RHG motif binds to arigid surface pocket in the BIR domains in the IAPs, and this binding interaction is strongly conserved between species.

A simplified schematic diagram highllighting the core death program involving Reaper, Grim, and Hid, the IAPs and caspases.

 

Reaper (RPR) reduces the stability of IAPs (DIAPI). Peptides or mimetics containing the RHG motif (stroped triangle) nhibits the binding of IAPs to capsases via the BIR domain (check­ered box) in the IAP

The BIR domains in IAPs alsobind caspases, and Dr. Steller's studies indicate that Reaper family proteins contribute to

apoptosis by displacing the caspases from the BIR domains, resulting in the liberation and thus the activation of caspases.

Reaper also reduces the stability of IAPs through the RING domain of the IAPs, which is a ubiquitin ligase that binds to ubiquitin-conjugating enzymes resulting in the addition of ubiquitin to the IAP itself. Thisprocess is termed "auto-ubiquitina­tion" and ubiquitin is a chemical tag that targets proteins to be destroyed by the cell. Therefore,Reaper family proteins have the dual role of liberating caspases and promoting IAP degradationin order to effect apoptosis. It isevident from this body of knowledge that IAPs are attractive targets for controlling apoptosis in order to treat cancer.

The process of apoptosis is highly conserved from the fly to the human. IAPs are known to beover-expressed in human cancers and promote tumor survival; therefore reduction or eliminationof IAP expression and function incancer cells by Reaper-like molecules may lead to the killing of tumor cells by apoptosis. Dr. Steller and his colleagues are pursuing this avenue of investigation by constructing Reaper/Hid/Grim­mimetics ("RHG-mimetics"), which disrupt the interaction between the IAPs and the caspases and neutralize the IAPs themselves. These molecules are small,cell-permeable synthetic peptides derived from Reaper, Hid, and Grim that will hopefully kill cancer cells with very high selectivity, which is always the desired outcome of a cancer therapeutic. They are currently obtaining proof of principle with peptides based on the native sequence of the Reaper protein,and studies are underway to test the peptides in mice.

If you are interested in learningmore about Dr. Steller's work atRockefeller University, pleasevisithis lab webpage at: http://www.rockefeller.edu/ labheads/steller/steller-lab.php