As a graduate student at University of California, San Francisco, I studied the genetic regulation of centrosome structure, function, and positioning and the mechanisms dictating internal cellular organization using the unicellular alga Chlamydomonas. I went on to characterize the role of the centrosome during epithelial polarization in C. elegans, working as a postdoctoral fellow at the Fred Hutchinson Cancer Research Center. I started my lab in the Biology Department in 2014. In my lab, we study structural changes that occur at the cellular level during normal development and in disease. In particular, we are interested in understanding how microtubules become spatially organized in different cell types during cell differentiation.
Honors & Awards
New Innovator Award, NIH (2015-2020)
Basil O’Connor Starter Scholar Research Award, March of Dimes Foundation (2015-2017)
Postdoctoral Fellowship, American Heart Association (2013)
Postdoctoral Fellowship, Helen Hay Whitney Foundation (2010-2013)
Predoctoral Fellowship, National Science Foundation
Ph.D., University of California, San Francisco, Cell Biology (2008)
B.A., Columbia University, Biology (2000)
Current Research and Scholarly Interests
We are interested in understanding design principles within cells that contribute to the diversification of cellular form and function. Using a combination of genetic, biochemical, and live imaging approaches, we are investigating how the microtubule cytoskeleton is spatially organized and the mechanisms underlying organizational changes during development.
Doctoral Dissertation Reader (AC)
Le Cai, KC Farrell, Julio Flores Servin, Ray Futia, Yan Gong, Melody Yung-Hsin Kao, Andrew Kennard, Miranda Stratton, Claire Venard
Postdoctoral Faculty Sponsor
Lauren Cote, James Ferguson, Jeremy Magescas, Melissa Pickett, Maria Sallee
Doctoral Dissertation Advisor (AC)
Alex Lessenger, Victor Naturale, Ariana Sanchez, Michael Tran
Victor Naturale, Ariana Sanchez, Michael Tran
Graduate and Fellowship Programs
Biology (School of Humanities and Sciences) (Phd Program)
SPD-2/CEP192 and CDK Are Limiting for Microtubule-Organizing Center Function at the Centrosome
2015; 25 (14): 1924-1931
The centrosome acts as the microtubule-organizing center (MTOC) during mitosis in animal cells. Microtubules are nucleated and anchored by γ-tubulin ring complexes (γ-TuRCs) embedded within the centrosome's pericentriolar material (PCM). The PCM is required for the localization of γ-TuRCs, and both are steadily recruited to the centrosome, culminating in a peak in MTOC function in metaphase. In differentiated cells, the centrosome is often attenuated as an MTOC and MTOC function is reassigned to non-centrosomal sites such as the apical membrane in epithelial cells, the nuclear envelope in skeletal muscle, and down the lengths of axons and dendrites in neurons. Hyperactive MTOC function at the centrosome is associated with epithelial cancers and with invasive behavior in tumor cells. Little is known about the mechanisms that limit MTOC activation at the centrosome. Here, we find that MTOC function at the centrosome is completely inactivated during cell differentiation in C. elegans embryonic intestinal cells and MTOC function is reassigned to the apical membrane. In cells that divide after differentiation, the cellular MTOC state switches between the membrane and the centrosome. Using cell fusion experiments in live embryos, we find that the centrosome MTOC state is dominant and that the inactive MTOC state of the centrosome is malleable; fusion of a mitotic cell to a differentiated or interphase cell results in rapid reactivation of the centrosome MTOC. We show that conversion of MTOC state involves the conserved centrosome protein SPD-2/CEP192 and CDK activity from the mitotic cell.
View details for DOI 10.1016/j.cub.2015.06.001
View details for Web of Science ID 000358465600033
Quantitative analysis and modeling of katanin function in flagellar length control
MOLECULAR BIOLOGY OF THE CELL
2014; 25 (22): 3686-3698
Flagellar length control in Chlamydomonas reinhardtii provides a simple model system in which to investigate the general question of how cells regulate organelle size. Previous work demonstrated that Chlamydomonas cytoplasm contains a pool of flagellar precursor proteins sufficient to assemble a half-length flagellum and that assembly of full-length flagella requires synthesis of additional precursors to augment the preexisting pool. The regulatory systems that control the synthesis and regeneration of this pool are not known, although transcriptional regulation clearly plays a role. We used quantitative analysis of length distributions to identify candidate genes controlling pool regeneration and found that a mutation in the p80 regulatory subunit of katanin, encoded by the PF15 gene in Chlamydomonas, alters flagellar length by changing the kinetics of precursor pool utilization. This finding suggests a model in which flagella compete with cytoplasmic microtubules for a fixed pool of tubulin, with katanin-mediated severing allowing easier access to this pool during flagellar assembly. We tested this model using a stochastic simulation that confirms that cytoplasmic microtubules can compete with flagella for a limited tubulin pool, showing that alteration of cytoplasmic microtubule severing could be sufficient to explain the effect of the pf15 mutations on flagellar length.
View details for DOI 10.1091/mbc.E14-06-1116
View details for Web of Science ID 000344236800030
View details for PubMedID 25143397
The Kinase Regulator Mob1 Acts as a Patterning Protein for Stentor Morphogenesis
2014; 12 (5)
Morphogenesis and pattern formation are vital processes in any organism, whether unicellular or multicellular. But in contrast to the developmental biology of plants and animals, the principles of morphogenesis and pattern formation in single cells remain largely unknown. Although all cells develop patterns, they are most obvious in ciliates; hence, we have turned to a classical unicellular model system, the giant ciliate Stentor coeruleus. Here we show that the RNA interference (RNAi) machinery is conserved in Stentor. Using RNAi, we identify the kinase coactivator Mob1--with conserved functions in cell division and morphogenesis from plants to humans-as an asymmetrically localized patterning protein required for global patterning during development and regeneration in Stentor. Our studies reopen the door for Stentor as a model regeneration system.
View details for DOI 10.1371/journal.pbio.1001861
View details for Web of Science ID 000336969200011
View details for PubMedID 24823688
Cell Interactions and Patterned Intercalations Shape and Link Epithelial Tubes in C. elegans
2013; 9 (9)
Many animal organs are composed largely or entirely of polarized epithelial tubes, and the formation of complex organ systems, such as the digestive or vascular systems, requires that separate tubes link with a common polarity. The Caenorhabditis elegans digestive tract consists primarily of three interconnected tubes-the pharynx, valve, and intestine-and provides a simple model for understanding the cellular and molecular mechanisms used to form and connect epithelial tubes. Here, we use live imaging and 3D reconstructions of developing cells to examine tube formation. The three tubes develop from a pharynx/valve primordium and a separate intestine primordium. Cells in the pharynx/valve primordium polarize and become wedge-shaped, transforming the primordium into a cylindrical cyst centered on the future lumenal axis. For continuity of the digestive tract, valve cells must have the same, radial axis of apicobasal polarity as adjacent intestinal cells. We show that intestinal cells contribute to valve cell polarity by restricting the distribution of a polarizing cue, laminin. After developing apicobasal polarity, many pharyngeal and valve cells appear to explore their neighborhoods through lateral, actin-rich lamellipodia. For a subset of cells, these lamellipodia precede more extensive intercalations that create the valve. Formation of the valve tube begins when two valve cells become embedded at the left-right boundary of the intestinal primordium. Other valve cells organize symmetrically around these two cells, and wrap partially or completely around the orthogonal, lumenal axis, thus extruding a small valve tube from the larger cyst. We show that the transcription factors DIE-1 and EGL-43/EVI1 regulate cell intercalations and cell fates during valve formation, and that the Notch pathway is required to establish the proper boundary between the pharyngeal and valve tubes.
View details for DOI 10.1371/journal.pgen.1003772
View details for Web of Science ID 000325076600033
View details for PubMedID 24039608
A Role for the Centrosome and PAR-3 in the Hand-Off of MTOC Function during Epithelial Polarization
2012; 22 (7): 575-582
The centrosome is the major microtubule organizing center (MTOC) in dividing cells and in many postmitotic, differentiated cells. In other cell types, however, MTOC function is reassigned from the centrosome to noncentrosomal sites. Here, we analyze how MTOC function is reassigned to the apical membrane of C. elegans intestinal cells.After the terminal intestinal cell division, the centrosomes and nuclei move near the future apical membranes, and the postmitotic centrosomes lose all, or nearly all, of their associated microtubules. We show that microtubule-nucleating proteins such as γ-tubulin and CeGrip-1 that are centrosome components in dividing cells become localized to the apical membrane, which becomes highly enriched in microtubules. Our results suggest that centrosomes are critical to specify the apical membrane as the new MTOC. First, γ-tubulin appears to redistribute directly from the migrating centrosome onto the lateral then apical membrane. Second, γ-tubulin fails to accumulate apically in wild-type cells following laser ablation of the centrosome. We show that centrosomes localize apically by first moving toward lateral foci of the conserved polarity proteins PAR-3 and PAR-6 and then move together with these foci toward the future apical surface. Embryos lacking PAR-3 fail to localize their centrosomes apically and have aberrant localization of γ-tubulin and CeGrip-1.These data suggest that PAR proteins contribute to apical polarity in part by determining centrosome position and that the reassignment of MTOC function from centrosomes to the apical membrane is associated with a physical hand-off of nucleators of microtubule assembly.
View details for DOI 10.1016/j.cub.2012.02.044
View details for Web of Science ID 000302844900018
View details for PubMedID 22425160
C. elegans Germ Cells Show Temperature and Age-Dependent Expression of Cer1, a Gypsy/Ty3-Related Retrotransposon
2012; 8 (3)
Virus-like particles (VLPs) have not been observed in Caenorhabditis germ cells, although nematode genomes contain low numbers of retrotransposon and retroviral sequences. We used electron microscopy to search for VLPs in various wild strains of Caenorhabditis, and observed very rare candidate VLPs in some strains, including the standard laboratory strain of C. elegans, N2. We identified the N2 VLPs as capsids produced by Cer1, a retrotransposon in the Gypsy/Ty3 family of retroviruses/retrotransposons. Cer1 expression is age and temperature dependent, with abundant expression at 15°C and no detectable expression at 25°C, explaining how VLPs escaped detection in previous studies. Similar age and temperature-dependent expression of Cer1 retrotransposons was observed for several other wild strains, indicating that these properties are common, if not integral, features of this retroelement. Retrotransposons, in contrast to DNA transposons, have a cytoplasmic stage in replication, and those that infect non-dividing cells must pass their genomic material through nuclear pores. In most C. elegans germ cells, nuclear pores are largely covered by germline-specific organelles called P granules. Our results suggest that Cer1 capsids target meiotic germ cells exiting pachytene, when free nuclear pores are added to the nuclear envelope and existing P granules begin to be removed. In pachytene germ cells, Cer1 capsids concentrate away from nuclei on a subset of microtubules that are exceptionally resistant to microtubule inhibitors; the capsids can aggregate these stable microtubules in older adults, which exhibit a temperature-dependent decrease in egg viability. When germ cells exit pachytene, the stable microtubules disappear and capsids redistribute close to nuclei that have P granule-free nuclear pores. This redistribution is microtubule dependent, suggesting that capsids that are released from stable microtubules transfer onto new, dynamic microtubules to track toward nuclei. These studies introduce C. elegans as a model to study the interplay between retroelements and germ cell biology.
View details for DOI 10.1371/journal.ppat.1002591
View details for Web of Science ID 000302225600040
View details for PubMedID 22479180
A Cell-Based Screen for Inhibitors of Flagella-Driven Motility in Chlamydomonas Reveals a Novel Modulator of Ciliary Length and Retrograde Actin Flow
2011; 68 (3): 188-203
Cilia are motile and sensory organelles with critical roles in physiology. Ciliary defects can cause numerous human disease symptoms including polycystic kidneys, hydrocephalus, and retinal degeneration. Despite the importance of these organelles, their assembly and function is not fully understood. The unicellular green alga Chlamydomonas reinhardtii has many advantages as a model system for studies of ciliary assembly and function. Here we describe our initial efforts to build a chemical-biology toolkit to augment the genetic tools available for studying cilia in this organism, with the goal of being able to reversibly perturb ciliary function on a rapid time-scale compared to that available with traditional genetic methods. We screened a set of 5520 compounds from which we identified four candidate compounds with reproducible effects on flagella at nontoxic doses. Three of these compounds resulted in flagellar paralysis and one induced flagellar shortening in a reversible and dose-dependent fashion, accompanied by a reduction in the speed of intraflagellar transport. This latter compound also reduced the length of cilia in mammalian cells, hence we named the compound "ciliabrevin" due to its ability to shorten cilia. This compound also robustly and reversibly inhibited microtubule movement and retrograde actin flow in Drosophila S2 cells. Ciliabrevin may prove especially useful for the study of retrograde actin flow at the leading edge of cells, as it slows the retrograde flow in a tunable dose-dependent fashion until flow completely stops at high concentrations, and these effects are quickly reversed upon washout of the drug.
View details for DOI 10.1002/cm.20504
View details for Web of Science ID 000288180600005
View details for PubMedID 21360831
ASQ2 Encodes a TBCC-like Protein Required for Mother-Daughter Centriole Linkage and Mitotic Spindle Orientation
2009; 19 (14): 1238-1243
An intriguing feature of centrioles is that these highly complicated microtubule-based structures duplicate once per cell cycle, affording the cell precise control over their number. Each cell contains exactly two centrioles, linked together as a pair, one of which is a mother centriole formed in a previous cell cycle and the other of which is a daughter centriole whose assembly is templated by the mother. Neither the molecular basis nor the functional role of mother-daughter centriole linkage is understood. We have identified a mutant, asq2, with defects in centriole linkage. asq2 mutant cells have variable numbers of centrioles and centriole positioning defects. Here, we show that ASQ2 encodes the conserved protein Tbccd1, a member of a protein family including a tubulin folding cochaperone and the retinitis pigmentosa protein RP2, involved in tubulin quality control during ciliogenesis. We characterize mitosis in asq2 cells and show that the majority of cells establish a bipolar spindle but have defects in spindle orientation. Few asq2 cells have centrioles at both poles, and these cells have properly positioned spindles, indicating that centrioles at the poles might be important for spindle orientation. The defects in centriole number control, centriole positioning, and spindle orientation appear to arise from perturbation of centriole linkage mediated by Tbccd1/Asq2p.
View details for DOI 10.1016/j.cub.2009.05.071
View details for Web of Science ID 000268530200034
View details for PubMedID 19631545
Katanin Knockdown Supports a Role for Microtubule Severing in Release of Basal Bodies before Mitosis in Chlamydomonas
MOLECULAR BIOLOGY OF THE CELL
2009; 20 (1): 379-388
Katanin is a microtubule-severing protein that participates in the regulation of cell cycle progression and in ciliary disassembly, but its precise role is not known for either activity. Our data suggest that in Chlamydomonas, katanin severs doublet microtubules at the proximal end of the flagellar transition zone, allowing disengagement of the basal body from the flagellum before mitosis. Using an RNA interference approach we have discovered that severe knockdown of the p60 subunit of katanin, KAT1, is achieved only in cells that also carry secondary mutations that disrupt ciliogenesis. Importantly, we observed that cells in the process of cell cycle-induced flagellar resorption sever the flagella from the basal bodies before resorption is complete, and we find that this process is defective in KAT1 knockdown cells.
View details for DOI 10.1091/mbc.E07-10-1007
View details for Web of Science ID 000262134800035
View details for PubMedID 19005222
The mother centriole plays an instructive role in defining cell geometry
2007; 5 (6): 1284-1297
Centriole positioning is a key step in establishment and propagation of cell geometry, but the mechanism of this positioning is unknown. The ability of pre-existing centrioles to induce formation of new centrioles at a defined angle relative to themselves suggests they may have the capacity to transmit spatial information to their daughters. Using three-dimensional computer-aided analysis of cell morphology in Chlamydomonas, we identify six genes required for centriole positioning relative to overall cell polarity, four of which have known sequences. We show that the distal portion of the centriole is critical for positioning, and that the centriole positions the nucleus rather than vice versa. We obtain evidence that the daughter centriole is unable to respond to normal positioning cues and relies on the mother for positional information. Our results represent a clear example of "cytotaxis" as defined by Sonneborn, and suggest that centrioles can play a key function in propagation of cellular geometry from one generation to the next. The genes documented here that are required for proper centriole positioning may represent a new class of ciliary disease genes, defects in which would be expected to cause disorganized ciliary position and impaired function.
View details for DOI 10.1371/journal.pbio.0050149
View details for Web of Science ID 000247173200013
View details for PubMedID 17518519
Retinoic acid signaling restricts the cardiac progenitor pool
2005; 307 (5707): 247-249
Organogenesis begins with specification of a progenitor cell population, the size of which provides a foundation for the organ's final dimensions. Here, we present a new mechanism for regulating the number of progenitor cells by limiting their density within a competent region. We demonstrate that retinoic acid signaling restricts cardiac specification in the zebrafish embryo. Reduction of retinoic acid signaling causes formation of an excess of cardiomyocytes, via fate transformations that increase cardiac progenitor density within a multipotential zone. Thus, retinoic acid signaling creates a balance between cardiac and noncardiac identities, thereby refining the dimensions of the cardiac progenitor pool.
View details for DOI 10.1126/science.1101573
View details for Web of Science ID 000226361900040
View details for PubMedID 15653502
- PCR-based assay for mating type and diploidy in Chlamydomonas BIOTECHNIQUES 2004; 37 (4): 534-536
Centrioles: Bad to be bald?
2004; 14 (16): R659-R660
In the alga Chlamydomonas, mutation of the gene encoding the novel centriolar component Bld10p results in seemingly acentriolar cells. Remarkably, bld10 cells are viable, highlighting the question of whether or not centrioles are essential.
View details for Web of Science ID 000223586900012
View details for PubMedID 15324683
The elongation factors Pandora/Spt6 and Foggy/Spt5 promote transcription in the zebrafish embryo
2002; 129 (7): 1623-1632
Precise temporal and spatial control of transcription is a fundamental component of embryonic development. Regulation of transcription elongation can act as a rate-limiting step during mRNA synthesis. The mechanisms of stimulation and repression of transcription elongation during development are not yet understood. We have identified a class of zebrafish mutations (pandora, sk8 and s30) that cause multiple developmental defects, including discrete problems with pigmentation, tail outgrowth, ear formation and cardiac differentiation. We demonstrate that the pandora gene encodes a protein similar to Spt6, a proposed transcription elongation factor. Additionally, the sk8 and s30 mutations are null alleles of the foggy/spt5 locus, which encodes another transcription elongation factor. Through real-time RT-PCR analysis, we demonstrate that Spt6 and Spt5 are both required for efficient kinetics of hsp70 transcription in vivo. Altogether, our results suggest that Spt6 and Spt5 play essential roles of comparable importance for promoting transcription during embryogenesis. This study provides the first genetic evidence for parallel functions of Spt6 and Spt5 in metazoans and establishes a system for the future analysis of transcription elongation during development.
View details for Web of Science ID 000175112300007
View details for PubMedID 11923199
- Genetic regulation of cardiac patterning in zebrafish Cold Spring Harbor Symposium on Quantitative Biology COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT. 2002: 19–25