Biochemische und molekularbiologische Charakterisierung von Spike, einem neuen BH3-Domänen Protein

During my PhD thesis I was working on the identification of novel
apoptosis-inducing genes by using a novel genetic expression screen
(Grimm and Leder, 1997). One of the identified genes turned out to
be a evolutionary conserved cDNA that codes for a novel BH3-only
protein of 219 amino acid residues which was named Spike, for Small
protein with inherent killing effect. Spike was then in the course
of my PhD thesis extensively characterised, molecularly and
biochemically. In summary, upon overexpression in mammalian cells
Spike efficiently leads to all features of apoptosis, such as
phenotypic alterations, cytochrome c release, caspase activation
and DNA degradation. It was shown that Spike localised to the
endoplasmic reticulum, where it interacts with a recently
identified apoptosis regulating protein complex, consisting of
Bap31, Bcl-2, Bcl-XL and an ER-specific isoform of caspase-8:
pro-caspase 8L (Breckenridge et al. 2002). Although no direct
interaction with anti-apoptotic members of the Bcl2-family could be
observed, the importance of the BH3-like sequence for the
apoptosis-inducing activity of Spike was demonstrated by using
point mutations of conserved amino acid residues of this motif
(Mund et al. 2003). Instead of directly interacting with
anti-apoptotic members of the Bcl-2 family Spike is able to
interfere with the complex formation between Bap31 and Bcl-XL.
Based on these data I proposed a model according to which the
complex on Bap31 is controlled by Bcl-2/Bcl-XL as long as they are
associated. Displacement of Bcl-2/Bcl-XL from Bap31 by Spike leads
to the formation of a pro-apoptotic complex. In addition, Spike
appears to be implicated in the cell death signal of the Fas
receptor. I observed that a dominant-negative version of Spike and
a highly effective anti-sense oligonucleotide significantly reduced
Fas-mediated DNA fragmentation whereas no reduction was detected in
TNFa-induced cell death (Mund et al. 2003).