© 2013. Published by The Company of Biologists Ltd | Development (2013) 140, 4844-4849 doi:10.1242/dev.103010
RESEARCH REPORT
STEM CELLS AND REGENERATION
Transdifferentiation and remodeling of post-embryonic C. elegans
cells by a single transcription factor
ABSTRACT
Terminally differentiated post-mitotic cells are generally considered
irreversibly developmentally locked, i.e. incapable of being
reprogrammed in vivo into entirely different cell types. We found that
brief expression of a single transcription factor, the ELT-7 GATA
factor, can convert the identity of fully differentiated, highly specialized
non-endodermal cells of the pharynx into fully differentiated intestinal
cells in intact larvae and adult Caenorhabditis elegans. Stable
expression of intestine-specific molecular markers parallels loss of
markers for the original differentiated pharynx state; hence, there is
no apparent requirement for a dedifferentiated intermediate during
the transdifferentiation process. Based on high-resolution
morphological characteristics, the transdifferentiated cells become
remodeled to resemble typical intestinal cells at the level of both the
cell surface and internal organelles. Thus, post-mitotic cells, though
terminally differentiated, remain plastic to transdifferentiation across
germ layer lineage boundaries and can be remodeled to adopt the
characteristics of a new cell identity without removal of inhibitory
factors. Our findings establish a simple model to investigate how cell
context influences forced transdifferentiation of mature cells.
germline cells can prematurely adopt somatic cell fates upon
removal of translational regulators (Ciosk et al., 2006) or inhibitory
chromatin remodeling factors (Tursun et al., 2011; Patel et al.,
2012). However, germline cells are uncommitted pluripotent cells
that are poised to differentiate shortly after gametogenesis, whereas
most somatic cells become terminally differentiated.
We report here that a transcription factor that regulates terminal
intestine differentiation violates the embryonic multipotency to
commitment transition (MCT) and can reprogram and remodel
differentiated cells in C. elegans larvae and adults into intestine-like
cells without removal of inhibitory factors. We found that a brief
pulse of the ELT-7 GATA transcription factor activates intestinespecific gene expression in diverse non-intestinal cells. Post-mitotic
pharyngeal cells maintain intestine gene expression, lose pharynx
gene expression, and become remodeled to resemble intestinal cells
at the fine ultrastuctural level. Our results suggests that post-mitotic,
terminally differentiated cells can be reprogrammed and remodeled
without prior removal of the initial cell identity and demonstrate that
susceptibility to in vivo reprogramming is influenced by
transcription factor identity and cellular context.
KEY WORDS: Transdifferentiation, Cellular reprogramming,
C. elegans
RESULTS AND DISCUSSION
ELT-7 activates and maintains intestine-specific gene
expression in differentiated pharyngeal cells at all stages of
development
INTRODUCTION
Cell fate is progressively restricted during development such that
most cells ultimately become post-mitotic, highly specialized and
terminally differentiated, with a fixed morphology and pattern of
gene expression. Early embryonic cells in C. elegans are capable of
being reprogrammed into cell types of all three germ layers by
appropriate transcription factors, but lose this ability during midembryogenesis, as they commit to particular pathways of
differentiation (Horner et al., 1998; Zhu et al., 1998; Gilleard and
McGhee, 2001; Quintin et al., 2001; Fukushige and Krause, 2005).
Eliminating polycomb complex (Yuzyuk et al., 2009) or Notch
signaling (Djabrayan et al., 2012) components extends the period for
multipotency during embryogenesis; however, post-mitotic somatic
cells in late embryos, larvae or adults cannot be reprogrammed by
the transcription factors reported to date (Horner et al., 1998; Zhu et
al., 1998; Gilleard and McGhee, 2001; Quintin et al., 2001;
Fukushige and Krause, 2005). Developing and proliferating
1
Department of Molecular, Cellular and Developmental Biology, and Neuroscience
Research Institute, University of California, Santa Barbara, CA 93106, USA.
2
Center for C. elegans Anatomy, Albert Einstein College of Medicine, Bronx,
NY 10461, USA. 3School of Biological Sciences, University of Auckland,
Auckland 1010, New Zealand.
‡
Present address: Department of Biology, Massachusetts Institute of Technology,
Cambridge, MA 02139, USA.
*Author for correspondence ([email protected])
Received 29 August 2013; Accepted 29 September 2013
4844
Although early embryonic cells in C. elegans are multipotent, cells
become refractory to reprogramming later in embryogenesis,
following the MCT (Horner et al., 1998; Zhu et al., 1998; Gilleard
and McGhee, 2001; Quintin et al., 2001; Fukushige and Krause,
2005). In contrast to other regulators of cell fate specification that
have been reported, we found that ELT-7, a GATA transcription
factor that regulates terminal intestinal differentiation (Maduro and
Rothman, 2002; Sommermann et al., 2010), activates an intestinal
marker (elt-2::lacZ::GFP) in non-intestinal cells when briefly
ectopically expressed via a heat-shock promoter at any embryonic,
larval, or adult stage (Fig. 1A-C; supplementary material Fig. S1).
We also detected IFB-2, an intermediate filament protein that is a
component of the terminal web of the fully differentiated intestine
(Bossinger et al., 2004), outside of the intestine in such animals
(Fig. 1D; supplementary material Fig. S2). By contrast, the END-1
GATA transcription factor, which specifies the endoderm progenitor
(Zhu et al., 1997), does not activate widespread intestinal gene
expression after the MCT (Fig. 1; supplementary material Fig. S2).
Thus, ELT-7 may violate the well-documented restriction to
developmental reprogramming that occurs during embryogenesis.
In the endogenous intestine, ELT-7 and ELT-2 expression is
maintained throughout larval development and adulthood via
cross- and auto-regulatory feedback mechanisms (Zhu et al., 1998;
Fukushige et al., 1999; Sommermann et al., 2010). Although
intestinal gene expression is never observed outside the intestine
in non-heat-shocked hs-elt-7 larvae (n=128) or in heat-shocked
Development
Misty R. Riddle1, Abraham Weintraub1,‡, Ken C. Q. Nguyen2, David H. Hall2 and Joel H. Rothman1,3,*
RESEARCH REPORT
Development (2013) doi:10.1242/dev.103010
larvae that do not contain the hs-elt-7 transgene (n=46), over 50%
(n=13) of larvae carrying the hs-elt-7 transgene express the elt-2
intestinal reporter in non-intestinal cells as early as 4 hours after
heat shock (Fig. 2C). Expression increases in intensity and cell
number rapidly: by 6 hours, broad expression is seen in 100% of
animals and remains high over the ensuing 24 hours (n=16).
Although expression in most cells is then progressively lost, we
found that cells in the neuromuscular feeding organ, the pharynx
(Fig. 2A-C), show strong continuous expression throughout the
life of the animal (Fig. 2B): by 72 hours, nearly all expression of
the intestinal marker is restricted to cells of the pharynx. The C.
elegans pharynx comprises seven different cell types that become
terminally differentiated and post-mitotic during embryogenesis
(Albertson and Thomson, 1976; Mango, 2007); the bulk of the
organ consists of muscle and marginal cells. We detected
immunoreactive ELT-2 (89% of animals; n=39) and IFB-2 (98%
of animals; n=98) in pharyngeal muscle and marginal cells of most
worms at 24-48 hours after ectopic ELT-7 expression (Fig. 2D;
supplementary material Table S1). Maintained expression of the
elt-2 reporter and immunoreactive ELT-2 and IFB-2 suggest that
the endogenous intestine ‘lockdown circuitry’ can be activated in
post-mitotic pharyngeal cells.
Pharyngeal cells lose their original differentiated identity
following activation of intestine differentiation
Several lines of evidence suggest that differentiated cells lose their
normal identities following ectopic ELT-7 expression. Although
animals exposed to brief END-1 expression grow and behave
normally, those subjected to brief ectopic ELT-7 arrest abruptly in
larval development (supplementary material Fig. S3, Table S2).
Despite this arrest, animals remain alive for an extended period (for
at least 8 days). Although active and motile, these non-developing
larvae are defective in chemosensation (supplementary material
Fig. S3) and molting (not shown), perhaps reflecting a switch in
gene expression states in the epidermis and neurons that transiently
express the elt-2 reporter after ectopic ELT-7 expression
(supplementary material Fig. S4).
Pharyngeal cells that maintain long-term ELT-2 expression lose
pharynx-specific gene expression (Fig. 3). Expression of a marker
for the myo-2 gene, which encodes a pharynx muscle-specific
myosin, is attenuated and undetectable only hours after forced
ubiquitous ELT-7 in embryos (supplementary material Fig. S5,
Movie 1). Further, expression of myo-2::GFP and ceh-22::GFP, an
NK-2 family homeobox transcription factor required for normal
pharynx muscle gene expression (Okkema and Fire, 1994)
progressively diminishes over several days following a pulse of
ectopic ELT-7 expression in larvae (Fig. 3A-D). Loss of pharynx
muscle-specific gene expression is not attributable to developmental
arrest, starvation, or general necrosis, as marker expression persists
under such conditions (Fig. 3B,D; supplementary material Fig. S6).
We found that structural components of fully differentiated
marginal cells are lost in parallel with acquisition of intestinal
markers following ectopic ELT-7 expression. We quantified the
intermediate filament protein IFA (Francis and Waterston, 1991) in
individual marginal cells of larvae based on average pixel intensities
of immunoreactive protein and found that marginal cells with strong
elt-2::lacZ::GFP-positive nuclei showed diminished IFA expression
compared with marginal cells with low or undetectable expression
in the same animal (Fig. 3E,F; eight worms analyzed, 24 total cells,
13 elt-2::lacZ::GFP-expressing cells). The disappearance of
otherwise stable pharynx-specific proteins, coincident with
maintenance of intestine-specific gene expression, is consistent with
bona fide transdifferentiation of differentiated, post-mitotic
pharyngeal marginal cells into intestinal cells.
Reprogrammed pharyngeal cells exhibit morphological
characteristics of intestinal cells
We found that reprogrammed marginal cells form organelles that
resemble those in intestinal cells. Within 3-5 days of a pulse of ELT7 expression, 11% of larvae (n=129) exhibit birefringent and
4845
Development
Fig. 1. ELT-7, but not END-1, activates endoderm
differentiation in differentiated non-endodermal cells.
(A) Percentage of embryos showing ectopic elt-2::lacZ::GFP
expression (>20 GFP+ nuclei) following heat shock at
various embryonic stages (based on E cell number). Total
number of embryos scored is shown above each bar.
(B) Embryos with widespread ectopic elt-2::lacZ::GFP
expression after ubiquitous END-1 or ELT-7 induction.
(C) Transient ectopic elt-2::lacZ::GFP in one cell outside the
pharynx (arrow) after ectopic END-1 expression and in
many cells throughout the body after ectopic ELT-7
expression. White line indicates the anterior end of the
intestine. Scale bars: 20 μm. (D) Expression of IFB-2
outside the intestine of hs-elt-7 but not hs-end-1 larvae.
Insets show region anterior to the endogenous intestine,
24 hours post-heat-shock.
RESEARCH REPORT
Development (2013) doi:10.1242/dev.103010
Fig. 2. Maintenance of intestinal differentiation markers
in pharyngeal cells. (A) The elt-2 reporter is normally
expressed only in intestinal nuclei (green) in non-heat
shocked hs-elt-7 larvae. Anterior (right) and posterior pharynx
regions in middle panel are enlarged in lower panels.
(B) 24 hours after ectopic ELT-7 expression, the elt-2 reporter
is expressed in cells inside and outside the pharynx. At
96 hours, bright signal is maintained only in cells within the
pharynx. Scale bars: 5 μm. (C) Percentage of worms
expressing elt-2::lacZ::GFP in non-intestinal cells outside the
pharynx (O), in the posterior pharynx (P) or in the anterior
pharynx (A). n, number of worms observed; t, time since heat
shock in hours. (D) Immunoreactive ELT-2 in non-heatshocked hs-elt-7 control larvae and hs-elt-7 larvae 48 hours
after heat shock.
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Specifically, cells in the pharynx exhibit accessory lumens (Fig. 4J)
that contain distinct microvilli with apparent glycocalyx on the
apical surfaces. We also observed a subjacent intestine-like terminal
web (Fig. 4K), as was indicated by immunofluorescence analysis
(Fig. 4G,H). Taken together with the expression data, our
observations suggest that several days after a 15-minute pulse of
ELT-7 expression, terminally differentiated pharyngeal cells become
transformed both at the gene expression and fine cellular structure
levels into differentiated intestine-like cells.
Pharynx to intestine transdifferentiation takes place
through mixed-identity post-mitotic cells
We found that a brief pulse of ectopic ELT-7 expression leads to
transdifferentiation of pharyngeal marginal cells, as evidenced by
maintained elt-2 reporter expression, stable ELT-2 protein, formation
of rhabditin granules, cellular remodeling to form a terminal web
and microvilli, as well as loss of normally stable marginal cell
proteins. Activation of endogenous ELT-2 by ectopic ELT-7, and
establishment of the intestine auto-regulatory circuitry, likely drives
transdifferentiation of marginal cells into intestinal cells. Increased
expression of intestine-specific proteins, and remodeling of the cell,
occur prior to repression of pharyngeal cell-specific identity,
suggesting that, unlike some examples of naturally occurring
transdifferentiation (Jarriault et al., 2008), this in vivo
reprogramming takes place through intermediate cellular states in
which characteristics of both the original and final cell types are
present. We found no evidence of cell division during the
reprogramming process based on cell number and position,
indicating that erasing original cell identity through cell division is
not a necessary step in forced transdifferentiation. Although some
morphological characteristics of pharynx tissue are present in the
animals with transdifferentiated cells, these ultrastructural features
are excluded in the regions that have undergone transdifferentiation
and that exhibit an intestine-like morphology.
Development
autofluorescent lysosome-related rhabditin granules (Clokey and
Jacobson, 1986) surrounding elt-2::LacZ::GFP-expressing nuclei
(Fig. 4A-E″; supplementary material Fig. S9, Table S3). Rhabditin
granules are normally present only in differentiated intestinal cells
and are never observed in the pharynx of untreated larvae (n>1000),
or in larvae ectopically expressing END-1 (n=36). Moreover, the
nuclei of cells with ectopic rhabditin granules are strikingly
reminiscent of typical differentiated intestinal nuclei (so-called ‘fried
egg nuclei’), with a clear nucleoplasm and prominent dense
nucleolus that are not typical of pharyngeal nuclei (Fig. 4C″-E″).
Formation of ectopic gut granules does not appear to be stage
dependent as we have observed them in the pharynx following
ectopic ELT-7 expression at later L1 (n=4), L2 (n=6), L3 (n=5) and
L4 stages (n=5) (n=144 worms observed).
The switch from pharynx-specific to intestine-specific gene
expression in fully differentiated cells is accompanied by
progressive remodeling of cellular structure. The marginal cells
accommodate the space between muscle cells along the length of the
pharynx and have a rigidly fixed wedge-like morphology supported
by radial bundles of intermediate filament proteins (Albertson and
Thomson, 1976; Francis and Waterston, 1991). We found that
immunoreactive IFB-2, an intestine-specific intermediate filament
that localizes to the intestine terminal web (Bossinger et al., 2004),
progressively localizes to the apical surface of marginal cells after
ectopic ELT-7 expression in L1 larvae and adults (Fig. 4F-H;
supplementary material Fig. S7). Expression and apical localization
of IFB-2 suggests that the cells may form an intestinal terminal web
structure following a brief pulse of ELT-7. Although apical surfaces
are narrow in normal marginal cells, they expand in area following
ELT-7 expression, consistent with remodeling to intestinal cell
structure (supplementary material Fig. S8).
Using transmission electron microscopy, we observed that the
fine-structure morphology of cells in the pharynx showed clear
evidence for transdifferentiation into intestinal cells (Fig. 4I-K).
RESEARCH REPORT
Development (2013) doi:10.1242/dev.103010
Transcription factor identity and cellular context influence
susceptibility to forced transdifferentiation
Although transcription factors that regulate specification of each of
the three germ layers can reprogram the fate of pre-gastrulation
blastomeres in C. elegans, they are unable to reprogram cells in later
stage embryos without prior removal of inhibitory factors (reviewed
by Joshi et al., 2010). We found that ELT-7 can initiate intestine
gene expression in many differentiated cell types in embryos, larvae,
and adults without removal of inhibitory factors, demonstrating that
not all transcription factors adhere to the MCT. A growing number
of in vitro studies demonstrate that transcription factors can redirect
differentiated cell identity without reversion to a pluripotent
intermediate; however, the context and factors that result in efficient
and stable transdifferentiation of any cell type remain relatively
unclear (reviewed by Ladewig et al., 2013).
We found that only pharynx cells appear to become fully
transdifferentiated and reprogrammed by ELT-7. We considered the
possibility that this reflects tissue-specific expression of the hsp-1641/2 promoter used in the study, which is expressed at somewhat
higher levels in the pharynx (Stringham et al., 1992). However, we
found that longer heat shocks (30 minutes and 60 minutes) also
failed to result in cellular reprogramming outside the pharynx,
although such a regimen results in higher heat-shock promoter
activity in epidermal and muscle cells than in the pharynx under the
brief (15 minutes) heat-shock conditions used throughout this study.
Thus, the cell-type specificity does not appear to be attributable to
differences in expression levels of the transgene.
Transdifferentiation of only pharynx marginal cells implies that
cellular context dictates susceptibility to reprogramming. With the
exception of two cells in the posterior pharynx, the marginal cells are
separated in cell lineage from intestinal cells at the first cell division
(Sulston et al., 1983); therefore, relatedness in cell lineage does not
appear to explain their particular susceptibility to reprogramming by
ELT-7. The intestine and pharynx are both epithelial tubes that are a
part of the digestive tract. It is conceivable that epithelial tube or
digestive tract identity has a role in establishing the context for direct
reprogramming into intestine per se; however, such an effect would
not extend to rectal epithelial cells, as we did not observe evidence for
transdifferentiation in that region. Although it is possible that marginal
cells might be uniquely developmentally plastic, the results of Gilleard
and McGhee (Gilleard and McGhee, 2001), Fukushige and Krause
(Fukushige and Krause, 2005), and Turson and Patel (Turson and
Patel, 2011) imply that differentiated marginal cells are not susceptible
to transdifferentiation into epidermis, muscle and neurons,
4847
Development
Fig. 3. Loss of pharynx-specific markers following a
pulse of ELT-7 expression. Expression of pharynx
muscle-specific ceh-22::GFP (A,B) or myo-2::GFP (C,D)
reporters diminishes by 3 days after ectopic ELT-7
expression compared with non-heat shocked hs-elt-7
control larvae. Scale bars: 5 μm. Exposure time is
indicated in the upper right-hand corner. (E) Decreased
IFA in cells expressing elt-2::lacZ::GFP compared with
non-GFP-expressing marginal cells within the same
section of the pharynx (boxed region shown in lower
panels). Bottom panel, overlay of IFA (red) and elt2::lacZ::GFP (green). Closed arrow indicates non-elt2::lacZ::GFP-expressing marginal cell nucleus; open
arrow indicates GFP-expressing nucleus. Faint green
signal over the non-expressing nucleus is the out-of-plane
signal from the positive nucleus of the third marginal cell.
(F) IFA average pixel intensity minus background was
normalized to the cell with the highest pixel intensity
within each worm. Normalized signal in anterior marginal
cells with strong elt-2 reporter expression (green
highlighted) was lower than in cells with weak or no
expression of elt-2 (white). Each row is an individual
animal and each column indicates the individual marginal
cells ordered based on increasing GFP signal. The
average overall IFA signal is reduced in the elt2::lacZ::GFP-positive marginal cells.
RESEARCH REPORT
Development (2013) doi:10.1242/dev.103010
Fig. 4. Reprogrammed pharyngeal cells have
intestine-specific morphology. Morphology of the
posterior (A) and anterior (B) bulbs of the pharynx in
non-heat shocked hs-elt-7 control animals and
morphology of the intestine (C), posterior bulb (D) and
anterior bulb (E) after heat shock (D, 72 hours; E,
96 hours) are seen using DIC. Boxed region (C′-E′)
outlines elt-2::lacZ::GFP-expressing nuclei in C′′-E′′.
Scale bars: 2 μm. (F-H) Overlay of IFB-2 (red) and elt2::lacZ::GFP (green) signal in the endogenous intestine
(F), and posterior (G) and anterior (H) pharynx 5 days
after ectopic ELT-7. (I-K) Morphology of the intestine and
remodeled pharynx by TEM. (Extensive images of the
ultrastructure of the wild-type pharynx for comparison
can be viewed at wormatlas.org.) (I) Cross-section of
normal L1 larva intestinal lumen. Microvilli (mv) on the
top are seen at nearly full length, whereas those at the
bottom are at oblique angles. Black arrow indicates the
terminal web. Scale bar: 1 μm. (J) Accessory lumen in
the anterior pharynx of hs-elt-7 L1 larva 6 days after
brief ectopic ELT-7 expression. Arrows show remaining
buccal cuticle. Several long microvilli (mv) are seen and
the circular profiles in the middle of the side lumen are
apparently additional microvilli extending at extreme
angles from other edges of the side lumen. Some E. coli
(Coli) lie within the buccal channel. Scale bar: 1 μm.
(K) Higher magnification of ectopic microvilli (mv). Scale
bar: 0.2 μm. Arrowheads indicate an electron-dense
line, an apparent terminal web structure, running
subjacent to the plasma membrane. The microvilli
appear surrounded by diffuse glycocalyx. Nuc, nucleus.
MATERIALS AND METHODS
C. elegans strains, maintenance, synchronization and heat
shock
Transgenic strains used were as follows: JR3410, wIs47 [hsp::end-1];rrIs1
[elt-2::lacZ::GFP] (Fukushige et al., 1998; Zhu et al., 1998); JR3373,
wIs125[hsp::elt-7] (Sommermann et al., 2010); rrIs1; JR3457, wIs125;
CuIs2 V [ceh-22::gfp]; and JR3471, unc-119(ed3); ruIsIII[unc-119(+), myo2::GFP; wIs125. Larvae were synchronized as described previously
(Stiernagle, 2006) and heat shocked at the desired stage in M9 buffer at
33°C for 15 minutes using a thermal cycler.
Immunofluorescence
Antibodies MH33 (anti-IFB-2) (Bossinger et al., 2004) and anti-ELT-2 were
gifts from J. McGhee (University of Calgary, Canada). Anti-IFA (Francis
and Waterston, 1991) was a gift from G. Gunderson (Columbia University,
NY, USA). Cy3 goat anti-rabbit and Cy3 goat anti-mouse were obtained
from Sigma. Fixation and permeabilization were carried out as described
previously (Sommermann et al., 2010). Animals were viewed with a Zeiss
Axioskop 2, Olympus BX60 or Nikon Eclipse Ti inverted microscope and
imaged with a MicroFire camera or Hamamatsu flash Orca 2.8. Brightness
and contrast of some images were modified using NIS Elements software or
PowerPoint to reveal relevant details more clearly in the figures.
IFA intensity measurement
Using NIS Elements software, individual marginal cells in the anterior
pharynx were outlined to create regions of interest (ROIs). Average intensity
of the IFA signal was quantified within each of the three anterior marginal
4848
cells in individual worms that contained one or two GFP-positive nuclei,
bright staining, and non-elongated or abnormal nuclei. We compared only
cells within individual worms, in order to control for differences in staining
efficiency between worms. Background was determined by measuring
average pixel intensity in region outside the worm.
Transmission electron microscopy (TEM)
L1 larvae of strain JR3373 were heat shocked and inspected by fluorescence
microscopy after 6 days. Worms with bright elt-2::lacZ::GFP-expressing
nuclei and intestine-like morphology in the anterior pharynx were placed on
ice, shipped to the Hall lab, and subsequently transferred into a small
planchette for the Bal-Tec HPM-010 High Pressure Freezing Machine using
E. coli as the surrounding matrix. Fixation and embedding procedures
generally followed Hall et al. (Hall et al., 2012). One or two animals were
fast frozen per planchette, after which frozen samples were placed into an
RMC FS-7500 Freeze Substitution unit, in a 1% osmium tetroxide solution
in 98% acetone, 2% dH2O. Samples were held at −90°C for 4 days, then
slowly warmed to 0°C, held for 3 days at 0°C, then rinsed in cold acetone
and gradually infiltrated with EmBed 812 resin. Samples were flat
embedded between Aclar sheets, then cured at 60°C for 2 days. Single
worms were viewed under the dissecting microscope, cut out of the Aclar
sandwich before re-embedding in fresh plastic resin and placed in a mold in
precise orientation before curing again at 60°C. The embedded sample was
serial thin-sectioned on an RMC PowerTome XL using a diamond knife,
mounted on slot grids, post-stained with uranyl acetate and viewed with a
Philips CM10 electron microscope. Digital images were collected using an
SIS camera system and viewed using iTEM or Photoshop software
platforms to analyze data and select images for illustrations.
Acknowledgements
We thank Andrew Sumner for assistance with chemosensation experiments, and
Leslie Gunther-Cummins and Geoff Perumal for help with the Bal-Tec HPF device.
We thank G. Gunderson and J. McGhee for reagents. Some strains were provided
by the CGC, which is funded by NIH Office of Research Infrastructure Programs
(p40 O8010440).
Development
respectively. Rather, these cells may express a factor or set of factors
that allow for transdifferentiation into intestine. Our current
observations establish an in vivo model to investigate the role of cell
context in stable transdifferentiation of somatic cells and reveal
remarkable plasticity in cellular differentiation in an organism with a
rigid pattern of cell division and identity.
Competing interests
The authors declare no competing financial interests.
Author contributions
M.R.R. designed and performed experiments, and wrote the manuscript draft.
A.W. performed experiments. K.C.Q.N. performed transmission electron
microscopy experiments. D.H.H. interpreted the electron micrographs. J.H.R.
conceived the project, and revised and approved the manuscript.
Funding
This work was supported by a training grant from the California Institute of
Regenerative Medicine, and grants from the National Institutes of Health [OD
010943 to D.H.H. and HD062922 to J.H.R.]. Deposited in PMC for release after
12 months.
Supplementary material
Supplementary material available online at
http://dev.biologists.org/lookup/suppl/doi:10.1242/dev.103010/-/DC1
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