REPRODUCTION, GERM PLASM, THE GERM LINE, AND GONADAL DEVELOPMENT

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Reproduction is the main theme of life.  Obviously, without reproductive success an individual (or species) is evolutionarily doomed.  All of the organs of the body and their functions may be viewed as structures and processes that enable us to survive in order to reproduce.  In this sense each adult individual is an elaborate device for producing eggs or sperm and for bringing them together in the process of fertilization.

            Sexuality (fertilization) has evolved because (1) it permits a substantial reshuffling of genes (via random segregation and recombination), and (2) it provides a mechanism by which the diploid state can be reproduced after the genetic diversity has been created. This allows an increased genetic variability in populations, and is thus advantageous in evolutionary terms.

            The concept of the germ line holds that only germ cells are passed from one generation to the next. They do not originate from a germ layer (in the embryonic sense). This idea arose from work in lower forms, but was demonstrated in vertebrates when Bounoure (1934) showed that the vegetal region of fertilized frog eggs contains a material with special staining properties.  He was able to trace this cytoplasm, called germ plasm, through cleavage and gastrulation to a few large cells, the primordial germ cells.  These cells were located initially in the prospective endoderm, and migrated to the genital ridge. When those cells were removed or destroyed by irradiation with ultraviolet light, the frogs became sterile. They could produce no germ cells, though they were otherwise normal. If fragments of endoderm from an unirradiated gastrula that contained some of these cells were transplanted to an irradiated host embryo, the resulting frogs were again fertile. The importance of the germ plasm was confirmed when it was shown that ultraviolet irradiation of the vegetal region of the zygote also resulted in sterile frogs. But when vegetal cytoplasm that contained the germ plasm was transferred with a micropipette to an irradiated zygote, it counteracted the effects of the irradiation. The frogs were fertile. Thus, the germ plasm in frog zygotes contains a determinant for germ cell formation that is sensitive to ultraviolet light and can be transferred with the fluid portion of the cytoplasm.

            There has been considerable controversy concerning whether primordial germ cells, PGCs, arise in the gonads during embryogenesis or are derived from extragonadal origin.  Once it was established that germ cell lines in mammals were derived from an extragonadal origin, investigations focused on two questions: Where and when does the germ line arise?  Two possibilities exist: (1) germ cells are set aside before embryonic cells become committed to specific pathways, (2) germ cell lines appear de novo from other cells at some point in development.

            All the evidence (based on following Green Fluorescent Protein-tagged cells) leads to the conclusion that mammalian germ cells are not morphologically distinct during early development and are induced to form by surrounding cells. There is only fragmentary data for humans, but in mouse the story is coming clear and so far human data are in line with the mouse data. During primitive streak formation, extraembryonic ectoderm cells near the posterior end of the epiblast begin secreting BMPs. This induces some nearby cells to differentiate and express the transmembrane protein fragilis. A subset of these cells then begins to express the protein stella which is thought to encode an RNA splicing factor (though that is yet unproven). It is this last set of cells that give rise the the PGCs.

            It was long thought that PGCs migrate out of the epiblast, into the extraembryonic mesoderm, then back into the embryo via the allantois. That appears not to be true. Based on visualizing fluorescent live cells, the PGCs  migrate directly into the embryonic endoderm from the posterior region of the primitive streak. There they find themselves in the developing hindgut. Migration then proceeds from the posterior end of the gut to the genital ridge where they take up residence and complete differentiation. They continue to grow and divide during the migration such that the 10-100 cells that begin the process are ~5000 cell by the time they reach the ridge.

            Two major questions concerning the migration of PCGs in mammals remain unanswered: 1) What cues PCGs to migrate specifically to the gonadal anlage (i.e. the developing gonadal ridge), and 2) what mechanisms do PCGs use to accomplish this migration?

            Chemotaxis, with movement being directed by a substance emitted by the developing gonads, is the long suggested cue for PCG migration, but evidence is incomplete.  In vitro TGF-b-like proteins secreted by the mouse gonadal ridge are able to induce migration of mouse PGCs. A similar story exists in zebrafish where the known lymphocyte chemoattractant, SDF-1, seems to be used by PGCs. The in vivo significance of these observations is still unknown. Alternative mechanisms for directed cell movement include contact guidance, and differential adhesion using molecules likefibronectin.  A theory based on contact guidance proposes that motile PCGs move along paths determined by endodermal and mesodermal tissues.  The differential adhesion theory proposes the PCGs move to maximize the strength of their adhesive contacts.  Most evidence indicates that PCGs move by ameboid motion during the time of active migration.  The cell propels itself using an actomyosin-based force generating mechanism with small membrane specializations (filopodia) on the PCG cell membrane acting as attachment points for the cells during migration.

            The number of PCGs, as defined by alkaline phosphatase staining, changes greatly during the migratory period.  After completing migration, PCGs undergo divisional arrest.  Regulation of the cell division (both mitotic and meiotic) in PCGs is poorly understood and a major area of research.

            The development of the gonads, either testes or ovary, is directly influenced by the localization of the PGCs.  And the chromosome complement of a given set of PGCs results in the production of specific gene products that directly effect gonadal differentiation.  During embryonic weeks 5-7, in prospective females (chromosomal XX), germ cells localize in the cortex of the indifferent gonad and participate in the formation of an ovary, while the medulla regresses.  In males, primordial germ cells (chromosomally XY) migrate deeper to the medullary region of the gonad and are incorporated into the forming testis, and stimulate the primary sex cords to form seminiferous tubules.