STEM CELLSMobile elements control stem cell potency
By Hidetoshi Hasuwa and Haruhiko Siomi

GRAPHIC: A. KITTERMAN/SCIENCE

Embryonic stem cells (ESCs) are of greatinterest for tissue replacement therapies and for the study of mammaliandevelopment. Cultured ESCs as well as
induced pluripotent stem cells (iPSCs),
which are generated from somaticcells, can self-renew indefinitely and giverise to all cell types of the body including
the germline. However, such stem cells are
not totipotent because they are inefficient
at generating extraembryonic tissues, suchas the placenta (1). What underlies this developmental restriction? On page 596 of thisissue, Choi et al. (2) report a mechanismthat prevents totipotency. The barrier involves mobile genetic elements embedded
throughout the genome and a small noncoding microRNA (miRNA).
Transposable elements are DNA insertions in the host genome that can rewire
gene expression networks and drive genome evolution and speciation (3, 4).
This is achieved because transposable
element sequences carry potential regulatory information such as transcription
factor–binding sites, and also because thesemobile elements are targets for epigenetic
modifications that can spread to flanking
regions and influence the expression ofnearby genes. Recently, a rare transient cell
population (~2%) within mouse ESC andiPSC cultures was shown to express highamounts of a transposable element known
as a murine endogenous retrovirus withleucine transfer RNA primer (MERVL) (5).
These stem cells also expressed transcripts
normally restricted to totipotent blastomeres of the two-cell (2C) embryo, andlacked three pluripotency factors: octamerbinding transcription factor 4 (Oct4), sex determining region Y-box 2 (Sox2), and Tir Na
Nog (Nanog). In 2C-stage mouse embryos,
more than 25% of the nearly 700 MERVL
copies are transcribed, representing 3% of
the total transcriptional output. Moreover,
307 protein-coding genes generate 626 chimeric transcripts containing MERVL sequences, suggesting that transcription ofmany genes in 2C-stage embryos is initiatedby MERVL elements. Remarkably, this rare
subpopulation of ESCs and iPSCs had theDepartment of Molecular Biology, Keio University School ofMedicine, Tokyo 160-8582, Japan. Email: awa403@keio.jptotipotent potential to produce both extraembryonic and embryonic cell lineages (5).
This highlights a critical role for mobile
elements in cell fate regulation by altering
the profile of transcribed genes, and also
indicates the existence of a molecular barrier that blocks the activity of MERVL andprevents pluripotent ESCs and iPSCs fromentering a totipotent state.
The efficiency of iPSC generation fromsomatic cells is decreased if a miRNA
called miR-34a is expressed, suggestingthat miR-34a antagonizes reprogramming(6). miRNAs are small noncoding RNAs
that associate with complementary sites on
messenger RNAs (mRNAs) to repress theirtranslation and promote their decay (7).
These small noncoding RNAs contributeto cell fate decisions by controlling regula-tory circuits that promote positive or negative feedback of gene expression (8). Choiet al. observed that miR-34a deficiency
induces MERVL expression in cultured
mouse ESCs (derived from the inner cellmass of the blastocyst embryo stage) andiPSCs. The authors found that teratomas
generated from miR-34a–deficient ESCsand iPSCs not only contain derivatives ofembryonic cell lineages but also display
features of extraembryonic trophoblastlike cells of the placenta (see the figure). To
corroborate these findings during early embryogenesis, Choi et al. microinjected miR34a–deficient ESCs [labeled with green
fluorescent protein (GFP)] into recipientmorula-stage mouse embryos ( just prior
to the blastocyst stage) and generated chimeric blastocysts. By tracing GFP-positive

Developmental potential
A microRNA (miR-34a) and embedded mobile genetic elements (MERVL) play a role
in restricting stem cells from generating nonembryonic (placenta) tissues.

Blastocyst

Somatic cell

Artifcialreprogramming

Inner cellmass

Induced pluripotent
stem cellEmbryonicstem cellTotipotent

Pluripotent
miR-34a is present

miR-34a is defcient

Gata2

MERVL

Gata2

Increased
developmental potential

Restricted
developmental potential• Embryonic cells• Nonembryonic cells• Embryonic cells

SCIENCE sciencemag.org

10 FEBRUARY 2017 • VOL 355 ISSUE 6325

Published by AAAS581

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A microRNA suppresses transposons and bars pluripotent stem cells from totipotency

INSIGHTS | P E R S P E C T I V E S

ylase as a negative regulator of MERVL in
2C blastomeres (9). The Gata2 transcript
harbors potential miR-34a binding sites,and Choi et al. observed that Gata2 mRNAamounts increased in miR-34a–deficient
iPSCs and decreased when miR-34a wasoverexpressed. Furthermore, decrease in
Gata2 in miR-34a–deficient ESCs abolisheddifferentiation into both embryonic and extraembryonic cell lineages. Thus, miR-34a
prevents pluripotent ESCs and iPSCs fromentering a totipotent state by repressing
Gata2, which otherwise activates the expression of MERVL.
It is not yet known how MERVL affects
the process that determines pluripotent
and totipotent cell states. It may be that
proteins encoded by these elements, which include reverse
transcriptase, and fusion proteins between MERVL andproximal genes are the key.
Approximately 700 copies ofMERVL exist in the mouse
genome; therefore, it is oneof the most abundant proteincoding genes in the genome.
It was recently shown that aminor population of “naïvelike” cells exists among human ESCs that express humanendogenous retrovirus type-H(HERV-H) transcripts (10).
These naïve cells are capable only of embryonic cell fates. In contrast to mouse ESCs,
human ESCs normally adopt a “primed”
pluripotent state, meaning that they can develop embryonic cell fates but not germline
cells. The expression of HERV-H appears todefine naïve ESCs; however, it is not known
how the HERV-H expression changes between naïve and primed cell states. Nonetheless, the findings of Choi et al. suggestthat transposable elements have been coopted to give rise to totipotent cells for early
host development. j“It is not yetknown how
MERVL affects
the process that
determines
pluripotent
and totipotent
cell states.”

582

REFERENCES

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10.1126/science.aam6589

BIOPHYSICS

Flipping
nanoscopy
on its head
Fluorescent imaging withminimal photon flux canachieve single-nanometer
resolution
By Jie Xiao1 and Taekjip Ha1,2,3,4

A

bout the smallest object we cansee with the naked eye is our own
hair. With a magnifying glass, wecan see about 10 times better, andlight microscopy, until relatively recently, could resolve features about
300 times thinner than human hair (~250nm). Recent developments in fluorescence“nanoscopy” made it possible to routinely
image cellular structures at 20- to 30-nm
resolution (1), but a gap remained at themolecular scale: Most proteins are smaller
than 5 nm across. On page 606 of this issue, Balzarotti et al. (2) report a new concept in nanoscopy, termed MINFLUX, thatachieves the true molecular resolution(2 to 3 nm) and dramatically reduces thenumber of photons required by “flipping” acommon wisdom in nanoscopy on its head.
In traditional optical imaging, even avery tiny object such as a single fluorophore(<1 nm) becomes blurred because light diffraction makes it appear much larger—
about half the wavelength of the lightused. Nonetheless, the center of the imaged
fluorescence spot can be determined withextremely high precision (down to 1.5 nm
if 10,000 photons are used) (3). The Abbe
diffraction limit can be overcome by determining the position of, or localizing, one
molecule at a time, with single fluorescentmolecules that can be switched on or off
stochastically (4, 5). This localization-based
strategy, together with other nanoscopy
approaches (6, 7), ushered in the “resolution revolution” that enabled breakthroughbiological discoveries in the past decade (1).
A common wisdom in nanoscopy is tolocalize a molecule near where the signal is
the strongest, which requires high-emission
photon flux that is often limited by the emission rate of the fluorophore. In MINFLUX,Balzarotti et al. devised the opposite strategy, in which they localize a molecule nearsciencemag.org SCIENCE

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ESC progeny cells, the authors observedthat more than 30% of injected miR-34a–
deficient ESCs contributed to the inner cellmass, which gives rise to embryonic celllineages, and to the trophectoderm, whichdevelops into extraembryonic cell lineages.
Thus, miR-34a–deficient ESCs had totipotent cell fate potential in vivo. Moreover,
injected miR-34a–deficient ESCs contributed to multiple, fully differentiated celllineages in the mouse embryo, yolk sac,and placenta at mid-gestation, demonstrating expanded cell fate potential that isfunctionally and molecularly distinct fromwild-type pluripotent ESCs.
As is the case for 2C blastomeres (5), Choiet al. observed that induction of transposable element expression in
miR-34a–deficient ESCs andiPSCs was largely specificto MERVL elements, andthat among the most differentially expressed genes
between miR-34a–deficientand wild-type iPSCs werethose proximal to MERVL
insertion sites, which generate chimeric transcripts containing MERVL sequences.
Overexpression of miR-34a in
miR-34a–deficient iPSCs decreased expression of thesechimeric transcripts. This
raises the question of whether this miRNAaffects cell fate potential by directly or indirectly regulating the activity of MERVL.
Choi et al. delineated a minimal fragment in the MERVL sequence required for
its expression and found that it contained
no substantial sequence complementary tomiR-34a. This suggests a model in whichtranscription of MERVL is by transcription
factors or chromatin modification factorsthat are repressed by miR-34a. Bioinformatic prediction identified binding sites
for 70 discriminatory transcription factors within the MERVL minimal fragment.
Among these factors, only GATA-binding
protein 2 (Gata2) exhibits an expression
pattern similar to that of MERVL during
early preimplantation (2C-stage embryo)development. Mutations in the putative
Gata2 binding site within the minimal fragment reduced its activity in miR-34a–deficient ESCs. Consistent with this, decreasing
Gata2 expression in miR-34a–deficient
ESCs abolished the expression of MERVLand MERVL-proximal genes. Thus, Gata2
directly promotes expression of MERVL
elements and their proximal genes in miR34a–deficient ESCs. However, Choi et al.
found no involvement of epigenetic modifications in this process, which is at odds
with an earlier report of a histone demeth-Mobile elements control stem cell potency
Hidetoshi Hasuwa and Haruhiko Siomi (February 9, 2017)
Science 355 (6325), 581-582. [doi: 10.1126/science.aam6589]

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