It is possible to consider presented research as first step of our group on the way of apoptosis research and, more exactly – their antagonist – antiapoptosis. We have reconstructed pathways of NFkB-dependent transcription activation - the main process defined antiapoptotic response. In the future exactly this research allowed us to start detailed kinetic model of one of the main unit of these processes.

The common scheme of pathways reconstructed in the framework of this project may be illustrated by following way (see the figure). As it may have seen, processes connected with NFkB–dependent transcription activation, adjoin narrowly not only with apoptosis but also with signal transmission pathways from hormones and growth factors (particularly, with processes dependent on secondary messenger - inositol triphosphate). It is very important because at the expanse of it regulatory network of this phenomenon essentially ramify. Moreover, scientists were found that NFkB-dependent transcription could be directly started by some interleukins (important elements of immune system). All this observation makes the researches of problems connected with antiapoptosis extremely interesting and perspective. Surely, the frames of this project do not allow investigating comprehensively described phenomena, limiting only by structuring and concentration of information.

An example of one element of this system reconstruction – activation and inhibition pathways of apoptotic reaction cascade development is given below. By this example we would like to show what the usual view of such research results presentation (scheme + biological description of various degree of detailing) is. For short description we significantly reduce the text given in order not to overload readers by very detailed biological information.

Apoptotic processes may be started in the cell by two methods. Firstly, because of violation of mitochondria functioning and action of special proteins of Bcl-2 family connected with it; secondly, during signal transmission from special intracellular regulators.

We stop in detail on the second method, i.e. on the processes being started by means of extracellular signal. There are several signal proteins - TNF, TRAIL, FasL. They act on the corresponding receptors, located in plasmatic cell membrane (TNFR, TRAIL-R1-4, FAS). At that, as a rule, binding together ligand and receptor results in ambiguous intracellular responses, i.e. both to activation and blocking of apoptosis. Thus, when TRAIL interact with TRAIL-R1 or with TRAIL-R2, special domain (death domain) is activated on the inner cytoplasmic side of receptor, and with it can bind various adaptor proteins FADD (FAS-associated death domain protein) or RIP (receptor interacting protein). Binding of one or another protein will define where signal will be directed. In case when FADD acts, signal will be transferred to the apoptotic pathways by means of procaspase-8 stimulation; in contrast, in case of RIP antiapoptotic pathways will be activated.

Quite interesting is process of procaspase-8 activation. Caspases are named the family of aspartate-specific cysteine proteinase, several of them are able to auto- and crossproteolysis. Exactly that happens with procaspase-8 after its binding with FADD. It cuts the piece of their own peptide sequence, rifts at that from FADD and goes to the cell cytoplasm as active caspase-8. This moment may be considered as transmission of external signal into cell and beginning of the caspase reaction cascade, and consequently, as starting apoptosis of the cell.

It is important to say some words about two other receptors of TRAIL-R family (third and forth type), which are located in plasmatic cell membrane. Both of them belong to the antiapoptotic elements i.e. during signal transmission from TRAIL they promote apoptosis inhibition, having different mechanism of inhibition at that. Thus, TRAIL-3 does not start any intracellular cascade, and only decreases the free concentration of ligand. This type of inhibition is named inductor exception (in this case, extracellular signal). In contrast, TRAIL-4 may start some intracellular processes, which have not been investigated up to now, but their final result is, as during the interaction TRAIL-R1,2 with RIP, blocking of apoptotic reaction cascade development by means of NFkB-dependent gene transcription activation.

Another variant of apoptosis inhibition is FADD interaction with с-FLIP (FLICE inhibitory protein). This protein is able to take procaspase-8 binding site on FADD. с-FLIP expression in healthy cells not liable to changes, which are observed during cancer cell formation, is NFkB-dependent.

Having come to the cytoplasm, caspase-8 may act in two ways. Firstly, to start caspase cascade by caspase-3 activation; secondly, lead to mitochondria destruction and mitochondrial apoptosis starting (see above) by means of proteins of Bcl-2 family. For that caspase-8 cut the part of small protein BID, transferring it to the active form - tBID. This activated agent promotes binding Bax (other protein from Bcl-2 family) with mitochondrial porine VDAC that lead to pore formation in the external mitochondrial membrane. Through these pores from intermembranous mitochondrial space citochrome c and regulatory protein Smac/DIABLO, taking part in the repulsing of antiapoptotic attack of IAP-1 and IAP-2, go to the cytoplasm. In the cytoplasm cytochrome c binds with proapoptotic adapter protein APAF-1. After that procaspase-9 joined to the cytochrome c and is activated. This complex named apoptosome. Apoptosome, as caspase-8 can start antiapoptitic reaction cascade (by means of caspase-3 activation) The result of this is activation of the great number of terminal caspase (types 2,3,6,7). These enzymes promote non-selective degradation of structural and regulatory proteins, such as lamine, cytokeratins and PARP (poly (ADP-ribose) polymerase), that irreversibly lead to cell death.

Mitochondrial apoptosis and ligand-dependent as well are inhibited by products of NFkB-dependent transcription. Firstly, by means of Bcl-2 and Bclxl (other representatives of Bcl-2 family regulatory proteins) pore formation in the external membranes of mitochondria is blocked and secondly, special proteins IAP-1 and –2 are synthesized, which are inhibitors of caspase –9 and -3, and decrease the rate of expansion of antiapoptotic reaction cascade by this.