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Proteomics characterization of Coelomic epithelium (CE) of the starfish Marthasterias glacialis

ABSTRACT: Coelomic epithelium (CE), or coelothelium, is a mesodermal-derived tissue lining coelomic cavities and enveloping inner organs in all coelomates, including vertebrates, where it forms both peritoneum and pericardium, as well as pleura, in mammals. Due to its histogenetic potential, CE is considered a key-tissue characterized by unique features. At present, in adult echinoderms, CE is supposed to play fundamental key roles acting as: 1) source of stem/multipotent cells (undifferentiated coelomocytes), particularly during regenerative phenomena (Candia Carnevali et al., 2009; Candia Carnevali and Burighel, 2010; Hernroth et al., 2010; García-Arrarás et al., 2011). 2) haematopoietic tissue, being the direct source of circulating elements (differentiated coelomocytes) including immune cells, indispensable elements during the first repair events of clotting and defence against pathogens (Boassche and Jangoux, 1976; Munoz-Chapuli et al., 2005; Pinsino et al., 2007). Despite these indications, the histological approach alone, microscopic or ultrastructural, is not sufficient to fully attribute a haematopoietic role to the CE and even the presumptive production/release process of stem/pluripotent cells from coelothelium during regeneration need to be confirmed as well. As a matter of fact, it is still unclear if the intense proliferation detectable in regenerating CE is functional to the production of wandering coelomocytes necessary to restore the post-traumatic fluid loss or rather to produce truly regeneration-competent cells used as building blocks of the regenerating tissues. In order to overcome some of these issues and deeply understand the complex physiological role of this epithelial tissue in echinoderm biology, it is important to integrate morphological evidences with detailed molecular data. In particular, proteomics is becoming a powerful tool to deepen the problem of cell and tissue functions during both tissue homeostasis and regeneration by depicting the existing protein activities involved in the regrowth processes. However, proteomics investigations focussed on echinoderm CE are quite limited. Specific molecular information are due to the works by Gabre and co-workers (2015) and by Kim and co-workers (2018), which are addressed to the characterization of CE transcriptome from the clonal starfish Coscinasterias muricata and the blue bat starfish Patiria pectinifera, respectively. The analysis of the biological processes associated to the transcripts identified in these studies suggests the involvement of CE in the development of anatomical units. More recently the CE proteome of the common starfish Asterias rubens was also published (Sharlaimova et al., 2021). Despite these evidences, there is a great lack of integrated multidisciplinary studies that can contribute to unravel the complex roles exerted by this tissue both in regeneration and other physiological processes. In order to partially unravel this gap, the present work is addressed to analyse the CE of the starfish Marthasterias glacialis in both standard physiological (homeostatic) and regenerating conditions, by combining histological/ultrastructural analyses with appropriate proteomics approaches, the former to provide a detailed cell and tissue perspective, the latter to contribute a base-line molecular view of CE multifunctional implications. This integrated approach can significantly help to shed new light into the intriguing role of this apparently simple but fundamental tissue.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Marthasterias Glacialis (spiny Starfish)

TISSUE(S): Coelomic Fluid, Coelomic Epithelium

SUBMITTER: Sandra Anjo

LAB HEAD: Ana Varela Coelho

PROVIDER: PXD027009 | Pride | 2023-01-24

REPOSITORIES: Pride

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Project description:A critical step in regeneration is recreating the cellular identities and patterns of lost organs long after embryogenesis is complete. In plants, perpetual (indeterminate) organ growth occurs in apical stem cell niches, which have been shown to re-establish quickly when damaged or removed (1,2). Here we ask whether the machinery of perpetual organ growth, stem cell activity, is needed for the phase of regeneration that leads to replenishing lost cell identities and patterning, or, whether organ re-establishment enlists a wider group of pluripotent cells. We adapt a root tip regeneration system to Arabidopsis that permits us to assess the molecular and functional recovery of specific cell fates during organ regeneration. These results suggest a rapid restoration of missing cell fate and function in advance of the recovery of stem cell activity. Surprisingly, plants with mutations that fail to maintain stem cell activity were able to re-pattern their distal tip and re-specify lost cell fates. Thus, although stem cell activity is required to resume indeterminate growth (3), our results show it is not necessary for cell re-specification and patterning steps. This implies a regeneration mechanism that coordinates patterning of the whole organ, as in embryogenesis, but is initiated from different starting morphologies. 1. Feldman, L. J. Denovo Origin of Quiescent Center Regenerating Root Apices of Zea-Mays. Planta 128, 207-212 (1976). 2. Xu, J. et al. A molecular framework for plant regeneration. Science 311, 385-8 (2006). 3. Gordon, S. P. et al. Pattern formation during de novo assembly of the Arabidopsis shoot meristem. Development 134, 3539-48 (2007). We adapted root tip excision techniques to Arabidopsis, enabling us to perform microarray profiling of regenerating root tissue. Excisions were performed at 4 days post-germination (dpg) at a distance of 130 um from the root tip, resulting in the complete excision of QC, all surrounding stem cells along with several tiers of daughter cells, and the root cap, including all of the columella and most of the lateral root cap. The tip section and then approximately 70 um of regenerating tissue was recut at different time points post cutting. We sampled regenerating stumps at 0hrs, 5 hrs, 13 hrs, 22 hrs, and 7 days after the excision for microarray analysis (Methods). We also sampled root sections immediately above the zone competent to regenerate at 270 um to approximately 340 um. Experiment Overall Design: 30 samples with 4 or 3 replicates for each condition representing a time course of regenerating root stumps and including controls for root tips (regeneration endpoint) at 4 dpg and 8 dpg and a wounded set of samples representing root tissue at 270-340 mm from the root tip for non-regeneration control

Project description:A large number of rodent studies have supported the developmental origins of health and disease (DOHaD) hypothesis that interauterine undernutrition (IU) is a risk factor for non-communicable diseases. The effect of IU is considered to be induced thorough epigenetic programming in the fetal tissues. We have recently carried out global transcriptome expression and promoter DNA methylation analyses on the fetal mouse liver following maternal 50% food restriction (FR) during late gestation, and reported a list of fetal liver genes that were transcriptionally or epigenetically regulated by IU (Ogawa et al., 2014, Congenital Anomalies). We have already reported the genes that were regulated oppositely between maternal and fetal livers (Ogawa et al., 2014, Congenital Anomalies). Here, we considered that the fetal liver is a hematopoietic organ, and exposed nine-week-old male mice (C57BL/6J from Japan SLC, Hamamatsu, Japan; n=8 / group) to 50% FR (chow; CE-2, CLEA, Tokyo Japan) for two weeks and carried out global gene expression analysis on the whole blood by a method reported previously (Ogawa et al., 2014, Congenital Anomalies). Male C57BL/6J mice at 9 weeks (Japan SLC, Hamamatsu, Japan ; n=8 / group) were exposed to 50% FR (chow; CE-2, CLEA, Tokyo Japan) for 2 weeks. After 2 weeks, the mice were anesthetized with somunopentylÂ®(Kyoritsu eiyaku, Tokyo) at a dose of 0.5 mL/kg, and blood was taken from the right ventricle of the heart. Blood samples were immediately immersed in liquid nitrogen and stored at -80Â°C. All animal care and experimental procedures were approved by the Institutional Animal Care and Use Committee of Showa University, and caried out at animal facilities in Showa University. DNA Microarray analysis was carried out on 3 pools, respectively. Pool 1 included samples from animal No. 1-8 in each group (control and 50% FR). Pool 2 included samples from animal No. 1-4 in each group. Pool 3 included samples from animal No. 5-8 in each group.

Project description:A critical step in regeneration is recreating the cellular identities and patterns of lost organs long after embryogenesis is complete. In plants, perpetual (indeterminate) organ growth occurs in apical stem cell niches, which have been shown to re-establish quickly when damaged or removed (1,2). Here we ask whether the machinery of perpetual organ growth, stem cell activity, is needed for the phase of regeneration that leads to replenishing lost cell identities and patterning, or, whether organ re-establishment enlists a wider group of pluripotent cells. We adapt a root tip regeneration system to Arabidopsis that permits us to assess the molecular and functional recovery of specific cell fates during organ regeneration. These results suggest a rapid restoration of missing cell fate and function in advance of the recovery of stem cell activity. Surprisingly, plants with mutations that fail to maintain stem cell activity were able to re-pattern their distal tip and re-specify lost cell fates. Thus, although stem cell activity is required to resume indeterminate growth (3), our results show it is not necessary for cell re-specification and patterning steps. This implies a regeneration mechanism that coordinates patterning of the whole organ, as in embryogenesis, but is initiated from different starting morphologies. 1. Feldman, L. J. Denovo Origin of Quiescent Center Regenerating Root Apices of Zea-Mays. Planta 128, 207-212 (1976). 2. Xu, J. et al. A molecular framework for plant regeneration. Science 311, 385-8 (2006). 3. Gordon, S. P. et al. Pattern formation during de novo assembly of the Arabidopsis shoot meristem. Development 134, 3539-48 (2007). We adapted root tip excision techniques to Arabidopsis, enabling us to perform microarray profiling of regenerating root tissue. Excisions were performed at 4 days post-germination (dpg) at a distance of 130 um from the root tip, resulting in the complete excision of QC, all surrounding stem cells along with several tiers of daughter cells, and the root cap, including all of the columella and most of the lateral root cap. The tip section and then approximately 70 um of regenerating tissue was recut at different time points post cutting. We sampled regenerating stumps at 0hrs, 5 hrs, 13 hrs, 22 hrs, and 7 days after the excision for microarray analysis (Methods). We also sampled root sections immediately above the zone competent to regenerate at 270 um to approximately 340 um. Keywords: time course, development, root regeneration

			Action	DRS
	Database_S160709.fasta	Fasta
	S160709_A1E1.wiff	Wiff
	S160709_A1E1.wiff.scan	Wiff
	S160709_A1E1_SWATH.wiff	Wiff
	S160709_A1E1_SWATH.wiff.scan	Wiff

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Proteomics characterization of Coelomic epithelium (CE) of the starfish Marthasterias glacialis

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