Mitosis–meiosis and sperm–oocyte fate decisions
are separable regulatory events
Clinton T. Morgana,b, Daniel Nobleb, and Judith Kimblea,b,c,d,1
aMedical Scientist Training Program,bIntegrated Program in Biochemistry,cHoward Hughes Medical Institute, anddDepartment of Biochemistry, University of
Wisconsin, Madison, WI 53706
Contributed by Judith Kimble, January 16, 2013 (sent for review November 27, 2012)
Germ cell fate decisions are poorly understood, despite their central
role in reproduction. One fundamental question has been whether
germ cells are regulated to enter the meiotic cell cycle (i.e., mitosis–
meiosis decision) and to be sperm or oocyte (i.e., sperm–oocyte de-
toward spermatogenesis or oogenesis, respectively. If two distinct
decisions are used, meiotic entry should be separable from specifica-
two decisions with tools uniquely available in the nematode Caeno-
rhabditis elegans. Specifically, we used a temperature-sensitive
Notch allele to drive germ-line stem cells into the meiotic cell cycle,
followed by chemical inhibition of the Ras/ERK pathway to repro-
meiotic prophase can nonetheless be sexually transformed from
a spermatogenic to an oogenic fate. This finding cleanly uncouples
the mitosis–meiosis decision from the sperm–oocyte decision. In ad-
dition, we show that chemical reprogramming occurs in a germ-line
region where germ cells normally transition from the mitotic to the
meiotic cell cycle and that it dramatically changes the abundance of
key sperm–oocyte fate regulators in meiotic germ cells. We conclude
that the C. elegans mitosis–meiosis and sperm–oocyte decisions are
separable regulatory events and suggest that this fundamental con-
clusion will hold true for germ cells throughout the animal kingdom.
their development, germ cells are regulated to transition from the
mitotic to the meiotic cell cycle, and they are regulated to produce
sperm in males or oocytes in females. A fundamental question in
the germ cell field has been whether the mitosis–meiosis and
If a single decision, germ cells might decide between male-specific
meiotic entry leading to spermatogenesis or female-specific mei-
otic entry leading to oogenesis. If two distinct decisions, germ cells
would be regulated to enter the meiotic cell cycle in a way that is
separable from their sexual fate decision. Major progress has been
made in understanding regulation of the mitosis–meiosis decision
(2), but the sperm–oocyte decision has been less tractable in
most organisms. Here we investigate the relationship between
the mitosis–meiosis and sperm–oocyte decisions in the nematode
Caenorhabditis elegans, in which molecular regulators of the two
decisions can be manipulated independently.
C. elegans adults exist as XO males or self-fertile XX her-
maphrodites. Males produce sperm continuously; hermaphrodites
make oocytes continuously in adults, after generating a limited
is organized linearly along its distal to proximal axis: at the distal
end, germ cells in the mitotic cell cycle occupy the “mitotic zone”
(MZ); more proximally, germ cells enter the meiotic cell cycle and
progress through meiotic prophase; germ cells terminally differ-
entiate as sperm or oocytes at the proximal end (Fig. 1A). Germ
cells move from distal to proximal as they progressively mature.
Regulators of the mitosis–meiosis decision have been identified
distal end (Fig. 1A, red) employsNotch signaling to maintain germ
erm cells make two major cell fate choices. One is a cell cycle
C. elegans Notch receptor GLP-1 (Germ Line Proliferation-1), all
mitotic germ cells enter the meiotic cell cycle and differentiate. A
glp-1 ts (temperature-sensitive) mutant maintains the MZ at per-
missive temperature (15 °C), but, at the restrictive temperature
(25 °C), all germ cells (including germ-line stem cells) enter the
meiotic cell cycle (Fig. 1B). Therefore, glp-1(ts) mutants provide a
Regulators of the sperm–oocyte decision have also been iden-
tified and can be manipulated in C. elegans (3, 4). C. elegans germ
cell sex relies on somatic signaling plus germ cell-specific sperm–
oocyte fate regulatorsthat respond tothesomatic signals(3,5).By
manipulating sperm–oocyte fate regulators with temperature-
sensitive mutants, RNA-mediated interference, or small molecule
intervention, adults making sperm can be transformed to make
oocytes without affecting somatic sex (e.g., refs. 4, 6, 7). Such
a transformation from sperm to oocyte production can be induced
in WT XO adult males or in XX adult hermaphrodites with an
(8). Germ-line sexual transformation does not appear to convert
mature sperm into oocytes or vice versa, but instead switches the
adult tissue from production of a gamete of one sex (e.g., sperm)
into production of the other (e.g., oocyte).
Onesperm–oocyte fate regulatoris the C. elegans homologueof
ERK/MAPK, called MPK-1 (9). We previously found that chem-
ical inhibitors of Ras/ERK signaling can reprogram adults from
mutant background (4) (Fig. 1C). XX puf-8;lip-1 adult germ lines
make sperm instead of oocytes, likely because of the dual loss of
the puf-8 oocyte fate regulator (10) and the lip-1 dual specificity
phosphatase, which leads to hyperactivation of the MPK-1/ERK
sperm fate regulator (9, 11). Importantly, this chemically induced
oogenesis generates functional oocytes that support embryogen-
esis (4). Treatment with Ras/ERK inhibitors therefore provides
a method to manipulate the sperm–oocyte decision quickly (ma-
ture oocytes are seen 24 h after treatment) and independently of
a temperature shift.
In this study, we manipulate the mitosis–meiosis and sperm–
oocyte decisions independently and find that they can be uncou-
pled: germ cells in meiotic prophase can be sexually transformed
from a spermatogenic toan oogenic cell fate. We also find that the
sperm-to-oocyte fate chemical reprogramming occurs in a region
in the distal germ line where cells transition from the mitotic to the
key sperm–oocyte fate regulators. Together these results provide
Author contributions: C.T.M. and J.K. designed research; C.T.M. and D.N. performed re-
search; C.T.M., D.N., and J.K. analyzed data; and C.T.M. and J.K. wrote the paper.
The authors declare no conflict of interest.
Freely available online through the PNAS open access option.
1To whom correspondence should be addressed. E-mail: email@example.com.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.
| February 26, 2013
| vol. 110
| no. 9
Biosystems) in a 7500 Fast Real-Time PCR System (Applied Biosystems). All
assays were normalized to the endogenous control, eft-3. The following
TaqMan assays were used: fog-1(L), Ce02415381_g1; fog-3, Ce02412829_g1;
gld-1, Ce02409901_g1; tra-1, Ce02407051_g1; fem-1, Ce02463926_g1; fem-3,
Ce02457444_g1; gld-2, Ce02408169_g1; rnp-8, Ce02413620_g1; eft-3,
Ce02448437_gH; and mpk-1, Ce02445290_m1.
ACKNOWLEDGMENTS. We thank Anne Helsley and Laura Vanderploeg for
help in manuscript and figure preparation; Dr. R. Lin (University of Texas
Southwestern Medical Center) and Dr. S. Ward (University of Arizona) for
OMA-2 and SP56 antibodies, respectively; Elena Sorokin for reading the
This work was supported by National Institutes of Health Grant GM069454 (to
J.K.). J.K. is an Investigator of the Howard Hughes Medical Institute.
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