Purushothaman, S. | Search bioRxiv

bioRxiv2021

Sonic hedgehog is essential for proximal-distal outgrowth of the limb bud in salamanders

The developing forelimb has been a foundational model to understand how specified progenitor cells integrate genetic information to produce the tetrapod limb bauplan (1, 2). Although the reigning hypothesis is that all tetrapods develop limbs in a similar manner, recent work suggests that urodeles have evolved a derived mode of limb development (3-5). Here we demonstrate through pharmacological and genetic inactivation of Sonic hedgehog (Shh) signaling in axolotls that Shh directs expansion and survival of limb progenitor cells in addition to patterning the limb across the proximodistal and antero-posterior axis. In contrast to inactivation of Shh in mouse or chick embryos where a humerus, radius and single digit develop (6-9), Shh crispant axolotls completely lack forelimbs. In rescuing limb development by implanting SHH-N protein beads into the nascent limb field of Shh-crispants, we show that the limb field is specified in the absence of Shh and that hedgehog pathway activation is required to initiate proximodistal outgrowth. When the derived nature of salamander limb development is placed in a phylogenetic context, it generates a new hypothesis where the ability to regenerate an entire tetrapod limb may have evolved uniquely among urodeles. TeaserShh is essential for salamander limb development

developmental biology↗

bioRxiv2020

Recurrent Dynamics of Rupture Transitions of Single Giant Vesicles at Solid Surfaces

Single giant vesicles (GVs) rupture spontaneously from their salt-laden suspension onto solid surfaces. At hydrophilic surfaces, they rupture via a recurrent burst-heal dynamics: during burst, single pores nucleate at the contact boundary of the adhering vesicles facilitating asymmetric spreading and producing a "heart" shaped membrane patch. During the healing phase, the competing pore closure produces a daughter vesicle. At hydrophobic surfaces, by contrast, the GVs rupture via a distinctly different, yet recurrent, bouncing ball rhythm: Rendered tense by the substrate interactions, GVs porate and spread monomolecular layer on the hydrophobic surface in a symmetric manner. Here too, the competition from pore closure produces a daughter vesicle, which re-engages with the substrate. In both cases, the pattern of burst-reseal events repeats multiple times splashing and spreading the vesicular fragments as bilayer patches at the solid surface in a pulsatory manner. These remarkable recurrent dynamics arise not because of the elastic properties of the solid surface but because the competition between membrane spreading and pore healing, prompted by the surface-energy dependent adhesion, determine the course of the topological transition. STATEMENT OF SIGNIFICANCEGiant lipid vesicles adhering to a solid surface experience strong mechanical stresses. The contacting membrane segment loses thermal fluctuations and accumulates mechanical tension, the equilibration of which can give rise to global shape changes, lipid phase separation, and traction forces. Beyond a threshold tension, vesicles porate, unravel, and spread. Here, we find that a competition from pore-healing can make rupture iterative, rather than a single all-or-nothing event. During burst, single pores expand, spreading a lipid bilayer on the hydrophilic surface and a monolayer on the hydrophobic one. During heal, pore-healing can produce daughter vesicles. This burst-reseal event reiterates "splashing" portions of single vesicles at the solid surface and "bouncing" the remainder as a secondary vesicle in multiple steps.

biophysics↗