Post-doctoral Fellow
kmsherra@u.washington.edu
I am interested in the cellular and mechanical basis for morphogenesis, a focus towards which I have spiraled inexorably if circuitously. I grew up in the Pacific Northwest and always expected to become a marine biologist, which I envisioned as studying whales and dolphins. I become side-tracked as an undergrad, receiving a B.A. in linguistics at Hampshire College in 1993, then rapidly came to my senses and remembered I wanted to study biology. While working through math and science pre-requisities for graduate school, I took invertebrate biology from Ken Lohman at UNC-Chapel Hill and discovered that small, squishy things that fit under a microscope were as bizarre as creatures from another planet and much more fascinating than whales. I went on to study the remnants of small squishy creatures, doing a master’s thesis in paleobiology with Peter Ward in the University of Washington’s geology department (1996-1998), in pursuit of which I spent much time in the Zoology department and at Friday Harbor Labs (FHL). I became fascinated by biomechanics, and thanks to courses with Tom Daniel and Garry Odell, my trajectory jumped definitively towards a mechanical approach to understanding biological function. I went on to study biomechanics with Mike LaBarbera at the University of Chicago, , where I worked on the fluid mechanics of suspension feeding in newly metamorphosed juvenile ascidians, and received my Ph.D. in 2003. I continued to work summers at FHL, and was there attracted to the Center for Cell Dynamics, where I have been a post-doc since 2005 (and where I can occasionally watch the orcas swim by). Inspired by Ray Keller’s opinion (see Keller et al. 2003) that what developmental biology needed was a biomechanical perspective, I set out to investigate endoderm invagination in ascidians . This entailed just moving my focus to a slightly smaller scale, and a little earlier in ontogeny; however it has also required that I educate myself in a whole new realm of cell and developmental biology. Fortunately, at this tiny center of my inward-spiraling trajectory I have found a realm vast enough to occupy and fascinate me for several lifetimes of study.
Vegetal view of ascidian embryo, 110-cell stage, early in gastrulation. Cell fates have largely been determined by the 32-cells stage: endoderm yellow, muscle orange, mesenchyme pink, notochord red, A-line neural blue, a-line neural purple, ectoderm cyan.
Figure 1. DIC timelapse invaginating Corella inflata embryo, in vegetal view. Ten endoderm cells which will invaginate outlined in black.
In contrast, the invagination at the start of ascidian gastrulation involves just 10 endoderm cells (each the size of a C. elegans embryo) in a 150 um embryo of c. 100 cells (Fig. 1); a few cell cycles later, a sheet of 80 cells neurulates and a mere 40 notochord cells converge and extend (Munro & Odell 2002, Munro et al. 2006). In all these cases, large cell size makes it possible to characterize the underlying cytoskeletal and adhesive dynamics at high resolution and to link individual cell shape changes unambiguously to global deformations of the embryo, while the low cell count facilitates making detailed computer models of the process. Several species of ascidian, such as Corella inflata, are transparent, making it possible to observe cell dynamics at high resolution with time lapse microscopy movie. Early cell fate specification, cell-autonomous morphogenetic behavior of explants, and the tolerance of intact embryos and explants to culture in seawater, make it possible to distinguish the contributions of cell-intrinsic behaviors from those that require adhesive coupling or other mechanical or biochemical interaction with neighbors. Three species genomes have been sequenced, facilitating the design of fluorescent probes and morpholino constructs, and these can be expressed or introduced by electroporation and/or microinjection. Because the ascidian genome is not duplicated, revealing the molecular underpinnings of specific behaviors is often simpler than in higher chordates. Thus, we can exploit the known genome sequence, the single-copy status of many genes, the early establishment of lineage-specific gene expression, and morpholino-based gene product knockdown screens to identify the proteins implicated in particular aspects of a morphogenetic mechanisms. Our current ascidian work focuses on invagination, but also positions us to study other classical morphogenetic events in these model chordate embryos.
To complement our experimental work Munro and Sherrard have created a computational simulation of morphogenesis in two dimensions, representing an entire cross-section of the embryo. This model is a direct outgrowth of Munro’s model of cell motility and convergent extension [LINK]. Previous models of tissue morphogenesis (e.g., Odell et al. 1981, Jacobsen et al. 1986, Hardin & Keller 1988, Clausi & Brodland 1993, Davidson 1995), though providing crucial tests of mechanical plausibility, have relied largely on continuum representations of cell mechanics and lack explicit reference to the underlying molecular machinery. We have developed a hybrid approach that combines a compartmental representation of the cell cortex with an agent-based model of cadherin-mediated adhesion (Fig. 2).
Figure 2. Computational model of invagination, representing two-dimensional cross-section of ascidian embryo (endoderm yellow, mesoderm orange, ectoderm red). Detail: the cortex is represented as a belt of viscous (dashpot symbols), elastic (zigzag symbols) and contractile elements and linkers (bound: pink; unbound: green) that bind and unbind from the cortex and each other stochastically and are transported by cortical flow. Different cell types can be endowed with distinct contractile, viscous, and elastic properties to mimic experimental observations (e.g., higher apical contractility on endoderm cells) or to explore the consequences of hypothetical manipulations. In addition, empirical observations, such as a feedback of contractility being downregulated in adhesively bound (i.e. basolateral) regions, can be built into the simulation. See also morphogenesis tutorial.
Two features distinguish our approach from all previous efforts: The first is a close correspondence between model building blocks and empirical observables at molecular and subcellular levels. The second is the complete absence of any top-down constraints on cell geometry or tissue organization. In our model, cell shapes, organization, and neighbor exchanges emerge solely from the local rules governing individual cortex parts. We can represent hypothetical local elevations in cortical tension, to correspond to data from kinematics and immunofluorescence observations of myosin activation (Fig. 3), and test whether our interpretations of invagination mechanism are feasible. We can also take a bottom-up approach, performing systematic searches of parameter space (using a simplified version of the model that subsumes adhesion kinetics into tension link to Sherrard's morphogenesis tutorial page) to discover, for a given starting geometry, the range of relative tension values in different cell types and locales that is capable of generating invaginations.
Figure 3. We saw elevated ser-19-phosphorylated myosin regulatory light chain on the endoderm apical surface (arrow) of control embryos in early gastrulation (A), contributing to invagination (B); the rho-kinase inhibitor Y-27632 greatly reduced this localization (arrow) (C), and prevented invagination (D). Computational simulations assuming strong apical contractility mimic the controls (E), and simulations with reduced apical contractility mimic Y-27632 treated embryos (F).
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