Associate Member, Institute for Stem Cell Biology and Regenerative Medicine (2013 - Present)
Honors & Awards
Predoctoral Fellowship for Science and Engineering, Department of Defense (1993-1996)
Predoctoral Fellowship, American Heart Association (1998-2001)
Postdoctoral Fellowship, American Heart Association (2006)
Genentech Foundation Fellow, Life Sciences Research Foundation (2007-2009)
Mentored Research Scientist Development Award (K01), NIH/NIDDK (2010-2015)
Gabilan Faculty Fellow, Stanford (2013)
Postdoctoral Scholar, UC Berkeley, Stem Cell Biology
Ph.D., UCSF, Biomedical Sciences / Cell Biology (2001)
B.A., Harvard University, Biochemistry (1992)
Current Research and Scholarly Interests
Animals live in dynamic environments where external conditions vary at cyclic or irregular intervals. When faced with environmental change, an individual’s physiological fitness requires that its organ systems functionally adapt. One type of organ adaptation occurs through tissue growth and shrinkage, tuning an organ's functional capacity to meet variable levels of physiologic demand. An “economy of nature", adaptive remodeling breaks the allometry of the body plan that was established during development. Unlike embryonic growth, adult organ remodeling is reversible and repeatable, suggesting that it occurs through different mechanisms. Stem cells are key players in at least some of these mechanisms. But, basic questions remain largely unanswered: How do stem cells sense different levels of functional demand? How do they help translate this information into appropriate changes in organ size?
We have developed the Drosophila midgut as a simple invertebrate model to uncover the rules that govern adaptive remodeling. In adult flies, the midgut is a stem cell-based organ analogous to the vertebrate small intestine. We have found that when dietary load increases, midgut stem cells activate a reversible growth program that increases total intestinal cell number and digestive capacity. The midgut is a uniquely tractable model to study adaptive growth; not only can gene expression be manipulated and lineages traced at single-cell and whole-tissue levels, but complete population counts of all cell types are possible. Our goal is to understand how this nutrient-driven mechanism regulates stem cell behavior for lifelong optimization of organ form and function.
Graduate and Fellowship Programs
Beyond the Niche: Tissue-Level Coordination of Stem Cell Dynamics
ANNUAL REVIEW OF CELL AND DEVELOPMENTAL BIOLOGY, VOL 29
2013; 29: 107-136
Adult animals rely on populations of stem cells to ensure organ function throughout their lifetime. Stem cells are governed by signals from stem cell niches, and much is known about how single niches promote stemness and direct stem cell behavior. However, most organs contain a multitude of stem cell-niche units, which are often distributed across the entire expanse of the tissue. Beyond the biology of individual stem cell-niche interactions, the next challenge is to uncover the tissue-level processes that orchestrate spatial control of stem-based renewal, repair, and remodeling throughout a whole organ. Here we examine what is known about higher order mechanisms for interniche coordination in epithelial organs, whose simple geometry offers a promising entry point for understanding the regulation of niche number, distribution, and activity. We also consider the potential existence of stem cell territories and how tissue architecture may influence niche coordination.
View details for DOI 10.1146/annurev-cellbio-101512-122319
View details for Web of Science ID 000326689300006
View details for PubMedID 23937350
Altered Modes of Stem Cell Division Drive Adaptive Intestinal Growth
2011; 147 (3): 603-614
Throughout life, adult organs continually adapt to variable environmental factors. Adaptive mechanisms must fundamentally differ from homeostatic maintenance, but little is known about how physiological factors elicit tissue remodeling. Here, we show that specialized stem cell responses underlie the adaptive resizing of a mature organ. In the adult Drosophila midgut, intestinal stem cells interpret a nutrient cue to "break homeostasis" and drive growth when food is abundant. Activated in part by niche production of insulin, stem cells direct a growth program through two altered modes of behavior: accelerated division rates and predominance of symmetric division fates. Together, these altered modes produce a net increase in total intestinal cells, which is reversed upon withdrawal of food. Thus, tissue renewal programs are not committed to maintain cellular equilibrium; stem cells can remodel organs in response to physiological triggers.
View details for DOI 10.1016/j.cell.2011.08.048
View details for Web of Science ID 000296573700015
View details for PubMedID 22036568
ERK and MMPs sequentially regulate distinct stages of epithelial tubule development
2004; 7 (1): 21-32
Epithelial cells undergo tubulogenesis in response to morphogens such as hepatocyte growth factor (HGF). To organize into tubules, cells must execute a complex series of morphogenetic events; however, the mechanisms that underlie the timing and sequence of these events are poorly understood. Here, we show that downstream effectors of HGF coordinately regulate successive stages of tubulogenesis. Activation of extracellular-regulated kinase (ERK) is necessary and sufficient for the initial stage, during which cells depolarize and migrate. ERK becomes dispensable for the latter stage, during which cells repolarize and differentiate. Conversely, the activity of matrix metalloproteases (MMPs) is essential for the late stage but not the initial stage. Thus, ERK and MMPs define two regulatory subprograms that act in sequence. By inducing these reciprocal signals, HGF directs the morphogenetic progression of tubule development.
View details for Web of Science ID 000222696300007
View details for PubMedID 15239951
Opinion - Building epithelial architecture: insights from three-dimensional culture models
NATURE REVIEWS MOLECULAR CELL BIOLOGY
2002; 3 (7): 531-537
How do individual cells organize into multicellular tissues? Here, we propose that the morphogenetic behaviour of epithelial cells is guided by two distinct elements: an intrinsic differentiation programme that drives formation of a lumen-enclosing monolayer, and a growth factor-induced, transient de-differentiation that allows this monolayer to be remodelled.
View details for DOI 10.1038/nrm859
View details for Web of Science ID 000176563500018
View details for PubMedID 12094219
Rac1 orientates epithelial apical polarity through effects on basolateral laminin assembly
NATURE CELL BIOLOGY
2001; 3 (9): 831-838
Cellular polarization involves the generation of asymmetry along an intracellular axis. In a multicellular tissue, the asymmetry of individual cells must conform to the overlying architecture of the tissue. However, the mechanisms that couple cellular polarization to tissue morphogenesis are poorly understood. Here, we report that orientation of apical polarity in developing Madin-Darby canine kidney (MDCK) epithelial cysts requires the small GTPase Rac1 and the basement membrane component laminin. Dominant-negative Rac1 alters the supramolecular assembly of endogenous MDCK laminin and causes a striking inversion of apical polarity. Exogenous laminin is recruited to the surface of these cysts and rescues apical polarity. These findings implicate Rac1-mediated laminin assembly in apical pole orientation. By linking apical orientation to generation of the basement membrane, epithelial cells ensure the coordination of polarity with tissue architecture.
View details for Web of Science ID 000170979600017
View details for PubMedID 11533663
Regional specificity in the Drosophila midgut: setting boundaries with stem cells.
Cell stem cell
2013; 13 (4): 375-376
Many organs consist of distinct subregions with specialized physiological roles, but how regional boundaries are upheld during cellular renewal is largely unknown. Recently, Buchon et al. (2013) and Marianes and Spradling (2013) showed that subregions of the Drosophila midgut are maintained by patterned transcription factors and compartmentalized stem cell progeny.
View details for DOI 10.1016/j.stem.2013.09.008
View details for PubMedID 24094316
STAT1 Is Required for Redifferentiation during Madin-Darby Canine Kidney Tubulogenesis
MOLECULAR BIOLOGY OF THE CELL
2010; 21 (22): 3926-3933
Tubule formation in vitro using Madin-Darby canine kidney (MDCK) epithelial cells consists mainly of two processes. First, the cells undergo a partial epithelial-mesenchymal transition (pEMT), losing polarity and migrating. Second, the cells redifferentiate, forming cords and then tubules with continuous lumens. We have shown previously that extracellular signal-regulated kinase activation is required for pEMT. However, the mechanism of how the pEMT phase is turned off and the redifferentiation phase is initiated is largely unknown. To address the central question of the sequential control of these two phases, we used MDCK cells grown as cysts and treated with hepatocyte growth factor to model tubulogenesis. We show that signal transducer and activator of transcription (STAT)1 controls the sequential progression from the pEMT phase to the redifferentiation phase. Loss of STAT1 prevents redifferentiation. Constitutively active STAT1 allows redifferentiation to occur even when cells are otherwise prevented from progressing beyond the pEMT phase by exogenous activation of Raf. Moreover, tyrosine phosphorylation defective STAT1 partially restored cord formation in such cells, suggesting that STAT1 functions in part as nonnuclear protein mediating signal transduction in this process. Constitutively active or inactive forms of STAT1 did not promote lumen maturation, suggesting this requires a distinct signal.
View details for DOI 10.1091/mbc.E10-02-0112
View details for Web of Science ID 000284216800043
View details for PubMedID 20861313
Morphological and biochemical analysis of Rac1 in three-dimensional epithelial cell cultures
METHODS IN ENZYMOLOGY, VOL 406, REGULATORS AND EFFECTORS OF SMALL GTPASES: RHO FAMILY
2006; 406: 676-691
Rho GTPases are critical regulators of epithelial morphogenesis. A powerful means to investigate their function is three-dimensional (3D) cell culture, which mimics the architecture of epithelia in vivo. However, the nature of 3D culture requires specialized techniques for morphological and biochemical analyses. Here, we describe protocols for 3D culture studies with Madin-Darby Canine Kidney (MDCK) epithelial cells: establishing cultures, immunostaining, and expressing, detecting, and assaying Rho proteins. These protocols enable the regulation of epithelial morphogenesis to be explored at a detailed molecular level.
View details for DOI 10.1016/S0076-6879(06)06053-8
View details for Web of Science ID 000235750600053
View details for PubMedID 16472697
beta 1-integrin orients epithelial polarity via Rac1 and laminin
MOLECULAR BIOLOGY OF THE CELL
2005; 16 (2): 433-445
Epithelial cells polarize and orient polarity in response to cell-cell and cell-matrix adhesion. Although there has been much recent progress in understanding the general polarizing machinery of epithelia, it is largely unclear how this machinery is controlled by the extracellular environment. To explore the signals from cell-matrix interactions that control orientation of cell polarity, we have used three-dimensional culture systems in which Madin-Darby canine kidney (MDCK) cells form polarized, lumen-containing structures. We show that interaction of collagen I with apical beta1-integrins after collagen overlay of a polarized MDCK monolayer induces activation of Rac1, which is required for collagen overlay-induced tubulocyst formation. Cysts, comprised of a monolayer enclosing a central lumen, form after embedding single cells in collagen. In those cultures, addition of a beta1-integrin function-blocking antibody to the collagen matrix gives rise to cysts that have defects in the organization of laminin into the basement membrane and have inverted polarity. Normal polarity is restored by either expression of activated Rac1, or the inclusion of excess laminin-1 (LN-1). Together, our results suggest a signaling pathway in which the activation of beta1-integrins orients the apical pole of polarized cysts via a mechanism that requires Rac1 activation and laminin organization into the basement membrane.
View details for DOI 10.1091/mcb.E04-05-0435
View details for Web of Science ID 000226563600001
View details for PubMedID 15574881
Formation of multicellular epithelial structures.
Novartis Foundation symposium
2005; 269: 193-200
The kidney is primarily comprised of highly polarized epithelial cells. Much has been learned recently about the mechanisms of epithelial polarization. However, in most experimental systems the orientation of this polarity is determined by external cues, such as growth of epithelial cells on a filter support. When Madin-Darby canine kidney (MDCK) cells are grown instead in a three-dimensional (3D) collagen gel, the cells form hollow cysts lined by a monolayer of epithelial cells, with their apical surfaces all facing the central lumen. We have found that expression of a dominant-negative (DN) form of the small GTPase Rac1 causes an inversion of epithelial polarity, such that the apical surface of the cells instead faces the periphery of the cyst. This indicates that the establishment of polarity and the orientation of polarity can be experimentally separated by growing cells in a 3D collagen gel, where there is no filter support to provide an external cue for orientation. DN Rac1 causes a defect in the assembly of laminin into its normal basement membrane network, and addition of a high concentration of exogenous laminin rescues the inversion of polarity caused by DN Rac1.
View details for PubMedID 16355541
Epithelial polarity and tubulogenesis in vitro
TRENDS IN CELL BIOLOGY
2003; 13 (4): 169-176
The most fundamental type of organization of cells in metazoa is that of epithelia, which comprise sheets of adherent cells that divide the organism into topologically and physiologically distinct spaces. Some epithelial cells cover the outside of the organism; these often form multiple layers, such as in skin. Other epithelial cells form monolayers that line internal organs, and yet others form tubes that infiltrate the whole organism, carrying liquids and gases containing nutrients, waste and other materials. These tubes can form elaborate networks in the lung, kidney, reproductive passages and vasculature tree, as well as the many glands branching from the digestive system such as the liver, pancreas and salivary glands. In vitro systems can be used to study tube formation and might help to define common principles underlying the formation of diverse types of tubular organ.
View details for DOI 10.1016/S0962-8924(03)00036-9
View details for Web of Science ID 000182394700004
View details for PubMedID 12667754
Hepatocyte growth factor switches orientation of polarity and mode of movement during morphogenesis of multicellular epithelial structures
MOLECULAR BIOLOGY OF THE CELL
2003; 14 (2): 748-763
Epithelial cells form monolayers of polarized cells with apical and basolateral surfaces. Madin-Darby canine kidney epithelial cells transiently lose their apico-basolateral polarity and become motile by treatment with hepatocyte growth factor (HGF), which causes the monolayer to remodel into tubules. HGF induces cells to produce basolateral extensions. Cells then migrate out of the monolayer to produce chains of cells, which go on to form tubules. Herein, we have analyzed the molecular mechanisms underlying the production of extensions and chains. We find that cells switch from an apico-basolateral polarization in the extension stage to a migratory cell polarization when in chains. Extension formation requires phosphatidyl-inositol 3-kinase activity, whereas Rho kinase controls their number and length. Microtubule dynamics and cell division are required for the formation of chains, but not for extension formation. Cells in the monolayer divide with their spindle axis parallel to the monolayer. HGF causes the spindle axis to undergo a variable "seesaw" motion, so that a daughter cells can apparently leave the monolayer to initiate a chain. Our results demonstrate the power of direct observation in investigating how individual cell behaviors, such as polarization, movement, and division are coordinated in the very complex process of producing multicellular structures.
View details for Web of Science ID 000182184300030
View details for PubMedID 12589067
Analysis of membrane traffic in polarized epithelial cells.
Current protocols in cell biology / editorial board, Juan S. Bonifacino ... [et al.]
2001; Chapter 15: Unit 15 5-?
Spatial asymmetry is fundamental to the structure and function of most eukaryotic cells. A basic aspect of this polarity is that the cell's plasma membrane is divided into discrete domains. The best studied and simplest example of this occurs in epithelial cells, which line exposed body surfaces. Epithelial cells use two pathways to send proteins to the cell surface. Newly made proteins can travel directly from the trans-Golgi network (TGN) to either the apical or basolateral surface. Alternatively, proteins can be sent to the basolateral surface and then endocytosed and transcytosed to the apical surface. Epithelial cells grown on porous filters adopt a typical polarized morphology; transfected epithelial cells can be used to biosynthetically characterize the trafficking patterns of a given protein. These cells can also be used to study delivery to a particular surface and to localize the protein by immunofluorescence.
View details for DOI 10.1002/0471143030.cb1505s12
View details for PubMedID 18228332
Exocyst is involved in cystogenesis and tubulogenesis and acts by modulating synthesis and delivery of basolateral plasma membrane and secretory proteins
MOLECULAR BIOLOGY OF THE CELL
2000; 11 (12): 4259-4275
Epithelial cyst and tubule formation are critical processes that involve transient, highly choreographed changes in cell polarity. Factors controlling these changes in polarity are largely unknown. One candidate factor is the highly conserved eight-member protein complex called the exocyst. We show that during tubulogenesis in an in vitro model system the exocyst relocalized along growing tubules consistent with changes in cell polarity. In yeast, the exocyst subunit Sec10p is a crucial component linking polarized exocytic vesicles with the rest of the exocyst complex and, ultimately, the plasma membrane. When the exocyst subunit human Sec10 was exogenously expressed in epithelial Madin-Darby canine kidney cells, there was a selective increase in the synthesis and delivery of apical and basolateral secretory proteins and a basolateral plasma membrane protein, but not an apical plasma membrane protein. Overexpression of human Sec10 resulted in more efficient and rapid cyst formation and increased tubule formation upon stimulation with hepatocyte growth factor. We conclude that the exocyst plays a central role in the development of epithelial cysts and tubules.
View details for Web of Science ID 000165955000016
View details for PubMedID 11102522