K. Rothbarth et al., Induction of apoptosis by overexpression of the DNA-binding and DNA-PK-activating protein C1D, J CELL SCI, 112(13), 1999, pp. 2223-2232
Apoptosis is induced in various tumor cell lines by vector-dependent overex
pression of the conserved gene C1D that encodes a DNA-binding and DNA-PK-ac
tivating protein. C1D is physiologically expressed in 50 human tissues test
ed, which points to its basic cellular function. The expression of this gen
e must be tightly regulated because elevated levels of C1D protein, e.g. th
ose induced by transient vector-dependent expression, result in apoptotic c
ell death. Cells transfected with C1D-expressing constructs show terminal d
eoxynucleotidyl transferase-mediated dUTP nick end-labeling of DNA ends. Tr
ansfections with constructs in which C1D is expressed in fusion with the (e
nhanced) green fluorescent protein from A. victoria (EGFP) allow the transf
ected cells to be identified and the morphological changes induced to be tr
aced. Starting from intense nuclear spots, green fluorescence reflecting C1
D expression increases dramatically at 12-24 hours post-transfection, Expre
ssion of C1D-EGFP protein is accompanied by morphological changes typical o
f apoptotic cell death, e.g. cytoplasmic vacuolation, membrane blebbing and
nuclear disintegration. Cell shrinkage and detachment from extracellular m
atrix are observed in monolayer cultures while suspension cells become prog
ressively flattened. The facility to differentiate between transfected and
nontransfected cells reveals that non-transfected cells cocultured with tra
nsfected cells also show the morphological changes of apoptosis, which poin
ts to a bystander effect.
C1D-dependent apoptosis is not induced in cells with non-functional p53, Ac
cordingly, C1D-induced apoptosis is discussed in relation to its potential
to activate DNA-PK, which has been considered to act as an upstream activat
or of p53.