The beginning of the beginning for the germline
Every cell in an individual's body contains identical genetic information,
but only a very few are able to use that information to contribute directly
to the creation of a new individual. Such cells, known as germ cells,
differ from the somatic cells that make up the rest of the body in several
very significant ways. Unlike somatic cells, the germ cells (which include
both eggs and sperm) have the ability to shed and reacquire their epigenetic
markings; to divide meiotically, thereby halving the normal complement
of chromosomes; and to fuse into a zygote that can then give rise to every
type of cell needed for the development of a new individual, a developmental
capacity known as totipotency.
Germ
cells in many species inherit their unique identities very early in development.
In mice, the process begins with a small cohort of a few dozen primordial
germ cells, or PGCs, which appear at the region that separates the extraembryonic
tissue from the embryo proper, then migrate inward, traversing the hindgut
endoderm on their way to colonize the gonads. The journey of these germline
founders begins very early in the embryo's history, at a stage when the
nascent mouse looks more like a tiny cylinder clinging to the uterine
wall. It is believed that the germ cells' destiny is determined in large
part in these early hours, with their initial segregation from their neighbors
and the repression of signals that would otherwise prompt them to adopt
a somatic fate.
In an article published in the 14 July 2005 edition of Nature
magazine, Mitinori Saitou (Team Leader, Laboratory for Mammalian Germ
Cell Biology), Yasuhide Ohinata and colleagues, working in collaboration
with labs in the UK and the US, reported the identification of a new factor
in the specification of PGCs. This molecule, Blimp1 (for B-lymphocyte
induced maturation protein) appears to function earlier than any known
factor in the determination of the germline in mice. A screen of cDNAs
from individual PGCs at embryonic stage 7.5 yielded a number of promising
candidates for germ lineage determinants, including Blimp1, first identified
as a factor in the differentiation of B cells into immunoglobulin-secreting
plasma cells, which circulate in the bloodstream and function at the frontlines
of the body's immune response. "We thought it was intriguing to find
that a factor functioning in such a terminal event as B cell differentiation
might also be at work in the very earliest days of the embryo's development,"
notes Saitou, "It's always interesting to find cases where the same
molecule turns up in two very different contexts."
The team next tracked Blimp1-positive cells by in situ hybridization,
which revealed Blimp1 expression in a thin swath of cells lying
at the border between the extraembryonic ectoderm and the embryo itself.
The onset of its expression was unexpectedly early, in the E6.25-stage
epiblast, even before the process of gastrulation begins. Studies in which
Blimp1 expression was monitored by a fluorescent protein expressed
under the control of Blimp1 upstream elements confirmed that
its expression began in a small number of cells in the epiblast which
proliferated to about 40 cells by E7.5, nicely matching previous estimates
of the number of founder PGCs. On tagging the Blimp1-expressing
cells with the antibody for a second protein, stella, which is a definitive
marker of nascent PGCs, the group found an almost perfect coincidence
of the two proteins' expression, pointing to the strong involvement of
Blimp1.
Turning next to Blimp1 function, Ohinata et al created homozygous and
heterozygous knockouts of the gene that let them study its full and partial
loss of function. The effects of Blimp1 deficiency were broadly dose-dependent,
with slightly more than half the normal number of PGCs in Blimp1+/-
embryos and no normal-appearing PGCs in the null mutants, an effect that
seemed to be linked to the failure of PGC genesis, rather than to cell
proliferation or survival.
Even those cells that did arise in the loss-of-function mutants behaved
differently than their wildtype counterparts, with stunted proliferation
and failure to disperse and migrate being typical behavior of the mutant
PGCs. The gene expression profiles of the Blimp1-deficient PGC-like cells
were also abnormal. In normal development, PGCs begin to express stella
while at the same time inhibiting the expression of Hox-family
genes; in the mutants, few stella-positive cells were detected
and Hox gene expression was prevalent.
"This is the earliest anyone has been able to trace back the origins
of the mammalian germ cell lineage," says Saitou, "we're still
not sure exactly how Blimp1 is causing this small cluster of cells to
adopt this highly specialized fate, but we're looking forward to finding
out whether there's any link to how it functions at the molecular level."
