Nuclear envelope (NE) cisternal Ca2+ and cytosolic ATP are required for nuc
lear-pore-complex (NPC-) mediated transport of DNAs, RNAs, transcription fa
ctors and other large molecules. Isolated cardiomyocyte nuclei, capable of
macromolecular transport (MMT), have intrinsic NPC ion channel behavior. Th
e large ion conductance (gamma) activity of the NPC channel (NPCC) is block
ed by the NPC monoclonal antibody mAb414, known to block MMT, and is also s
ilenced during periods of MMT. In cardiomyocytes, neither cytosolic Ca2+ no
r ATP alone directly affects NPCC gating. To test the role of Ca2+ and ATP
in NPCC activity, we carried our the present patch-clamp study with the pip
ette attached to the outer NE membrane of nuclei isolated from cultured Dun
ning G prostate cancer cells. Our investigations demonstrate that in these
isolated nuclei neither cytosolic Ca2+ nor ATP alone directly affects NPCC
gating. However, when simultaneously applied to the bath and pipette, they
transiently silence NPCC activity through stimulation of MMT by raising the
Ca2+ concentration in the NE cisterna ([Ca2+](NE)). Our fluorescence micro
scopy observations with nuclear-targeted macromolecular fluorochromes (B-ph
ycoerythrin and plasmid for the enhanced green fluorescence protein EGFP, p
EGFP-C1) and with FITC-labeled RNA support the view that channel silence ac
companies MMT. Repeated Ca2+ loading of the NE with Ca2+ and ATP, after unl
oading with 1-5 mu M inositol 1,4,5-trisphosphate (IP3), thapsigargin (TSG)
or 5 mM BAPTA or EGTA, failed to affect channel gating. This result indica
tes that other factors are involved in this phenomenon and that they are ex
hausted during the first cycle of NE Ca2+ loading/unloading - in agreement
with current theories of NPC-mediated MMT. The results explain how Ca2+ and
IF, waves may convert the NE into an effective Ca2+ barrier and, consequen
tly, affect the regulation of gene activity and expression through their fe
edback on MMT and NPCC gating. Thus, [Ca2+](NE) regulation by intracellular
messengers is an effective mechanism for synchronizing gene activity and e
xpression to the cellular rhythm.