September 15, 1999
Learning How Organs Tell Left From Right
From all outward appearances, the human body is symmetrical. If one
were to divide the body into two halves, for example, each side would
have a single arm and leg. But a look inside at the internal organs
shows that there is not perfect symmetry throughout the body. The heart
and spleen normally reside on the left side and the liver and
gallbladder are on the right. Furthermore, the organs themselves are
asymmetrical.
Understanding how genes control the shape and spatial orientation of
organs is one of the goals of developmental biologists. In the mouse,
as in other vertebrates, cells begin receiving and responding to
instructions about their positional fate during the earliest stages of
development, when the embryo resembles little more than a flat sheet of
cells. Two proteins in particular, called Nodal and Pitx2, are produced
predominantly on the left side of the mouse embryo and they appear to
direct the growth of organs in a way that is appropriately
"left-sided." Understanding how these two proteins interact may allow
scientists to understand a great deal more about how organs are
segregated into left-side and right-side.
“Cell proliferation stops. There is a failure to progress past the initial determination of organ identity.”
Michael G. Rosenfeld
In attempting to unravel the genetic program that determines
left/right orientation, Howard Hughes Medical Institute (HHMI)
investigator Michael
Rosenfeld and colleagues at the University of California, San
Diego, and The Salk Institute, engineered knockout mice that lacked the
Pitx2 gene. The mice exhibited a number of developmental
abnormalities, including inappropriate position of the heart and
lungs.
Humans with one defective copy of the Pitx2 gene have a
constellation of problems called Rieger syndrome, the signs of which
include irregularly shaped eyes, lack of tooth growth, craniofacial
deformities, and, more rarely, problems with growth hormone
production.
Rosenfeld's team found that Pitx2 knockout mice had defective
growth in their teeth and pituitary glands, the source of growth
hormone. "Cell proliferation stops. There is a failure to progress past
the initial determination of organ identity," Rosenfeld said.
Summarizing the results of these studies, which are published in the
September 16, 1999, issue of the journal Nature, Rosenfeld said,
"the work shows that the control of the left/right orientation of
organs is more complicated than we thought."
Researchers knew that Nodal is a signal-carrying protein, and Pitx2
is a transcription factor—a protein that turns on other genes.
Once the Nodal signal arrives, it induces Pitx2 to help implement its
directives. One testable issue in the current study, says Rosenfeld,
"was to determine whether Pitx2 implements the entire Nodal program or
whether Nodal acts via additional targets."
The results of earlier experiments that attempted to untangle the
functions of Nodal and Pitx2 were intriguing, but did not produce a
clear answer. When researchers added Nodal or Pitx2 to the right side
of a mouse embryo, for example, the right side developed as a mirror
image of the left. In other experiments, researchers added extra Nodal
or Pitx2 to embryos only to find that the two proteins had similar
effects. And knocking out the Nodal gene alone resulted in
either mirror-image or randomized organization of multiple organs.
Given these results, Rosenfeld says he would have expected the
knockout of Pitx2 to cause the same sort of organ abnormalities
reported by scientists who performed the Nodal knockout
experiments.
The actual result was a mixed bag, but it appears that Pitx2 is
crucial in determining the "leftness of the lung," say Rosenfeld and
his colleagues. Without Pitx2, both sides of the lungs of the
knockout mice looked like the right side of a normal lung. The left
side of a normal pair of lungs is usually smaller to accommodate the
position of the heart.
Studies of the heart orientation in the Pitx2 knockout mice proved
less conclusive. In the Pitx2 knockout mice, the heart was
located on the right side of the body, not in its normal location on
the left side. Despite its improper location, however, the direction of
cardiac looping was normal.
"Everyone might well have predicted the result would be the same as
after removal of Nodal," said Rosenfeld. "But now we know that
this single transcription factor in the pathway does not account for
the whole of left/right asymmetry."
Rosenfeld is now searching for other transcription factors that
respond to Nodal signals. These transcription factors may be involved
in a condition called situs invertus, which affects approximately 1 in
10,000 humans and gives rise to a mirror-image organization of the
internal organs. In many cases the disorder has no adverse
consequences, though it can be associated with heart abnormalities.
Rosenfeld also hopes to begin identifying genes that are switched on
by Pitx2 when organs grow. Such genes are likely to provide additional
insight into understanding left/right determination, he says.
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