Human Bak Induces Cell Death in Schizosaccharomyces pombe with Morphological Changes Similar to Those with Apoptosis in Mammalian Cells

Abstract

Apoptosis as a form of programmed cell death (PCD) in multicellular organisms is a well-established genetically controlled process that leads to elimination of unnecessary or damaged cells. Recently, PCD has also been described for unicellular organisms as a process for the socially advantageous regulation of cell survival. The human Bcl-2 family member Bak induces apoptosis in mammalian cells which is counteracted by the Bcl-x L protein. We show that Bak also kills the unicellular fission yeast Schizosaccharomyces pombe and that this is inhibited by coexpression of human Bcl-x L . Moreover, the same critical BH3 domain of Bak that is required for induction of apoptosis in mammalian cells is also required for inducing death in yeast. This suggests that Bak kills mammalian and yeast cells by similar mechanisms. The phenotype of the Bak-induced death in yeast involves condensation and fragmentation of the chromatin as well as dissolution of the nuclear envelope, all of which are features of mammalian apoptosis. These data suggest that the evolutionarily conserved metazoan PCD pathway is also present in unicellular yeast.Programmed cell death (PCD) in metazoans is an essential homeostatic mechanism permitting the removal of surplus cells during morphogenesis and tissue maintenance and the deletion of cells that present a risk to the organism because they are mutated or infected (10,19,35,36,40). For vertebrates, the descriptive name commonly given to the process of PCD is apoptosis. Classical apoptosis is characterized by membrane blebbing, cell shrinkage, chromatin condensation, and nuclear and cellular fragmentation, and it results from the activation of an intrinsic suicide program (47). Recent studies implicate the dysregulation of PCD in the pathophysiology of several human diseases, including AIDS (12, 28), neurodegenerative disease (25,37,44), and cancer (for a review, see reference 42).The basal machinery responsible for metazoan PCD is highly evolutionarily conserved and, at its execution level, involves the action of a discrete class of cysteine proteases, of which the prototypes are the interleukin-1␤-converting enzyme in humans and Ced-3 in the nematode (49). Also conserved are key regulators of apoptosis: in Caenorhabditis elegans the Ced-9 protein and in humans the Bcl-2 protein family (18, 46), which comprises both suppressors (e.g., Bcl-2 and Bcl-x L ) and promoters (e.g., Bax and Bak) of PCD (17,30,46).Recently, there have been several reports describing apparent PCD in the unicellular eukaryotes Tetrahymena thermophila, Dictyostelium discoideum, Trypanosoma brucei rhodsiense, and Trypanosoma cruzi and even in bacteria (2, 6, 45, 48; for an overview, see reference 1). PCD in unicellular organisms might facilitate constant selection for the fittest cell in the colony or optimal adaptation of cell numbers to the environment or might serve as a means for altruistic cell death to prevent the spread of virus in the event of infection.It has been shown that expression of the mammalian Bax protein in th...

Sequence and partial functional analysis of canine Bcl-2 family proteins

“…Here, we successfully performed a functional analysis of five canine Bcl-2 family members in this model organism, namely the pro-apoptotic members Bak and Bax and the anti-apoptotic members Bcl-w, BclxL and Mcl-1. As previously described for the human and/or murine orthologs (Ink et al, 1997;Tao et al, 1997;Ligr et al, 1998), expression of canine Bak and Bax showed a lethal effect in yeast. Moreover, this effect was abrogated on co-expression of the anti-apoptotic members Bcl-xL, Mcl-1 and Bcl-w as previously demonstrated for large part of their human orthologs in yeast (Tao et al, 1997;Beaumont et al, 2013).…”

Apoptosis without caspases: an inefficient molecular guillotine?

“…152 Yeast can be killed by various means such as oxidative stress, metabolic block, irradiation and other toxic substances, but cell death does not resemble apoptosis and there is no endogenous expression of known apoptosis regulators of the CED-3/caspase, CED-4 or Bcl-2/Bax families. Nevertheless, forced overexpression of Bax, Bak or CED-4 provokes vacuolarization and focal chromatin c o n d en s at i o n a s s ee n i n m am m al i an a po p t osis, 28,150,151,154,155 indicating these death factors can indeed provoke a caspase-independent form of apoptosis via components that are already present in a unicellular organism. To identify these components, a mammalian expression cDNA library has recently been screened for clones that rescue yeast from Bax-mediated cell death.…”

Subcellular and submitochondrial mode of action of Bcl-2-like oncoproteins

“…Transfection of yeast cells (Saccharomyces cerevisiae or Schizosaccharomyces pombe), whose entire genome is sequenced and lacks Bcl-2-like proteins, has shown that Bax and Bak clearly have a killer activity which can be neutralized by simultaneous expression of Bcl-2 or Bcl-X L (Table 4) (Ink et al, 1997;Jurgensmeier et al, 1997;Manon et al, 1997;Sato et al, 1994;Torgler et al, 1997;Zha et al, 1996a). Physical interaction between Bcl-2 and Bax is not required for the Bcl-2-mediated inhibition of Bax-induced apoptosis (Zha and Reed, 1997).…”

“…Note that certain Bcl-2 homology regions (BH1 through BH4, in solid) or C-terminal hydrophobic membrane insertion sequences (tinted) are missing in determined members of the family. Some structural features (a domains) and functions determined by cristallographic or mutational analysis are indicated (Longo et al, 1997) Bcl-X L ; Mcl-1; A1 Prevent Bax/Bak-mediated killing (Manon et al, 1997;Sato et al, 1994;Tao et al, 1997) S. Pombe Bax Cell killing with chromatin condensation and vacuolization (Jurgensmeier et al, 1997) Bak Slow growth phenotype and killing with chromatic condensation and lamin dissolution (Ink et al, 1997;Jurgensmeier et al, 1997;Torgler et al, 1997) Bcl-X L , Bcl-2 Prevent Bax/Bak-mediated e ects (Jurgensmeier et al, 1997;Ink et al, 1997;Torgler et al, 1997) CED-4 Causes chromatin condenstaion and death CED-9 Prevents CED-4-induced death lular distribution of di erent members of the Bcl-2 family can depend on the cell type as well as on the family member. Thus, Bcl-2, Bcl-X L , Bax, and the Epstein ± Barr virus gene product BHRF tend to be particularly abundant in the outer mitochondrial membrane (Cory, 1995;Gonzalez-Garcia et al, 1994;Krajewski et al, 1993;Reed, 1994;Riparbelli et al, 1995;Yang and Korsmeyer, 1996;Zha et al, 1996a).…”