In <i>Caenorhabditis</i> nematodes, sperm compete for position in the spermathecae.

<p>(A) Diagram of a hermaphrodite (red) mating with a male (blue). The hermaphrodite gonad is bilaterally symmetric with a central uterus. The male is ejaculating larger sperm (blue) into the uterus, and they outcompete the smaller hermaphrodite sperm (pink) in the race to repopulate each spermatheca (purple) after sperm are dislodged during ovulation. (B) Diagram of a female (red) mating sequentially with two males (one green and the other blue). The female's gonad resembles that of the hermaphrodite in (A). The spicules from the blue male have penetrated the vulva, and he is ejaculating (blue) sperm into the uterus. These sperm will compete with those from the first male (green) for positions in the two spermathecae (purple), where they wait for the chance to fertilize oocytes. Although the sperm from the first male have already taken the best positions (shown in the right spermatheca), they will be displaced into the uterus each time an oocyte is ovulated (shown in the left spermatheca) and must compete with those from the second male to reestablish their positions. Although displacement has been directly observed, additional factors that remain unknown might help influence competition among these sperm.</p

Male and female cues involved in gamete behavior.

<p>The male ejaculates sperm (blue) and seminal fluid (green) into the uterus. The seminal fluid contains signals that activate the sperm so that they extend pseudopods and are able to crawl. The seminal fluid is probably complex and might contain additional signals. In addition, active sperm release small membrane-bound packets (blue circles) that contain major sperm protein (MSP), which stimulates oocytes to mature. MSP also causes the gonad sheath cells (orange) to contract and force oocytes through the distal spermathecal valve into the spermatheca (purple). Finally, the oocytes release a complex mixture of prostaglandins (PGs) that guide the sperm. Each of these signals has the potential to mediate sperm competition and/or cryptic female choice.</p

Hermaphrodites have evolved in three independent lineages in <i>Caenorhabditis.</i>

<p>Only species with sequenced genomes are shown. Androdioecious species (comprised of males and hermaphrodites) are marked with a red symbol, and the others are dioecious (comprised of males and females). The two species in blue are able to interbreed and produce fertile offspring, and the outgroup for the elegans group is orange. Modified from Kiontke et al. <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001916#pbio.1001916-Kiontke1" target="_blank">[14]</a> and Félix et al. <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001916#pbio.1001916-Flix1" target="_blank">[38]</a>. See main text for details on the types of male sperm.</p

Cloning the <i>C. briggsae</i> Fog gene <i>trr-1</i> by SNP mapping.

<p>(A) Genetic map of LG<i>II</i>, showing the location of <i>trr-1</i> based on three-factor crosses (Methods). The positions of potential sex-determination genes on the top are from WormBase, and those of the marker genes are from <a href="http://www.briggsae.org" target="_blank">www.briggsae.org</a>. (B) The region extending from 7100 to 7800 kb and the position of each SNP were downloaded from WormBase. The structure of critical recombinant chromosomes is indicated by color, and the frequency of these recombinants is shown at the right. (C) The predicted genes in the region extending from 7560 to 7720 kb were downloaded from WormBase. (D) The predicted structure of <i>trr-1</i> was downloaded from WormBase, The location of each missense allele is marked by an arrow, and the extent of each deletion by a line.</p

A new <i>C. briggsae</i> Fog gene.

<p>Young adult animals of the following genotypes were photographed using DIC optics. A. Wild-type <i>XX</i> hermaphrodite. B. Wild-type <i>XO</i> male. C. <i>v76 XX</i> female. D. <i>v76 XO</i> Fog male. Expanded images of the boxed regions are shown on the right. Anterior is left and ventral down. The hermaphrodite vulva is indicated with a black arrow, and the male tail with a blue oval. Oocytes are marked with a red “o”, sperm with a blue arrow, and a one-celled embryo in the wild type with and “e”.</p

Evolutionary Change within a Bipotential Switch Shaped the Sperm/Oocyte Decision in Hermaphroditic Nematodes

<div><p>A subset of transcription factors like Gli2 and Oct1 are bipotential — they can activate or repress the same target, in response to changing signals from upstream genes. Some previous studies implied that the sex-determination protein TRA-1 might also be bipotential; here we confirm this hypothesis by identifying a co-factor, and use it to explore how the structure of a bipotential switch changes during evolution. First, null mutants reveal that <i>C. briggsae</i> TRR-1 is required for spermatogenesis, RNA interference implies that it works as part of the Tip60 Histone Acetyl Transferase complex, and RT-PCR data show that it promotes the expression of <i>Cbr-fog-3</i>, a gene needed for spermatogenesis. Second, epistasis tests reveal that TRR-1 works through TRA-1, both to activate <i>Cbr-fog-3</i> and to control the sperm/oocyte decision. Since previous studies showed that TRA-1 can repress <i>fog-3</i> as well, these observations demonstrate that it is bipotential. Third, TRR-1 also regulates the development of the male tail. Since <i>Cbr-tra-2 Cbr-trr-1</i> double mutants resemble <i>Cbr-tra-1</i> null mutants, these two regulatory branches control all <i>tra-1</i> activity. Fourth, striking differences in the relationship between these two branches of the switch have arisen during recent evolution. <i>C. briggsae trr-1</i> null mutants prevent hermaphrodite spermatogenesis, but not <i>Cbr-fem</i> null mutants, which disrupt the other half of the switch. On the other hand, <i>C. elegans fem</i> null mutants prevent spermatogenesis, but not <i>Cel-trr-1</i> mutants. However, synthetic interactions confirm that both halves of the switch exist in each species. Thus, the relationship between the two halves of a bipotential switch can shift rapidly during evolution, so that the same phenotype is produce by alternative, complementary mechanisms.</p></div

Components of the Tip60 HAT complex control the sperm/oocyte decision.

<p>A. Analysis of the PCAF/SAGA HAT complex. B. Analysis of the Tip60/NuA4 HAT complex. The phenotypes are Fog (feminization of the germ line), Emb (embryonic lethal) and Ste (Sterile). <i>C. briggsae</i> genes were identified by BLAST. For details, see methods and <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003850#pgen.1003850.s008" target="_blank">Table S5</a>.</p

TRR-1 acts through TRA-1 to promote spermatogenesis and <i>fog-3</i> expression.

<p>A–D. The null allele <i>tra-1(nm2)</i> is epistatic to mutations in <i>trr-1</i>. Animals in A were produced by <i>trr-1(v76)</i> mothers, except for the <i>cby-15; tra-1</i> control at the bottom. Those in B and C were produced by self-fertilization from heterozygous mothers. Blue colored bars indicate spermatogenesis, pink bars indicate oogenesis, and mixed colors represent animals that make both sperm and oocytes. Abnormal germ lines are gray. E. Western blot showing that the levels of TRA-1¹⁰⁰ are not altered in a <i>trr-1</i> mutant. The absence of TRA-1¹⁰⁰ in a <i>tra-1(v56)</i> mutant served as a negative control. (Full-length TRA-1 at the top is obscured by a non-specific band, marked by a star). F. RT-PCR analyses of hand-picked worms. Each age and genotype was run with independent samples, which are presented side-by-side.</p

Tip60 and the FEM complex play complementary roles in germ line regulation.

<p>A. Knocking down Tip60 genes in a <i>Cel-fem-1(ts)</i> mutant. B. Knocking down Tip60 genes in a <i>Cel-fem-2(ts)</i> mutant. C. Analysis of double mutants using weak alleles of <i>trr-1</i> in <i>C. briggsae</i>.</p>*<p>— <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003850#pgen.1003850-Hill1" target="_blank">[22]</a>;</p>†<p>— The Cby progeny of <i>cby-15 trr-1(v104)/++</i> mothers;</p>‡<p>— The Cby progeny of <i>cby-15 trr-1(v128)/++</i> mothers.</p

Mutations in <i>trr-1</i> and <i>tra-2</i> act synthetically to produce male tails.

<p>A, B. Photomicrographs of typical <i>XX</i> male tails, prepared using DIC optics. The arrows indicate the small, spindly rays typical of <i>tra-2</i> mutants, and the stars mark the large rays seen in <i>tra-2 trr-1</i> double mutants. C. Number of rays detected in single and double mutants. (The wild-type male produces 18 rays). Error bars represent 95% confidence intervals, and the indicated probabilities were calculated using the Mann Whitney U test.</p