COM M E N T Open Access
Emerging links between m
6
A and
misregulated mRNA methylation in cancer
Samie R. Jaffrey
1*
and Michael G. Kharas
2*
Editorial summary
N
6
-methyladenosine (m
6
A) in mRNA has emerged as a
crucial epitranscriptomic modificatio n that controls
cellular differentiation and pluripotency. Recent
studies are pointing to a role for the RNA methylation
program in cancer self-renewal and cell fate, making
this a new and promising therapeutic avenue for
investigation.
m
6
A, an epitranscriptomic mark that influences
cellular differentiation
One of the hallmark features of cancer is misregulated
gene expression. A newly recognized concept in the
regulation of gene expression is that mRNAs contain a
diverse set of modified nucleotides, and the location and
identity of these modifications within the transcriptome
constitute an ‘epitranscriptomic’ code. The initial con-
cept of the epitranscriptome was introduced as a result
of transcriptome-wide mapping of N
6
-methyladenosine
(m
6
A), which revealed that m
6
A is found in at least a
quarter of all mRNAs, typically near stop codons [1].
RNA methylation is mediated by a multiprotein ‘writer’
complex comprising RBM15–WTAP–METTL3–
METTL14 [2]. METTL3 is the sole methyltransferase re-
sponsible for forming m
6
A, whereas RBM15 couples the
methylation complex to mRNA to methylate adjacent
m
6
A residues [2]. WTAP acts as an adaptor, coupling
RBM15 to METTL3, whereas METTL14 positions RNA
substrates for methylation by METTL3. Notably, adeno-
sine methylation is reversible. Although FTO (fat mass
and obesity-associated protein) was reported to be an
m6A demethylase, it is now known that AlkB family
member 5 (ALKBH5) is the only enzyme to show physio-
logically relevant demethylation activity in vivo [3].
* Correspondence: srj2003@med.cornell.edu; kharasm@mskcc.org
1
Department of Pharmacology, Weill Cornell Medical College, Cornell
University, New York, NY 10065, USA
2
Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center,
New York, NY, USA
Under normal conditions, the most prominent effect
of the presence of m
6
A is to induce mRNA degradation,
but, in response to certain types of cellular stress, the
m
6
A distribution across the transcriptome can change,
with the most notable effect being increases in the abun-
dance of m
6
A marks in the 5′-untranslated region of se-
lect mRNAs [4]. This methylation confers to the mRNA
the ability to be translated in a manner that does not re-
quire the canonical cap-binding protein eIF4E [4].
eIF4E-independent translation is activated in diverse dis-
ease states, especially cancer.
A connection between m
6
A and cancer-relevant pro-
cesses is suggested from studies linking m
6
A to differen-
tiation pathways that control stem cell fate [5].
Pluripotent stem cells depleted of m
6
A show mark ed re-
sistance to stimuli that promote differentiation. These
cells retain pluripotency markers and fail to acquire gene
expression patterns seen in differentiated cells. By con-
trast, primed stem cells, which lack the ability to con-
tribute to blastocyst chimeras, are more prone to
differentiate and show enhanced and abnormal expres-
sion of differentiation markers upon depletion of m
6
A
[5]. These studies show that alterations in m
6
A levels
can alter differentiation pathways. As the pathways in-
volved in embryonic stem cell maintenance and differen-
tiation have been directly linked to the acquisition of
stem cell properties in both solid and hematological ma-
lignancies, m
6
A alterations might have a role in cancer
development (Fig. 1). Hypoxic environments and dysreg-
ulation of hypoxia-inducible factors (HIFs) have been
implicated in a variety of cancers, including brain, lung,
pancreatic, colon, ovarian, and many other cancers.
ALKBH5 and m
6
A depletion as a driver of cancer
stem cell formation
In line with the above, recent studies point to a link be-
tween alterations in m
6
A levels and the abnormal cellu-
lar differentiation states present in cancer. In a variety of
tumors , cancer stem cell populations are readily detected
in hypoxic niches. Semenza and colleagues showed that
© The Author(s). 2017 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
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reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
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Jaffrey and Kharas Genome Medicine (2017) 9:2
DOI 10.1186/s13073-016-0395-8
hypoxia was associated with increased breast cancer
stem cell formation and elevated levels of ALKBH5 in
breast cancer [6]. Notably, ALKBH5 was required for ac-
quisition of the breast cancer stem cell phenotype. The
authors observed that hypoxia increased the stability of
NANOG mRNA and its protein levels. This could reflect
ALKBH5-mediated NANOG mRNA demethylation or
an indirect effect of ALKBH5 expression on m
6
A levels
in NANOG mRNA [6]. NANOG is a key transcription
factor that is associated with pluripotency. In addition to
the effects of hypoxia on NANOG mRNA, hypoxia in-
duces the expression of the zinc-finger protein ZNF217,
which inhibits nuclear methylation [6]. Induction of
ZNF217 also reduces m
6
A levels in NANOG, as well a s
in KLF4 [7]. The KLF4 transcription factor is a pluripo-
tency factor required for the maintenance of breast can-
cer stem cells. Thus, hypoxia reduces m
6
A le vels to
promote the formation of breast cancer cells (Fig. 1).
m
6
A elevations in hematologic malignancies
While hypoxia decreases m
6
A levels, increases in the
abundance of m
6
A might also predispose to cancer. This
is supported by evidence that genes that encode proteins
that contribute to the RNA methylation complex are
upregulated in myeloid leukemia (Fig. 1). Analysis of
The Cancer Genome Atlas (TCGA; https://cancergen-
ome.nih.gov/) shows that METTL3 , METTL14, and
RBM15 are highly expressed in myeloid leukemia com-
pared with other cancers. These proteins appear to be
required for maintaining the abnormal differentiation
state seen in myeloid leukemia. A role for m
6
A in mye-
loid leukemias is supported by studies of WTAP deple-
tion. Bansal and colleagues found that WTAP expression
was ele vated in cells derived from 32% of patient s with
acute myeloid leukemia [8]. WTAP knockdown results
in reduced proliferation, increased differentiation, and
increased apoptosis in a leukemia cell line [8]. WTAP
knockdown is a highly efficient approach to deplete m
6
A
from mRNA. Thus, m
6
A depletion might account for
the anti-leukemia effe cts observed upon WTAP
depletion.
RBM15, another component of the m
6
A writer com-
plex, is also linked to myeloid leukemia. In this case,
RBM15 has a clear driver role in the development of
hematologic malign ancy. Acute megakaryoblastic leuke-
mias were shown to be mediated by a chromosomal
translocation t(1;22) of RBM15 (also called OTT1) with
the MAL gene [9]. RBM15 has crucial roles in
Fig. 1 Cancer can be promoted by upregulating either N
6
-methyladenosine (m
6
A) demethylases or methyltransferase proteins. In breast cancer,
hypoxia increases the expression of ALKBH5 or ZNF217 through the activation of hypoxia-inducible factors (HIFs). ALKBH5 is an m6A demethylat-
ing enzyme, and ZNF217 inhibits the RNA methylation writer complex (RBM15–WTAP–METTL3–METTL14), resulting in a reduction of the levels of
the m
6
A modification in the mRNA of breast cancer pluripotency transcripts NANOG and KLF4, promoting their stability and increased expression.
This contributes to the reacquisition of the breast cancer stem cell phenotype in these cells. In myeloid leukemia, by contrast, increased levels of
components of the m
6
A methylation machinery proteins (RBM15–WTAP–METTL3–METTL14) are present, suggesting misregulated and increased
mRNA methylation. Thus, the increase in these proteins might alter the normal differentiation trajectory of hematopoietic stem cells, leading to
abnormal fates, including leukemic blasts. (Arrows indicate activation; ‘lightning bolts’ indicate misregulation of the RNA methylation program)
Jaffrey and Kharas Genome Medicine (2017) 9:2 Page 2 of 3
maintaining quiescence in hematopoietic stem cells and in
megakaryocyte leukemia cell line differentiation by con-
trolling the splicing of key differentiation genes, including
GATA1, RUNX1, TAL1,andc-MPL [10]. Because RBM15
directs m
6
A formation in the transcriptome [2], the onco-
genic effects of RBM15 overexpression and RBM15-MAL
translocation might reflect aberrant m
6
Aformation.
Although each of the major proteins in the m
6
A methy-
lation complex—that is, RBM15, WTAP, METTL3, and
METTL14—show alterations in myeloid leukemias, de-
finitive demonstration of the role of m
6
Awillrequire
mechanistic evidence linking m
6
A alterations to leukemia
phenotypes in these cancers.
Conclusions
The modification m
6
A is an epitranscriptomic mark that
influences a wide variety of RNA processing steps, in-
cluding splicing , mRNA stability, and translation. Genes
associated with pluripotency and lineage-specific dif-
ferentiation are cont rolled by m
6
A levels , and reduced
m
6
A levels can lead to a misregulation of these genes
and the acquisition of stem cell characteristics. Alter-
natively, increases in m
6
A levels are expe cted to
stabilize these transcripts and would therefore be par-
ticularly problematic in tissues that are continuously
replenished from a stem cell population, such as the
hematopoietic lineage. Hematopoietic stem c ells
traverse through distinct differentiation intermediates
in order to achieve their final differentiated state. Ele-
vations in m
6
A might alter the normal differentiation
pathway, resulting in cells being trapped i n a progeni-
tor cell state.
Many unanswered questions remain. How conse rved
are these pathways in other cancer types? Many cancer
subtypes are associated with abnormal differentiation
states or cancer stem cells, making it likely that inter-
ventions that influence m
6
A levels could therapeutically
alter the differentiation program. Will a systematic ana-
lysis of the marked transcripts in cancer reveal new tar-
gets for therapeutic intervention? Can pharmacologic
modulation of the RNA methylation program in various
cancers push cells toward differentiation? Another im-
portant question is whether targ eting m
6
A would have
unwanted side-effects. As m
6
A might be used in every
cell for the regulation of gene expression, targeting m
6
A
might not provide a suitable therapeutic index. Finally,
the high reliance of myeloid leukemia cells on methyla-
tion complex proteins raises the hope that these cells
will show higher sensiti vity to m
6
A pathway inhibitors
than other cell types.
Abbreviations
m
6
A: N
6
-methyladenosine; TCGA: The Cancer Genome Atlas
Authors’ contributions
SRJ and MGK drafted the manuscript, and both authors approved the final
manuscript.
Competing interests
The authors declare that they have no competing interests.
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