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The retinoblastoma protein (pRB) is a critical regulator of cell proliferation and differentiation and an important tumor suppressor. In the G 1 phase of the cell cycle, pRB localizes to perinucleolar sites associated with lamin A/C intranuclear foci. Here, we examine pRB function in cells lacking lamin A/C, finding that pRB levels are dramatically decreased and that the remaining pRB is mislocalized. We demonstrate that A-type lamins protect pRB from proteasomal degradation. Both pRB levels and localization are restored upon reintroduction of lamin A. Lmna -/- cells resemble Rb -/- cells, exhibiting altered cell-cycle properties and reduced capacity to undergo cell-cycle arrest in response to DNA damage. These findings establish a functional link between a core nuclear structural component and an important cell-cycle regulator. They further raise the possibility that altered pRB function may be a contributing factor in dystrophic syndromes arising from LMNA mutation.

In primary mammalian cells, DNA replication initiates in a small number of perinucleolar, lamin A/C-associated foci. During S-phase progression in proliferating cells, replication foci distribute to hundreds of sites throughout the nucleus. In contrast, we find that the limited perinucleolar replication sites persist throughout S phase as cells prepare to exit the cell cycle in response to contact inhibition, serum starvation, or replicative senescence. Proteins known to be involved in DNA synthesis, such as PCNA and DNA polymerase delta, are concentrated in perinucleolar foci throughout S phase under these conditions. Moreover, chromosomal loci are redirected toward the nucleolus and overlap with the perinucleolar replication foci in cells poised to undergo cell cycle exit. These same loci remain in the periphery of the nucleus during replication under highly proliferative conditions. These results suggest that mammalian cells undergo a large-scale reorganization of chromatin during the rounds of DNA replication that precede cell cycle exit.

The brain of Alzheimer's disease (AD) patients contains deposits of amyloid beta-peptide (A beta). Recent studies have shown that A beta is a substrate for tissue transglutaminase (TGase), which induces the formation of cross-linked dimers and polymers, and that tacrine, indomethacin and deferoxamine, which have widely different chemical structures, attenuate the progression of symptoms of AD. This report evaluated the potential of a total of ten different pharmacological agents to inhibit TGase-induced cross-linking of A beta, including known TGase inhibitors (dansylcadaverine, spermine), non-steroidal anti-inflammatory drugs (indomethacin, meclofenamic acid, diflunisal, salicylic acid), monoamine oxidase inhibitors (tranylcypromine, phenelzine), an acetylcholinesterase inhibitor (tacrine), and an iron chelating agent (deferoxamine). All but one (salicylic acid) of these ten agents had an inhibitory effect on TGase-induced A beta cross-linking. These results suggest that inhibition of TGase-induced cross-linking of A beta is a potential pharmacologic target for the treatment of AD. A method is also presented for the determination of percent inhibition of TGase-induced A beta cross-linking based on the separated monomer, dimer and polymer bands on SDS-PAGE gels.