Dynamic and Mechanical Role of Spectrin in Membrane-Cytoskeleton Interplay

Informazioni relative al progetto

MechanoSpectrin

ID dell’accordo di sovvenzione: 796547

DOI

10.3030/796547

Progetto chiuso

Data della firma CE 26 Marzo 2018

Data di avvio 1 Giugno 2018

Data di completamento 31 Maggio 2020

Finanziato da

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Costo totale

€ 168 277,20

Contributo UE

€ 168 277,20

Coordinato da

IFOM-ISTITUTO FONDAZIONE DI ONCOLOGIA MOLECOLARE ETS
Italy

Periodic Reporting for period 1 - MechanoSpectrin (Dynamic and Mechanical Role of Spectrin in Membrane-CytoskeletonInterplay)

Periodo di rendicontazione: 2018-06-01 al 2020-05-31

Sintesi del contesto e degli obiettivi generali del progetto

The spectrin-based membrane skeleton is a major component of the cell cortex. While expressed by all metazoans, its dynamic interactions with the other cortex components, including the plasma membrane or the acto-myosin cytoskeleton, are poorly understood. Here, I investigated how spectrin re-organizes spatially and dynamically under the membrane during changes in cell mechanics. We found spectrin and acto-myosin to be spatially distinct but cooperating during mechanical challenges, such as cell adhesion and contraction, or compression, stretch and osmolarity fluctuations, creating a cohesive cortex supporting the plasma membrane. Actin territories control protrusions and contractile structures while spectrin territories concentrate in retractile zones and low-actin density/inter-contractile regions, acting as a fence that organize membrane trafficking events. During MECHANOSPECTRIN, I unveiled the existence of a dynamic interplay between acto-myosin and spectrin necessary to support a mesoscale organization of the lipid bilayer into spatially-confined cortical territories during cell mechanoresponse.
This line of research contributes to open new directions in the fields of morphodynamic regulation of cell shape and cell matrix interactions. Thus, it might lead to the identification of novel molecular target for cell signalling mechanisms linked to mechanoresponse during tissue/organ development or pathophysiological conditions with altered cell/tissue morphogenesis.

Lavoro eseguito dall’inizio del progetto fino alla fine del periodo coperto dalla relazione e principali risultati finora ottenuti

The results of MECHANOSPECTRIN show that, despite its well-known organization in erythrocytes and neurons, the spectrin-based membrane skeleton displays a remarkable complementarity with the actin cytoskeleton in mammalian cells of different origins. The project provided in-depth investigation of live specimens at high spatio-temporal resolution, while most of previous knowledge is extrapolated from ex-vivo analysis. For this purpose, the project made use of custom-developed devices: the cell stretching and cell compression devices were indeed patented previously by MSCA-IF grantee Dr. Qingsen Li at IFOM and were successfully applied to address these relevant mechanobiological questions. Moreover, spectrin organizations have been resolved by an innovative approach (Expansion Microscopy) that was fostered during a training visit at HHMI-Janelia Advanced Imaging Center. Briefly, spectrin is able to adopt a dual conformation in structural cells (i.e. fibroblasts): an erythroid-like triangular organization and a neuron-like periodic patter. Intriguingly, these two distinct conformations can co-exist within the same cell. In live specimens it was not possible to achieve this level of resolution, but spectrin condensation in response to either intrinsic or extrinsic mechanical stimuli suggested a functional relevance of the two conformations that depended on subcellular localization. Ultimately, mechanical stress within the spectrin meshwork was measured by a FRET-based tension sensor in response to osmolarity fluctuations. These measurements were complemented by the application of a membrane tension probe instead of the proposed optical tweezer-based assay, which represent the only deviation from the initial proposal. The results and developed novel techniques have been disseminated at various conferences and workshops, including public and scientific communication initiatives.

Progressi oltre lo stato dell’arte e potenziale impatto previsto (incluso l’impatto socioeconomico e le implicazioni sociali più ampie del progetto fino ad ora)

MECHANOSPECTRIN has achieved most of its objectives. Despite being a well-known player in cell mechanoprotection, particularly in erythrocytes and neurons, we found a novel role of spectrin in complementing acto-myosin cytoskeleton dynamics as well as organizing key membrane trafficking events. A manuscript describing these novelties has been submitted in a peer-reviewed journal (second revision). The spectrin-based membrane skeleton has been implicated in many pathophysiological conditions. The dynamic interplay with actin newly described by MECHANOSPECTRIN, shed lights on the mechano-dependent mechanisms potentially leading to such conditions. Beyond that, the original and innovative techniques designed for the project will be highly valuable to fulfil future scientific tasks aimed at describing regulation of cell shape and dynamic interactions between complementary molecular scaffolds.

Spectrin and actin defining distinct and complementary plasma membrane territories