[Show abstract][Hide abstract] ABSTRACT: Lamin A (LaA) is a component of the nuclear lamina, an intermediate filament meshwork that underlies the inner nuclear membrane (INM) of the nuclear envelope (NE). Newly synthesized prelamin A (PreA) undergoes extensive processing involving C-terminal farnesylation followed by proteolysis yielding non-farnesylated mature lamin A. Different inhibitors of these processing events are currently used therapeutically. Hutchinson-Gilford Progeria Syndrome (HGPS) is most commonly caused by mutations leading to an accumulation of a farnesylated LaA isoform, prompting a clinical trial using farnesyltransferase inhibitors (FTI) to reduce this modification. At therapeutic levels, HIV protease inhibitors (PI) can unexpectedly inhibit the final processing step in PreA maturation. We have examined the dynamics of LaA processing and associated cellular effects during PI or FTI treatment and following inhibitor washout. While PI reversibility was rapid, with respect to both LaA maturation and associated cellular phenotype, recovery from FTI treatment was more gradual. FTI reversibility is influenced by both cell type and rate of proliferation. These results suggest a less static lamin network than has previously been observed.
[Show abstract][Hide abstract] ABSTRACT: Sun-2 and Nesprin-3 are present in abnormal cytoplasmic aggregates during mitosis and early G1 during Lop treatment. A 48hr treatment of Saos-2 cells with Lop led to the accumulation of Sun 2 and Nesprin-3 at LaA/C immunoreactive cytoplasmic aggregates in metaphase (upper panels) and early G1 (lower panels). DNA is labeled by Hoechst dye in blue. Bar, 10µm.
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[Show abstract][Hide abstract] ABSTRACT: Inhibition of protein synthesis does not impair PreA processing following PI washout. Saos-2 cells were treated with cyclohexamide at the time of Lop washout. PreA levels dramatically disappear by 3 hrs following washout. Percent of mature LaA listed below.
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[Show abstract][Hide abstract] ABSTRACT: In HGPS cells, HDJ-2 exhibits prolonged maturation following FTI washout. (A) An anti-HA immunoblot of extracts from WT human fibroblasts expressing exogenous HA-progerin were either treated with DMSO (control) or FTI-277 for 48 hrs immediately following transfection. The FTI-277 treated progerin migrates more slowly. (B) In HGPS cells treated with FTI-277 for 96hrs, HDJ-2 is incompletely processed by 24hrs following FTI-277. Percent of mature HDJ-2 listed above.
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[Show abstract][Hide abstract] ABSTRACT: Multiple FTIs failed to permit rapid processivity of PreA following washout. As detected by anti-LaA immunoblots of Saos-2 cell lysates, PreA was refractory to processing following washout of 10µm L-744, 832 or 1µm BMS-214662. Percent of mature LaA listed below.
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[Show abstract][Hide abstract] ABSTRACT: Endochondral ossification is the process of skeletal bone growth via the formation of a cartilage template that subsequently undergoes mineralization to form trabecular bone. Genetic mutations affecting the proliferation or differentiation of chondrocytes result in skeletal abnormalities. Activating transcription factor-2 (ATF-2) modulates expression of cell cycle regulatory genes in chondrocytes, and mutation of ATF-2 results in a dwarfed phenotype. Here we investigate the regulatory role that ATF-2 plays in expression of the pocket proteins, cell cycle regulators important in cellular proliferation and differentiation. The spatial and temporal pattern of pocket protein expression was identified in wild type and mutant growth plates. Expression of retinoblastoma (pRb) mRNA and protein were decreased in ATF-2 mutant primary chondrocytes. pRb mRNA expression was coordinated with chondrogenic differentiation and cell cycle exit in ATDC5 cells. Type X collagen immunohistochemistry was performed to visualize a delay in differentiation in response to loss of ATF-2 signaling. Chondrocyte proliferation was also affected by loss of ATF-2. These studies suggest pRb plays a role in chondrocyte proliferation, differentiation and growth plate development by modulating cell cycle progression. ATF-2 regulates expression of pRb within the developing growth plate, contributing to the skeletal phenotype of ATF-2 mutant mice through the regulation of chondrocyte proliferation and differentiation.
Full-text · Article · Sep 2008 · Mechanisms of development