Lens epithelial cell apoptosis is an early event in the development of UVB-induced cataract

Li, Wancheng; Spector, Abraham

doi:10.1016/0891-5849(96)02050-3

Cited by 142 publications

(138 citation statements)

References 60 publications

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“…DNA fragmentation analyses were conducted as previously described [21,22]. Control and okadaic acid-treated N/N1003 cells (2ϫ10 6 cells per sample) were collected and washed with NaCl/P i twice.…”

Section: Methodsmentioning

confidence: 99%

Okadaic acid‐induced lens epithelial cell apoptosis requires inhibition of phosphatase‐1 and is associated with induction of gene expression including p53 and bax

Fass

Huizar

et al. 1998

European Journal of Biochemistry

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It is well established that phosphorylation and dephosphorylation are key cellular events which regulate important metabolic activities such as gene expression, cell cycle progression, and apoptosis. The polyether fatty acid, okadaic acid has been shown previously to activate apoptosis in a variety of cell lines. Although this marine sponge toxin is known to inhibit protein phosphatase (PP)-2A and PP-1, it is not certain in most cases whether inhibition of PP-1 or PP-2A is necessary to activate apoptosis. Furthermore, it is not clear how inhibition of these phosphatases leads to apoptosis. Here we present evidence that inhibition of PP-2A by okadaic acid does not activate apoptosis in the lens system. However, when PP-1 is inhibited by okadaic acid, rabbit lens epithelial cells undergo rapid apoptosis. Associated with this process is the several-fold up-regulation of the tumor suppressor gene p53 and the pro-apoptotic gene bax at both mRNA and protein levels. Analyses of the temporal pattern of expression of the two genes reveal that the up-regulation is maximized in a few hours after treatment with okadaic acid, when the majority of the treated cells become committed to apoptosis. A brief treatment of the cells with a protein synthesis inhibitor can abolish okadaic acid-induced up-regulation of both P53 and Bax proteins. Concomitant with this inhibition, okadaic acid-induced apoptosis is also temporarily blocked. These results suggest that okadaic acid-induced expression of p53, bax, and other genes are necessary for the activation of the apoptotic programs in lens systems.Keywords : apoptosis; p53 ; bax ; phosphatase-1; lens.Apoptosis is a distinct form of cell death regulated by internal genetic programs [1,2]. The morphological changes associated with apoptosis include blebbing of cytoplasmic membranes, condensation of nucleoplasm and cytoplasm, and fragmentation of the dying cell into apoptotic bodies which are phagocytosed by neighboring cells [3,4]. The biochemical hallmarks of apoptosis are marked by DNA fragmentation into nucleosomal fragments [4,5], activation of the interleukin-1β-converting enzyme family of proteases [6], and cleavage of various substrates of that family of proteases such as poly(ADPribose) polymerase [7,8] and nuclear lamin [9].Apoptosis normally occurs as a physiological process during certain stages of animal development or during tissue homeostasis in an adult organism [10,11]. It can also be triggered by various external signals such as hormone [4], γ-irradiation [12] and withdrawal of growth factors [13] and other factors (reviewed in [2]). Normal physiological apoptosis helps to remove

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Section: Methodsmentioning

confidence: 99%

Okadaic acid‐induced lens epithelial cell apoptosis requires inhibition of phosphatase‐1 and is associated with induction of gene expression including p53 and bax

Fass

Huizar

et al. 1998

European Journal of Biochemistry

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“…For analysis of the response to okadaic acid and calyculin A, the HLE cells with or without p53 silenced were treated with 0.01% DMSO (control), 20-75 nM okadaic acid or 1-6 nM calyculin A for 12 h; the viability of these samples were analysed with cell flow cytometry, Hoechst staining and DNA fragmentation as described previously (Li and Spector, 1996;Li et al, 1998Li et al, , 2001aLi et al, , 2005.…”

Section: Transfection Of P53à/à Mlecs and Pc-3 Cellsmentioning

confidence: 99%

Protein serine/threonine phosphatase-1 dephosphorylates p53 at Ser-15 and Ser-37 to modulate its transcriptional and apoptotic activities

et al. 2006

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We have previously demonstrated that the serine/threonine protein phosphatase-1 (PP-1) plays an important role in promoting cell survival. However, the molecular mechanisms by which PP-1 promotes survival remain largely unknown. In the present study, we provide evidence to show that PP-1 can directly dephosphorylate a master regulator of apoptosis, p53, to negatively modulate its transcriptional and apoptotic activities, and thus to promote cell survival. As a transcriptional factor, the function of p53 can be greatly regulated by phosphorylation and dephosphorylation. While the kinases responsible for phosphorylation of the 17 serine/threonine sites have been identified, the dephosphorylation of these sites remains largely unknown. In the present study, we demonstrate that PP-1 can dephosphorylate p53 at Ser-15 and Ser-37 through co-immunoprecipitation, in vitro and in vivo dephosphorylation assays, overexpression and silence of the gene encoding the catalytic subunit for PP-1. We further show that mutations mimicking constitutive dephosphorylation or phosphorylation of p53 at these sites attenuate or enhance its transcriptional activity, respectively. As a result of the changed p53 activity, expression of the downstream apoptosis-related genes such as bcl-2 and bax is accordingly altered and the apoptotic events are either largely abrogated or enhanced. Thus, our results demonstrate that PP-1 directly dephosphorylates p53, and dephosphorylation of p53 has as important impact on its functions as phosphorylation does. In addition, our results reveal that one of the molecular mechanisms by which PP-1 promotes cell survival is to dephosphorylate p53, and thus negatively regulate p53-dependent death pathway.

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“…[1][2][3] In lens system, our previous work [4][5][6] has suggested that induced lens epithelial cell apoptosis appears to be a common cellular mechanism mediating stress-induced, noncongenital cataractogenesis. Subsequently, several laboratories have reported that lens epithelial cell apoptosis is indeed actively involved in lens pathogenesis as demonstrated from in vivo animal model studies.…”

Section: Introductionmentioning

confidence: 99%

Human αA- and αB-crystallins bind to Bax and Bcl-XS to sequester their translocation during staurosporine-induced apoptosis

et al. 2004

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aA-and aB-crystallins are distinct antiapoptotic regulators. Regarding the antiapoptotic mechanisms, we have recently demonstrated that aB-crystallin interacts with the procaspase-3 and partially processed procaspase-3 to repress caspase-3 activation. Here, we demonstrate that human aA-and aBcrystallins prevent staurosporine-induced apoptosis through interactions with members of the Bcl-2 family. Using GST pulldown assays and coimmunoprecipitations, we demonstrated that a-crystallins bind to Bax and Bcl-X S both in vitro and in vivo. Human aA-and aB-crystallins display similar affinity to both proapoptotic regulators, and so are true with their antiapoptotic ability tested in human lens epithelial cells, human retina pigment epithelial cells (ARPE-19) and rat embryonic myocardium cells (H9c2) under treatment of staurosporine, etoposide or sorbitol. Two prominent mutants, R116C in aA-crystallin and R120G, in aB-crystallin display much weaker affinity to Bax and Bcl-X S . Through the interaction, a-crystallins prevent the translocation of Bax and Bcl-X S from cytosol into mitochondria during staurosporine-induced apoptosis. As a result, a-crystallins preserve the integrity of mitochondria, restrict release of cytochrome c, repress activation of caspase-3 and block degradation of PARP. Thus, our results demonstrate a novel antiapoptotic mechanism for a-crystallins. Keywords: aA-crystallin; R116C; aB-crystallin; R120G; Bax; Bcl-X S Abbreviations: a-crystallin, alpha-crystallin; GFP, green fluorescence protein; HaA, human aA-crystallin; HaB, human aB-crystallin; GFP-HaA, green fluorescence and human aAcrystallin fusion protein; GFP-HaB, green fluorescence and human aB-crystallin fusion protein; MEM, Eagle's minimal essential medium; PAGE, polyacrylamide gel electrophoresis; PBS, phosphate-buffered saline; PMSF, phenylmethylsufonyl fluoride; HLE, human lens epithelial cells; SDS, sodium dodecylsulfate; TBS, tris-buffered saline; TBS-T, tris-buffered saline with tween-20.

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Lens epithelial cell apoptosis is an early event in the development of UVB-induced cataract

Cited by 142 publications

References 60 publications

Okadaic acid‐induced lens epithelial cell apoptosis requires inhibition of phosphatase‐1 and is associated with induction of gene expression including p53 and bax

Okadaic acid‐induced lens epithelial cell apoptosis requires inhibition of phosphatase‐1 and is associated with induction of gene expression including p53 and bax

Protein serine/threonine phosphatase-1 dephosphorylates p53 at Ser-15 and Ser-37 to modulate its transcriptional and apoptotic activities

Human αA- and αB-crystallins bind to Bax and Bcl-XS to sequester their translocation during staurosporine-induced apoptosis

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