The Scientist - 03.2020

donors didn’t seem to be a mosaic. “This one person really stood out,”

recalls Tukiainen. Across all her tissues, “the active X chromosome

was always the same.”

Tukiainen checked her data several times to confirm it wasn’t

an artifact. Such skewing of the mosaic can happen naturally in

blood, especially as women age and some cell lineages die out, but

this was an extreme example, in a woman who died of asphyxi-

ation at the age of 21. And it was a lucky break for Tukiainen’s

research. She could easily identify escapees by looking for genes

that expressed the alleles of both X chromosomes, to compare

expression patterns in 16 different tissues.

In this woman, 23 percent of 186 genes assessed in the GTEx

dataset escaped. Of those, 43 percent escaped in a majority of tis-

sues, while 11 escaped in only one tissue. “The thing that really

amazed me is how heterogeneous X inactivation escape really is,”

says Tukiainen, who now runs a lab at the University of Helsinki’s

Institute for Molecular Medicine Finland.

Genetic clues to why X genes escape

Escapee lists provide important clues to what Brown says is

now the big question in the field: How do these genes avoid or

reverse silencing?

Researchers suspect there are genetic markers that help main-

tain silencing or promote escape, and patterns in the sequences of

known escapees suggest what the markers might be. For exam-

ple, says Brown, sequences rich in long, repeating sequences called

LINEs tend to be silenced, while those with shorter repeat elements

(SINEs) are more likely to escape. In addition, geneticist Christine

Disteche and colleagues at the University of Washington reported in

a 2015 paper that a transcription factor called CCCTC-binding fac-

tor (CTCF) clustered around escapees and their promoters, leading

the researchers to propose that the factor might promote esca pe.^8

In addition to sequence-gazing, scientists use genetic manipu-

lation experiments to move known escapees and normally silent

sequences around the X chromosome, in search of factors that the

genes carry with them. For example, molecular biologist Laura

Carrel and colleagues at the Penn State College of Medicine relo-

cated the mouse escapee Kdm5c (which also evades silencing in

humans), along with flanking genes that are normally inactivated,

to parts of the mouse X chromosome that are normally silenced.

Kdm5c persisted in escaping. The flanking genes, meanwhile,

maintained their silence.^9 The individual genes appeared to rely

on local sequences that block or favor escape, rather than depend-

ing on their position on the X chromosome to control expression.

Further experiments pointed to genetic sequences that act as

barriers between expressed and silenced regions. In a different

cell line with an out-of-place sequence that included Kdm5c, the

DNA was truncated, deleting everything after Kdm5c including the

downstream, normally inactivated gene. Kdm5c still escaped, but

so did three genes downstream of its insertion site.^10 That is, once

one gene found its voice, it passed on the ability. Normally, scien-

tists presume, some barrier between genes prevents this.

CTCF might serve as one such barrier when it binds between

escapees and silenced genes, as Disteche’s group reported in a 2005

paper.^11 Indeed, CTCF binds just upstream ofKdm5c. Brown’s group

and collaborators found evidence for another candidate escape marker

X ESCAPE PATTERNS IN HUMANS

There are about 1,150 known genes on human X chromosomes. Genes in the pseudoautosomal regions (PARs) at an

X chromosome’s tips pair with corresponding genes on the Y chromosome in XY cells and are expressed from both X

chromosomes in XX cells. Outside of these regions, hundreds of genes are inactivated to avoid giving XX cells a double

dose of genes that XY cells have in only one copy. However, dozens of X-chromosome genes escape this silencing.

Escape

X chromosome

Variable

PAR1 Silenced

(n=22)

Escape
(n=29)

Mostly escape

(n=26) Mostly

silenced

(n=131)

Variable

(n=91)

Silenced

(n=331)

PAR1 PAR2

0 50 100 150 Mb

THE SCIENTIST

STAFF

GENE EXPRESSION PATTERNS ON THE

“INACTIVE” X CHROMOSOME

DISTRIBUTION OF ESCAPEES ON THE

“INACTIVE” X CHROMOSOME