Microsome-associated proteome modifications of Arabidopsis seedlings grown on board the International Space Station reveal the possible effect on plants of space stresses other than microgravity

11p.-2 fig.-6 tab.Growing plants in space for using them in bioregenerative life support systems during long-term human spaceflights needs improvement of our knowledge in how plants can adapt to space growth conditions. In a previous study performed on board the International Space Station (GENARA A experiment STS-132) we evaluate the global changes that microgravity can exert on the membrane proteome of Arabidopsis seedlings. Here we report additional data from this space experiment, taking advantage of the availability in the EMCS of a centrifuge to evaluate the effects of cues other than microgravity on the relative distribution of membrane proteins. Among the 1484 membrane proteins quantified, 227 proteins displayed no abundance differences between µ g and 1 g in space, while their abundances significantly differed between 1 g in space and 1 g on ground. A majority of these proteins (176) were over-represented in space samples and mainly belong to families corresponding to protein synthesis, degradation, transport, lipid metabolism, or ribosomal proteins. In the remaining set of 51 proteins that were under-represented in membranes, aquaporins and chloroplastic proteins are majority. These sets of proteins clearly appear as indicators of plant physiological processes affected in space by stressful factors others than microgravity.The authors would like to thank the National Aeronautics and Space Administration (NASA) who successfully performed the spaceflight experiment; they also thank the astronauts for performing the required tasks on board the ISS. We acknowledge the Norwegian User Support and Operations Center team (NUSOC) for the ground and space preparation of the GENARA-A experiment and we thank the European Aeronautic Defense and Space Company (Astrium EADS) for the design and building of the hardware. We also thank the European Space Agency (ESA) and the Centre National d’Etudes Spatiales(CNES) for their scientific and financial support.Peer reviewe

Plants in space: GRAVI-2 experiment

The Gravi-2 experiment was launched on flight SpaceX3 in April 2014 (http://lensesinspace.wordpress.com/) and was performed on-board the ISS in May 2014 with EMCS (European Modular Cultivation System).Gravity is considered to be an important environmental factor in the orientation of plant growth. In the case of roots, signal gravity is perceived by specialized cells located in the root apex and, so called statocytes. Even if it is well known that the movements of amyloplasts (located in the statocytes) induce a series of signalling pathways, the role of amyloplasts displacement is not clearly elucidated. The objective of the GRAVI-2 project is to study the impact of amyloplasts displacement on the calcium-dependant pathways and thus, better understanding the gravity perception in lentil roots. The Gravi-2 experiment was launched on flight SpaceX3 in April 2014 (http://lensesinspace.wordpress.com/) and was performed on-board the ISS in May 2014 with EMCS (European Modular Cultivation System). Lentil seeds were germinated on the International Space Station (ISS) in different situations: (1) continuously in microgravity conditions (e.g. 10-4 g), (2) 10-2 g during 8 hours after a growth period of 23 hours in microgravity, (3) 2g during 5 minutes after a growth period of around 31 hours in microgravity, and (4) 2g during 15 minutes after a growth period of around 31 hours in microgravity. The studies will be performed using complementary approaches including the analysis of amyloplast positioning, calcium localisation and the level of expression of genes

GRAVI-2 space experiment: investigating statoliths displacement and location effects on early stages of gravity perception pathways in plant roots

GRAVI-2 space experiment: investigating statoliths displacement and location effects on early stages of gravity perception pathways in plant roots. ASGSR meetin

Both gravistimulation onset and removal trigger an increase of cytoplasmic free calcium in statocytes of roots grown in microgravity

Gravity is a permanent environmental signal guiding plant growth and development. Gravity sensing in plants starts with the displacement of starch-filled plastids called statoliths, ultimately leading to auxin redistribution and organ curvature. While the involvement in gravity sensing of several actors such as calcium is known, the effect of statolith displacement on calcium changes remains enigmatic. Microgravity is a unique environmental condition offering the opportunity to decipher this link. In this study, roots of Brassica napus were grown aboard the International Space Station (ISS) either in microgravity or in a centrifuge simulating Earth gravity. The impact of short simulated gravity onset and removal was measured on statolith positioning and intracellular free calcium was assessed using pyroantimonate precipitates as cytosolic calcium markers. Our findings show that a ten-minute onset or removal of gravity induces very low statolith displacement, but which is, nevertheless, associated with an increase of the number of pyroantimonate precipitates. These results highlight that a change in the cytosolic calcium distribution is triggered in absence of a significant statolith displacement

Microgravity Induces Changes in Microsome-Associated Proteins of Arabidopsis Seedlings Grown on Board the International Space Station

International audienceThe ''GENARA A'' experiment was designed to monitor global changes in the proteome of membranes of Arabidopsis thaliana seedlings subjected to microgravity on board the International Space Station (ISS). For this purpose, 12-day-old seedlings were grown either in space, in the European Modular Cultivation System (EMCS) under microgravity or on a 1 g centrifuge, or on the ground. Proteins associated to membranes were selectively extracted from microsomes and identified and quantified through LC-MS-MS using a label-free method. Among the 1484 proteins identified and quantified in the 3 conditions mentioned above, 80 membrane-associated proteins were significantly more abundant in seedlings grown under microgravity in space than under 1 g (space and ground) and 69 were less abundant. Clustering of these proteins according to their predicted function indicates that proteins associated to auxin metabolism and trafficking were depleted in the microsomal fraction in mg space conditions, whereas proteins associated to stress responses, defence and metabolism were more abundant in mg than in 1 g indicating that microgravity is perceived by plants as a stressful environment. These results clearly indicate that a global membrane proteomics approach gives a snapshot of the cell status and its signaling activity in response to microgravity and highlight the major processes affected

Proteins specifically identified in one extraction fraction and common to the three gravity conditions (i.e. 1 g ground, 1 g space and µg space).

<p>Blue bars indicate the total number of proteins identified. Red bars show the number of proteins with at least one transmembrane segment and the green bars the number of proteins which are annotated as “membrane” in Gene Ontology (The number of proteins is indicated at the top of the bars).</p

Comparative distribution of under and over-represented proteins according to functional categories.

<p>Black bars = over-represented, Grey bars = under-represented.</p

Proteins specifically identified in one extraction fraction and common to the three gravity conditions (i.e. 1 g ground, 1 g space and µg space).

<p>Blue bars indicate the total number of proteins identified. Red bars show the number of proteins with at least one transmembrane segment and the green bars the number of proteins which are annotated as “membrane” in Gene Ontology (The number of proteins is indicated at the top of the bars).</p

Workflow of microsome preparation and sequential extraction of proteins with salts and detergents.

<p>¹ Volumes of buffer are expressed as µl per mg of initial fresh material used. * Centrifugation 100 000 g, 20 min; ** Centrifugation 15 000 g 15 min.</p