Analysis of the Xist RNA isoforms suggests two distinctly different
forms of regulation
Mingchao Ma, William M. Strauss
MCD Biology, University of Colorado, 347 UCB, Boulder Colorado 80309, USA
Received: 28 December 2004 / Accepted: 24 February 2005
The noncoding RNA Xist has been shown to direct
the mammalian dosage compensation pathway.
Expression of the Xist RNA is regulated through an
uncharacterized post-transcriptional mechanism,
thought to involve Xist RNA stability. We have
previously demonstrated that Xist RNA isoforms
contain different 3¢ ends. In this report we analyze
the expression patterns of Xist RNA isoforms and
show the Xist RNA long form (L-isoform) is the
predominant form in early development. Significant
amounts of both the short form (S-isoform) and the
L-isoform were found in the female soma. We also
define the precise sequence structure of the Xist
RNA isoforms 3¢ ends and show the S-isoform and
the L-isoform are structurally dissimilar. Our data
show both the S-isoform and L-isoform are cleaved
from the same primary transcript. However, the S-
isoform is subsequently post-transcriptionally poly-
adenylated, while the L-isoform is not post-trans-
criptionally polyadenylated. Sequence organization
of the L-isoform shows that there are at least five
different nonadenylated L-isoforms in the female
soma and only one in embryonic stem (ES) cells.
This stem cell–and somatic cell–specific processing
may suggest a role for Xist RNA processing in the
regulation of Xist RNA expression.
The large (>17 kb) noncoding RNA Xist/XIST is the
initiator of the mammalian dosage compensation
pathway; the Xist RNA is essential for X-chromo-
some inactivation in females (Gartler and Riggs
1983; Brown 1991; Lyon 1994; Heard et al. 1997;
Goto and Monk 1998; Panning and Jaenisch 1998;
Mlynarczyk and Panning 2000; Avner and Heard
2001; Brown and Chow 2003; Chadwick and Willard
2003; Chow and Brown 2003; Brockdorff and Duthie
1998). The Xist/XIST gene is encoded on an interval
of the X-chromosome called the Xic. Transfer of
yeast artificial chromosome (YAC) DNA spanning
the Xic into male murine ES cells resulted in the
recapitulation of the entire process of X chromosome
inactivation (Lee et al. 1996). Deletions of the Xist
gene result in the inability to silence an X chromo-
some (Penny et al. 1996; Marahrens et al. 1997). The
Xist RNA has been shown to be expressed exclu-
sively from and coat the inactive X chromosome in
somatic tissues (Brown et al. 1992; Clemson et al.
1996). Expression of the Xist gene alone is insuffi-
cient to result in X chromosome silencing instead,
the Xist RNA must physically associate with an X
chromosome, in a developmentally appropriate
context, to direct X chromosome inactivation
(Beletskii et al. 2001).
The expression of the Xist RNA is developmen-
tally regulated in both male and female animals.
Previously we have shown that there are two differ-
ent Xist RNA isoforms that are different at their 3¢
ends (Hong et al. 1999, 2000; Memili et al. 2001). In
male somatic tissue all Xist transcription is lost by an
incompletely characterized repressive mechanism
involving chromatin methylation (Li 2002; Reik et al.
2003). In the pre-implantation embryo both sexes
transcribe Xist at similar rates but the RNA is
unstable, being degraded very rapidly (half-life <
30 min) (Panning et al. 1997; Sheardown et al. 1997).
After implantation the RNA is stable (half-life > 8 h)
in females (Panning et al. 1997; Sheardown et al.
1997). The post-transcriptional mechanism that
determines the shift in Xist RNA stability is un-
known. A number of models have been put forward
to explain the change in Xist RNA stability and
consequent upregulation of Xist expression (Brown
and Chow 2003). In this article we present molecular
Correspondence to: William M. Strauss; E-mail: William.
DOI: 10.1007/s00335-004-2464-3 Volume 16, 391–404 (2005) Ó Springer Science+Business Media, Inc. 2005