FBN-1, a fibrillin-related protein, is required for resistance of the epidermis to mechanical deformation during C. elegans embryogenesis

Abstract

During development, biomechanical forces contour the body and provide shape to internal organs. Using genetic and molecular approaches in combination with a FRET-based tension sensor, we characterized a pulling force exerted by the elongating pharynx (foregut) on the anterior epidermis during C. elegans embryogenesis. Resistance of the epidermis to this force and to actomyosin-based circumferential constricting forces is mediated by FBN-1, a ZP domain protein related to vertebrate fibrillins. fbn-1 was required specifically within the epidermis and FBN-1 was expressed in epidermal cells and secreted to the apical surface as a putative component of the embryonic sheath. Tiling array studies indicated that fbn-1 mRNA processing requires the conserved alternative splicing factor MEC-8/RBPMS. The conserved SYM-3/FAM102A and SYM-4/WDR44 proteins, which are linked to protein trafficking, function as additional components of this network. Our studies demonstrate the importance of the apical extracellular matrix in preventing mechanical deformation of the epidermis during development.DOI: http://dx.doi.org/10.7554/eLife.06565.001

FRET-based Molecular Tension Microscopy

“…This result suggests that the intracellular environment may affect FRET. Interestingly, tension-insensitive FRET standards inserted into b-spectrin appeared to exhibit similar FRET to that of their host-protein-free counterparts previously measured in other conditions [23,46]. Therefore, a reasonable possibility is that protein-bound and free control sensors may effectively be under different mechanical loads in different cellular localizations, despite the lack of protein binding sites.…”

“…The terminally-tagged a-actinin sensor was insensitive to Focal Adhesion recruitment and myosin activity inhibition [40] and to subcellular localization in adhesion-confined cultured cells [41]. Tension-insensitive standards within b-spectrin also showed no FRET change in C. elegans after axotomy, and across tissues during morphogenesis [23,46]. Finally, sensor-alone controls appeared to exhibit insignificant FRET changes in cultured cells through time, under thrombin treatment, osmotic shock [11] and fluid shear flow [38].…”

“…For instance, terminally-tagged sensor in vinculin appeared to be insignificantly sensitive to substrate biochemistry, Focal Adhesion assembly, disassembly and position within the cell [10], to laser ablation targeting actin fibers [35], to myosin pharmacological perturbation [21,45] and to secretagogue stimulation in chromaffin cells [24]. Similarly, terminally-tagged sensors in b-spectrin were not sensitive to embryonic tissue localization in C. elegans [46] and to cell migration and cytoskeleton drugs in cultured cells [12]. In E-cadherin, terminaly-tagged sensor was insensitive to EGF stimulation in epithelial cells [47], and to the expression of a DN Rac and to its localization in migrating border cell clusters in Drosophila [29].…”

“…Similarly, differential FRET in distinct subcellular or tissue regions during spontaneous cell or animal activity may not be accounted for global measurement drift. For instance, vinculin is under higher tension in Focal Adhesions of protruding cell regions than it is in retracting cell regions in cultured migrating cells [10] and generally exhibit temporal and spatial variations across Focal Adhesions population [35,45], E-cadherin exhibits a tension differential from front to back border cell cluster in Drosophila [29], a-actinin is under higher tension in growing Focal Adhesions that around [40], and b-spectrin exhibit differential tension across tissues during C. elegans embryonic development [46].…”

“…Finally, tension in b-spectrin of C. elegans epidermis appeared to depend on the contact of the neighboring pharynx during embryo morphogenesis [46].…”