Introduction to germ cell development in Caenorhabditis elegans

Abstract

A central feature of the continuum of life in sexually reproducing metazoans is the cycle of the germline from one generation to the next. This volume describes the cycle of the germline for Caenorhabditis elegans through chapters that are focused on distinct aspects or processes in germ cell development. Topics include sequential and dependent processes such as specification of germ cells as distinct from somatic cells, sex determination, stem cell proliferative fate versus meiotic development decision, recombination/progression through meiotic prophase, contemporaneous processes such as gametogenesis, meiotic development and apoptosis, and continuing the cycle into the next generation through fertilization and the oocyte-to-embryo transition. Throughout germ cell development, translational control and epigenetic mechanisms play prominent roles. These different aspects of germ cell development are seamlessly integrated under optimal conditions and are modified in the different reproductive strategies that are employed by C. elegans under harsh environmental conditions. In this chapter, we set the stage by providing a brief background on the C. elegans system and germ cell development, indicating processes in the cycle of the germline that are covered in each chapter.

Figure 1. Schematic of adult C. elegans hermaphrodite and gonad

(Top) Adult C. elegans hermaphrodite, highlighting the reproductive system, which contains two U-shaped gonad arms connected by a common uterus. The distal end of each gonad arm is capped by a somatic distal tip cell (DTC) that covers the distal end of the germline, containing the proliferative zone (yellow). A surface view of the left side U-shaped gonad arm shows the five pairs of somatic gonadal sheath cells covering the area from the transition zone to the spermatheca. On the right side, the gonad arm is shown without the sheath cells. The green cells represent the germ cells in meiotic prophase I, the purple cells represent the developing oocytes, the proximal darker blue area is the spermatheca, and the clear embryos are found within the uterus. (Bottom) A detailed view of one adult hermaphrodite gonad arm is shown. The upper part of the arm is shown as a surface view without the covering sheath cells. The transition zone is visible as a light green color. The lower part of the gonad is an internal view of the proximal region including the sheath cells. The oocytes closest to the loop are connected to the central rachis (light blue, also see Figure 4), which allow cytoplasmic material to enter, while the proximal 4 – 5 oocytes closest to the spermatheca are fully cellularized. See text and wormatlas.org < http://www.wormatlas.org/ > for details.

Figure 2. Morphology from Nomarski differential interference contrast (DIC) microscopy

Live C. elegans adult hermaphrodite showing an interior view of the U-shaped gonad arm in the posterior half of the worm. The gonad is outlined in dark gray where the distal tip cell caps the distal end and developing oocytes populate the proximal end. In the distal half of the gonad, on the dorsal surface, germ cell nuclei are situated along the outer surface of the gonadal tube whereas the central rachis is devoid of germ cells/nuclei. Germ cells move from the very distal end to the proximal end via bulk flow. Overt oocyte development occurs from the loop region into the proximal half of the gonad on the ventral side. The spermatheca lies adjacent to the final oocyte. As an oocyte is ovulated into the spermatheca, a sperm fertilizes it and early development initiates in the uterus. The developing embryos are then released through the vulva to the external environment, where the remainder of embryogenesis occurs. Also see Figure 4.

Figure. 3. Cycle of the germline: hermaphrodite gonadogenesis

Gonadogenesis begins at L1 stage with four gonad precursor cells. Z2 and Z3, the germline progenitors, are sandwiched between the somatic precursors Z1 and Z4 (red), which will divide and form the DTCs and the remaining somatic gonad cells. Prior to the L3 stage, all germ cells proliferate mitotically (yellow). Beginning in early L3, germ cells that are farthest from the DTC leave the proliferative zone, enter meiotic development (green) and progress through meiotic prophase and gametogenesis in an assembly-line fashion. Germ cells that switch from the proliferative fate to meiotic development at this point will become sperm, by the end of the L4 stage in the proximal end of the gonad arm, while those that switch in L4 and adulthood will become oocytes. Migration of the two DTCs generates the U-shape of the gonad: starting in the L3 stage, the DTCs each migrate away from the centrally located gonad primordium along the ventral surface; at the L3/L4 molt, the DTCs each migrate centripetally to the dorsal surface; and in the L4 stage, each DTC migrates back toward the center of the animal. Germ cell proliferation and gametogenesis mediated germ cell volume expansion fills the gonadal tube as the DTCs migrate. Gonads in the schema are for illustrative purpose and are not to scale.

Figure 4. Fluorescence image of C. elegans adult hermaphrodite

Live C. elegans hermaphrodite highlighting the U-shaped gonad arm in the posterior half of the worm. The gonad is outlined in yellow where the distal tip cell caps the distal end and developing oocytes populate the proximal end. The strain, OD95 (Green et al. 2011), allows visualization of plasma membranes (green), with a GFP tag fused to the PH-domain from PLC delta, expressed in the germline and early embryo from the pie-1 promoter, and chromatin (red), with an mCherry tag fused to histone H2B, also expressed from the pie-1 promoter. Interior view shows, in the distal half of the gonad, germ cells that are situated along the outer surface of the gonadal tube with openings to the central rachis that is largely devoid of germ cells/nuclei. A single file row of growing oocytes in diakinesis is shown in the proximal half of the gonad. Asterisks indicate highly condensed nuclei that are indicative of apoptosis.

Fig. 5. Life Cycle of C. elegans and Potential Diapauses

C. elegans adults lay embryos that pass through gastrulation, comma stage, 2-fold and 3-fold embryos before hatching in the L1 larval form. The larvae develop through L2, L3, and L4 molts before becoming adults. The cycle takes about 3.5 days at 20°C under rich nutritional conditions. When the food source becomes scarce, worms at different stages of development can enter diapause where reproductive development is halted and their metabolism is slowed (red arrows). If L1 larvae hatch in the absence of food, they enter L1 diapause, blocking initiation of further development. When food becomes available, L1 larvae reinitiate development. If L2 worms encounter environments with reduced/no food, overcrowding and/or high temperatures, they enter an alternate developmental form called dauer, where development of the reproductive system is arrested, which is resistant to stress and desiccation but is motile and active in searching for food. If the dauer finds food, then the worm enters the developmental program in the L3 stage and proceeds to adulthood. The third diapause occurs when L4/adults find themselves without food. The adult reproductive diapause halts reproduction by degrading most of the germline, but leaving proliferative zone cells that are apparently cell cycle arrested. Upon refeeding, worms in adult reproductive diapause resume germ cell proliferation, meiotic development and oogenesis and can become fertile. Bagging is another strategy to overcome starvation conditions. Here the adult worm stops laying embryos, and any embryos within the mother continue to develop. At hatching, these embryos eat the mother from the inside out. The mother does not survive, but ensures that the embryos have enough food to reach larval diapauses. See Hubbard et al (2012)(Chapter 5) for further discussions of the response of the reproductive system to alternative environments.