Brain Development: Normal Processes and the Effects of Alcohol ...

94 NORMA L DEVELOPMENT
but lac k BH4-e.g., Bax , Bcl-x s, Bak , and Bok/Mtd ,
or (2) the BH3-only subgroup that includes Bid , Bini/
Bod, Bad, Bmf, Bik/Nbk, Blk, Noxa, Puma/Bbc3, an d
Hrk/DP5.
Bel family members form dimers . Homodimers of
two anti-apoptotic members , o r heterodimers formed
by one pro-apoptotic an d one anti-apoptotic member ,
for example , Bcl-2-Bax , ar e pro-survival . I n contrast ,
homodimers o f pro-apoptoti c members , o r het -
erodimers o f tw o pro-apoptoti c members , resul t i n
death. The BH 3 domain appears to be the prime suspect
fo r inducin g apoptosis . Protein s wit h th e BH 3
domain have two functions: (1 ) to bind to and inactivate
the survival-inducing Bel proteins, and (2 ) to activate
th e pro-apoptoti c Be l protein s wit h multipl e B H
domains, such a s bax and ba k (Desagher e t al, 1999 ;
Wei et al, 2000, 2001; Degterev et al, 2001). According
to on e mode l o f apoptosis , Ba k i s a trans-membran e
protein in the outer mitochondrial membran e an d Bax
is i n th e cytoplas m (Lucken-Ardjomand e an d Marti -
nou, 2005) . An apoptoti c signa l cause s a conforma -
tional change in both proteins that results in exposure
of their N terminal s an d translocatio n o f Bax to th e
mitochondrial oute r membrane . Onc e i n the mem -
brane, Bak and Bax form oligomers that induce mem -
brane permeabilit y an d thu s allo w movemen t o f
molecules suc h a s cytochrorne C , HtrA2/Omi , an d
Smac/DIABLO.
Caspases
Caspases ar e a group of structurally related enzyme s
in mammals. Genes fo r caspases have homology with
the C. elegans gene Ced3 (e.g., Jiang and Wang, 2004;
Shiozaki an d Shi , 2004) . Lik e th e gen e produc t o f
Ced3, caspase s are importan t fo r cell death . I n con -
trast to Ced3, however, caspases are no t require d for
death (se e below) . Caspases ar e proteases ; a t leas t 1 4
caspases have been identified. They have cysteine s i n
their activ e site s an d cleav e thei r targe t protei n adja -
cent to an asparti c aci d residue . Caspase s ar e synthe -
sized a s inactiv e isoform s an d ar e cleave d int o larg e
and small subunits. Active caspases are tetramers com -
posed of two large and two small subunits.
Caspases ar e groupe d int o thre e subfamilies :
interleukin-lp-converting enzyme-lik e caspases , ini -
tiator caspases (e.g., caspase 8 or caspase 9), and effec -
tor caspase s (e.g. , caspas e 3 ) (Shi , 2002) . Initiato r
caspases can cleav e themselves and effecto r caspase s
are cleave d b y othe r enzymes , includin g initiato r
caspases. Activatin g initiato r caspase s ca n induc e a
caspase cascade i n which effecto r caspase s are cleave d
and the n othe r apoptosis-relate d downstrea m mole -
cules are activated. Caspase s ca n cleave more than 40
target proteins ; o f particula r importanc e ar e a n in -
hibitor of DNases, nuclea r laminins, and cytoskeletal
proteins. Inactivatio n o f DNas e inhibitor s allow s
DNases to cut DNA into fragments that are multiples
of 180-200 bp. The resul t of this serial snipping is the
stereotypical DNA ladder on a Southern blot that is associated
with apoptosis. Cleavage o f nuclear laminins
results i n nuclea r fragmentation . Similarly, cleavage
of cytoskeleta l element s result s i n membran e bleb -
bing and, ultimately, cell fragmentation.
The importanc e o f caspase s i n norma l develop -
ment i s reflected in mice that lack effective caspas e 3
or 9 . Bot h mutant s hav e craniofacia l malformations
and abnorma l brai n morpholog y (Kuid a et al., 1996 ,
1998; Ceccon i e t al , 1998 ; Hake m e t al , 1998) .
Specifically, these animals exhibit enlarged prolifera -
tive zones , heterotopi c cluster s o f neurons , an d in -
creased number s o f neurons. Interestingly , caspas e 3
deficiency ca n b e offse t b y deficienc y i n th e anti -
apoptotic gen e Bcl-x L (Rot h e t al. , 2000) , implyin g
that Bcl-x L can ac t through a caspase-3-independent
mechanism.
Apoptosis Protease Activating Factor 1
APAF-1, homologous to Ced4, exists in the cytoplasm
as an inactiv e protein (Ferrar o et al, 2003 ; Shiozaki
and Shi, 2004). Interaction with cytochrome C in the
presence of adenosine triphosphate (ATP) results in a
conformational chang e i n APAF- 1 that allow s seven
molecules o f APAF-1 to assemble an d for m th e cen -
tral portion o f the apoptosome . I n addition, th e con -
formational chang e expose s the caspas e recruitmen t
domain, which bind s procaspase 9 . In the absence of
an inhibitory apoptotic protein (IAP ) such as X-linked
IAP (XIAP) , caspas e 9 i s activate d vi a a n unknow n
mechanism. Activ e caspase 9 recruits and activates effector
caspases 3 and 7 .
APAF-1 knockou t mic e ar e embryoletha l (Cec -
coni e t al, 1998) . The y have craniofacia l an d CN S
malformations tha t appear to result from insufficien t
cell deat h durin g development . Interestingly , their
limbs ar e differentiall y affected . O n gestationa l da y
(G) 15.5 , the limbs are less mature those than in wildtype
animals, as evidenced by the persistence of interdigital
webbing . B y G16.5 , however , thei r limb s