142 research outputs found
Interactions between nuclear lamins and their binding partners in EDMD fibroblasts
No full textLamins are components of the nuclear lamina and are divided In A and B-types, which Interact with proteins of the inner nuclear membrane like emerin. Mutations in emerin (X-linked) and A-type lamins (Autosomal Dominant) has been linked to the Emery-Dreifuss Muscular Dystrophy (EDMD), which conduced to the hypothesis that these two proteins might interact in the nucleus. I examined the interaction between A and B-type lamins with emerin using a panel of deletion mutants of lamin Bl and full-length lamins A, C and B1 in a yeast two-hybrid assay, where emerin interacted with all lamins and the preferred region of interaction was the globular tail domain of lamin Bl. Ectopic expression of tagged proteins in human dermal fibroblasts confirmed that emerin remains attached to the inner nuclear envelope through its association with lamin 81, as aggregation of tagged A-type lamins did not miss localize endogenous emerin or lamin Bl. In addition, methanol-acetone fixation showed higher number of cells presenting characteristic morphological abnormalities called "honeycombs". A-type lamins and their associated protein emerin co- localized in these structures. Lamin Bl depletion from the honeycombs was accompanied by depletion of nuclear pore complexes. In the honeycombs, A- type lamins segregated from the B-type lamins, forming homo-filaments. On the other hand, AD-EDMD cell lines showed a characteristic pattern as a high sub-population of cells presented nesprin 1 (amino-terminal) in stress fibres co- localizing with a-S-Actin fibres, which was enhanced by growth inhibition induced by serum starvation. Re-stimulation of fibroblasts by normal serum concentrations increased the appearance of honeycombs by up to 2.5 fold in the AD-EDMD cell, lines. Late passage cultures of AD-EDMD entered a senescence state reminiscent of the induced quiescence state induced by serum starvation. Finally, differential allelic expression was evidenced using a specific set of ARMS-primers in the cell lines studied, indicative of transcript imbalance, and bioinformatics analysis demonstrated the presence of SNPs in the coding region of the wild type LMNA gene. The results of these study confirm that lamins interact with emerin and suggest that the interacting region is the tail domain of lamins; honeycomb structures might have a biological meaning in patient cells; other proteins might be involved in EDMD, like nesprins; and heterozygosis is presented with transcript imbalance, which might have a negative impact in the correct assembly of the nuclear lamina
The Transcriptomic and Genomic Analysis of Lamin A/C Expression in the Colon and in Colorectal Cancer
No full textLamins A and C, also known as A-type lamins, are type V nuclear intermediate filament proteins which form an interlacing meshwork of filaments subjacent to the inner nuclear membrane termed the nuclear lamina. A-type lamins have been implicated in DNA replication, gene transcription regulation, apoptosis, regulation of growth promoters and nuclear migration. Traditionally, expression of A-type lamins has been associated with differentiated cells. As such, mutations in A-type lamins have been associated with a diverse range of genetic diseases, including premature ageing syndromes and with increased proliferation, especially in tumours.
In colorectal cancer, expression of A-type lamins, have been shown to impart an adverse prognosis. In order to understand the underlying biological processes responsible for this adverse outcome in patients with colorectal cancer, I sought to clarify the expression profile of A-type lamins and their binding partners in normal colonic/rectal mucosa, prior to investigating the expression of A-type lamins in colorectal cancers. I used fresh tissue specimens obtained from patients with colorectal cancer for my experiments. A unique finding was the expression of lamin A in the putative stem cell niche of colonic / rectal mucosal crypts.
Further studies in the form of a microarray analysis, revealed a very complex picture of up regulation involving various signalling cascades in the cancer samples expressing A-type lamins. There was no evidence to suggest a direct involvement of A-type lamins influencing the Wnt signalling cascade, however, direct involvement of other signalling cascades, such as the IGF signalling cascade, Shh signalling cascade and TGF-β signalling cascades were noted. These signalling cascades were known to influence the Wnt signalling cascades and hence could play a crucial role in the pathogenesis of colorectal cancers expressing A-type lamins.
In addition to these important signalling cascades, other key genes involved in apoptosis, growth promoters, cell adhesion, stem cell regulation, oncogenes and tumour suppression, were noted to have a unique expression profile in the cancer sample expressing A-type lamins, not observed in the cancer sample lacking A-type lamin expression. These observations were suggestive of A-type lamins having a diverse range of actions via, as yet, undefined pathways. It would appear that A-type lamins were imparting a more motile, less adherent phenotype with stem cell like features in colorectal cancers expressing A-type lamins. This could explain the observed poor prognosis of patients with colorectal cancers expressing A-type lamins.
Creatine kinase brain (CKB), was also identified as an additional, potential, prognostic indicator in the Duke’s B group of patients with colorectal cancer expressing A-type lamins. This potential marker, in conjunction with A-type lamin expression could be used to identify a sub group of Duke’s B patients at high risk. Whether adjuvant therapy in this group would help improve their long term survival is unknown since no study has been done to assess this
Lecture: Lamin Sanneh of Yale University delivers annual Cole Lecture
No full textIncludes descriptive metadata provided by producer in MP3 file: "Listen to historian and author Lamin Sanneh, the D. Willis James Professor of Missions and World Christianity and professor of history at Yale University, as he delivers the first of the 2006 Vanderbilt Cole Lectures on Oct. 19 at Benton Chapel. Sanneh, an important voice in inter-religious dialogue, was raised an orthodox Muslim before converting to Christianity. The Cole Lectures were established in 1892 by Col. E.W. Cole for 'the defense and advocacy of the Christian religion.'" The title of his talk is: Has Christianity outlived the Enlightenment
The effects of National culture and Geographic proximity on the innovation performance of Industry-University collaborations
No full textBackground: Innovation has been recognized by most firms as critical for the development
of novel or improved products, processes, and services to remain competitive. In light of the
importance of Industry-University collaborations (IUCs) towards enhancing the competitive
advantage of firms via innovation, it is vital to ensure successful outcomes of IUCs.
Objectives: The objectives of this paper are: (i) to determine whether national cultural values
impact the innovation performance of IUCs, (ii) to determine the effects of the differences in
national cultural values between the industry and university partners on the innovation performance of IUCs, and (iii) to determine the effects of geographical proximity on the innovation
performance of IUCs
Modelling The Socio-Demographic and Spatial Determinants of Undernutrition in Zambia: A Comparison of Full Bayesian with Penalised Structured Additive Regression
No full textFull Bayes Markov Chain Multi Carlo (MCMC) and Penalised Structured Additive Regression (STAR) models were compared for an underenutrition (measured as stunting) study in Zambia. Spatial correlated effects were specified as a Markov random field prior, continuous covariates were modelled using Bayesian penalised splines and diffused priors were assigned to fixed effects.
A Bayesian Structured Additive Regression Model was developed for the Zambia data. Model estimation and inference was based on both fully Bayesian MCMC and Empirical Bayes (based on mixed method methodology). In a frequentist setting, EB inference is closely related to penalized likelihood estimation. (Approximate) restricted maximum likelihood are used to estimate Variance components which correspond to inverse smoothing parameters. Both inference procedures were then compared based on the results from the Zambia study and were found to be very similar.
The results indicate spatial variations in stunting among the districts of Zambia. Continuous covariates Age and BMI have a significant effect on stunting. There is also significant difference among the factors of all categorical variables except for mother's employment status where no difference was found in stunting between children of employed and unemployed mothers.
Keywords: MCMC, STAR, Full Bayes, Empirical Bayes, spatial, continuous, categorical, model estimation, inference
The effects of National culture and Geographic proximity on the innovation performance of Industry-University collaborations
No full textBackground: Innovation has been recognized by most firms as critical for the development
of novel or improved products, processes, and services to remain competitive. In light of the
importance of Industry-University collaborations (IUCs) towards enhancing the competitive
advantage of firms via innovation, it is vital to ensure successful outcomes of IUCs.
Objectives: The objectives of this paper are: (i) to determine whether national cultural values
impact the innovation performance of IUCs, (ii) to determine the effects of the differences in
national cultural values between the industry and university partners on the innovation performance of IUCs, and (iii) to determine the effects of geographical proximity on the innovation
performance of IUCs
Modelling The Socio-Demographic and Spatial Determinants of Undernutrition in Zambia: A Comparison of Full Bayesian with Penalised Structured Additive Regression
No full textFull Bayes Markov Chain Multi Carlo (MCMC) and Penalised Structured Additive Regression (STAR) models were compared for an underenutrition (measured as stunting) study in Zambia. Spatial correlated effects were specified as a Markov random field prior, continuous covariates were modelled using Bayesian penalised splines and diffused priors were assigned to fixed effects.
A Bayesian Structured Additive Regression Model was developed for the Zambia data. Model estimation and inference was based on both fully Bayesian MCMC and Empirical Bayes (based on mixed method methodology). In a frequentist setting, EB inference is closely related to penalized likelihood estimation. (Approximate) restricted maximum likelihood are used to estimate Variance components which correspond to inverse smoothing parameters. Both inference procedures were then compared based on the results from the Zambia study and were found to be very similar.
The results indicate spatial variations in stunting among the districts of Zambia. Continuous covariates Age and BMI have a significant effect on stunting. There is also significant difference among the factors of all categorical variables except for mother's employment status where no difference was found in stunting between children of employed and unemployed mothers.
Keywords: MCMC, STAR, Full Bayes, Empirical Bayes, spatial, continuous, categorical, model estimation, inference
Repo-Man/PP1 SUMOylation mediates binding to lamin A and Serine 22 dephosphorylation
No full textData accessibility. The microscopy data are available from the corre- sponding author upon request and will be released via Figshare. Electronic supplementary material is available online at https://doi.org/10.6084/m9.figshare.c.5912878.ORCiD: Paola Vagnarelli, 0000-0002-0000-2271Copyright © 2022 The Authors. Lamin A phosphorylation/de-phosphorylation is an important process during cells division as it allows for nuclear envelope (NE) disassembly at mitotic entry and its re-assembly during mitotic exit. Several kinases have been identified as responsible for these phosphorylations, but no protein phosphatase has been implicated in their reversal. One of the mitotic phosphosites in lamin A responsible for its dynamic behaviour is serine 22 (S22) which is de-phosphorylated during mitotic exit. Recent evidence has also linked the nuclear pool of lamin A S22ph in interphase to gene expression regulation. Previous work suggested that the phosphatase responsible for lamin A S22 de-phosphorylation is chromatin bound and interacts with lamin A via SUMO-SIM motives. We have previously reported that Repo-Man/protein phosphatase 1 (PP1) is a chromatin-associated phosphatase that regulates NE reformation. Here we propose that Repo-Man/PP1 phosphatase mediates lamin A S22 de-phosphorylation. We indeed show that depletion of Repo-Man leads to NE defects, causes hyperphosphorylation of lamin A S22 that can be rescued by a wild-type but not a SUMOylation-deficient mutant. Lamin A and Repo-Man interact in vivo and in vitro, and the interaction is mediated by SUMOylation. Moreover, the localization of Repo-Man/PP1 to the chromatin is essential for lamin A S22 de-phosphorylation.Wellcome Trust Investigator award 3 210742/Z/18/Z to Paola Vagnarelli
The role of emerin and LEM domain proteins in nuclear envelope assembly and cytoskeleton organisation
No full textThe nuclear envelope (NE) plays a fundamental role in the cell by separating nuclear from cytoplasmic activities, and mutations in NE proteins have been associated with a diverse array of diseases. In the present study the Xenopus cell-free system was used to investigate the function of the inner nuclear membrane protein, emerin, which is associated with the Emery-Dreifuss muscular dystrophy (X-EDMD).Initially, the order and dynamics of NE assembly in Xenopus egg extracts have been investigated. Using a panel of antibodies it was shown that NE assembly proceeds by the ordered recruitment of two membrane populations, Nuclear Envelope Precursor vesicles -A and -B (NEP-A and NEP-B), to chromatin. As shown by immunofluorescence NEP-B vesicles, together with nucleoporins (Nups), appear first around chromatin at about ten minutes after initiation of NE assembly while NEP-A vesicles appear at a later stage, at about twenty minutes. To investigate the role of different emerin domains in this process, four human emerin peptides consisting of amino acids (aa) 1-70, 1-176, 1-220 and 73-180 were added individually to Xenopus nuclear assembly reactions at different concentrations and the effect on nuclear vesicle recruitment and NPC formation was monitored. Immunofluorescence analysis showed that peptides containing the LEM domain of emerin interfere with a correct NE assembly by inhibiting chromatin decondensation and recruitment of membranes to chromatin. This inhibitory effect was shown to be exerted mainly on NEP-A membranes and on Nup62 and Nupl53. By the use of two antibodies, raised against the LEM domain of human emerin and LAP2ß, two proteins of 30 and 36 kD, respectively, were identified in Xenopus. Both proteins were shown to reside in the NEP-A membrane population providing an explanation for the preferential inhibition of NEP-A recruitment to chromatin by exogenously added LEM domain containing emerin peptides. To further investigate whether the domain specific inhibitory effects of emerin on nuclear assembly correlate with specific interacting proteins, co-precipitation experiments were performed to identify emerin binding proteins in the Xenopus cytosol. From these experiments ß -tubulin was identified as a protein able to interact with emerin peptides 1-70 and 73-180. Staining of X-EDMD cells, which lack emerin, with a ß -tubulin antibody revealed no alterations in the organisation of the microtubule (MT) network. The most prominent effect of emerin mutations regarding MTs was the position of the Microtubule Organising Centre (MTOC) relative to the NE. Staining for the centrosomal protein pericentrin revealed a mis-localisation of the MTOC away from the NE in X-EDMD cell lines at distances at least double compared to control cells
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Lamin A/C modulates spatial organization and function of the Hsp70 gene locus via nuclear myosin I.
No full textThe structure-function relationship of the nucleus is tightly regulated, especially during heat shock. Typically, heat shock activates molecular chaperones that prevent protein misfolding and preserve genome integrity. However, the molecular mechanisms that regulate nuclear structure-function relationships during heat shock remain unclear. Here, we show that lamin A and C (hereafter lamin A/C; both lamin A and C are encoded by LMNA) are required for heat-shock-mediated transcriptional induction of the Hsp70 gene locus (HSPA genes). Interestingly, lamin A/C regulates redistribution of nuclear myosin I (NM1) into the nucleus upon heat shock, and depletion of either lamin A/C or NM1 abrogates heat-shock-induced repositioning of Hsp70 gene locus away from the nuclear envelope. Lamins and NM1 also regulate spatial positioning of the SC35 (also known as SRSF2) speckles - important nuclear landmarks that modulates Hsp70 gene locus expression upon heat shock. This suggests an intricate crosstalk between nuclear lamins, NM1 and SC35 organization in modulating transcriptional responses of the Hsp70 gene locus during heat shock. Taken together, this study unravels a novel role for lamin A/C in the regulation of the spatial dynamics and function of the Hsp70 gene locus upon heat shock, via the nuclear motor protein NM1.This article has an associated First Person interview with the first author of the paper
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