166 ■ CHAPTER 09 What Genes Are
GENETICS
however, changing a few nucleotides in a gene’s
DNA sequence has little or no effect. In such
cases, a mutation is said to be “silent” because
it produces no change in the function of the
protein, and therefore no change in the pheno-
type of the organism. (We further explore muta-
tions and their effect on the resulting proteins in
Chapter 10.)
Insertions and deletions can be point muta-
tions, but they can also involve more than one
nucleotide; sometimes thousands may be added
or deleted. Large insertions and deletions almost
always result in the synthesis of a protein that
cannot function properly. Güell’s goal was to
use CRISPR in the pig genome to delete bits of
DNA in the pol gene so that the PERVs could
no longer function. The pol gene is essential for
PERVs to replicate, so a large mutation should
halt the formation of viral particles. In addition
to being essential to the virus, pol is present
in all 62 copies of PERVs but not elsewhere in
the pig genome, so targeting CRISPR to mutate
pol would spare damage to other parts of the
pig DNA. “My biggest concern is always not to
destroy the genome,” says Güell. So, he and the
team designed two CRISPR RNA guides to
target the complex right to pol, and to mutate the
heck out of it.
After much trial and error, the team
inserted its CRISPR-Cas9 construct into pig
embryos, and the system began its work. The
CRISPR system made 455 different cuts in
the PERV pol genes throughout the genome,
resulting in deletions ranging from 1 to 148
base pairs. About 80 percent of the engineered
mutations were small deletions of fewer than
nine base pairs—tiny, but enough to disrupt
the gene.Pigs Are People Too?
In October 2015, Güell, Church, and the team
presented the results of their initial work:
Using CRISPR, they had inactivated 62 PERVs
in pig embryos. Recall that in previous non-
CRISPR-edited experiments, PERVs from the
pig cells infected human cells when put in closeproximity. This time, when the team placed the
CRISPR-edited pig cells next to human cells,
there was zero transmission of the virus. “The
gene editing completely disrupted the ability of
the virus DNA to replicate and form viral parti-
cles,” says Güell.
Although the team has successfully destroyed
the PERV genes in the pig genome, there are still
a few more steps before pig organs will be ready
and safe for human transplantation. Other
genes need to be manipulated, says Güell, such
as immune system proteins so that the human
immune system doesn’t reject the organs as
foreign. However, the next immediate step, says
Güell, is to edit an embryo, implant it back into
a female pig, and raise a live, genetically engi-
neered animal. “We’re trying hard,” says Güell.
“It could be in the next year.”
Other researchers are pursuing a different
path: growing actual human organs in pigs.
Again, CRISPR is the workhorse for the experi-
ment. In 2016, a team at UC Davis used CRISPR
to remove the genes that encode for the pig
pancreas in a pig embryo. The procedure created
a void in the embryo, which they then filled with
human stem cells. Stem cells have the poten-
tial to develop into almost all human cells and
organs—including, in this case, a fully functional
human pancreas.
Some research groups are trying to use this
method to grow other organs. Figure 9.12
shows the process by which scientists could
use CRISPR and human stem cells to grow a
pig with human kidneys, which would then be
used to donate a kidney to a patient in need of a
transplant. No researchers have yet to complete
this process, and the work remains controver-
sial, especially because of concerns that the
human cells might migrate to the developing
pig’s brain and make the pig, even in some small
way, more human.
Overall, CRISPR has revitalized the idea of
a safe, clean, limitless source of healthy organs.
“It opens up the possibility of not just transplan-
tation from pigs to humans but the whole idea
that a pig organ is perfectible,” Church told the
BBC in 2016. “Gene editing could ensure the
organs are very clean, available on demand,
and healthy, so they could be superior to human
donor organs.”