Reproduction is one of the major biological processes characteristic of all living species. In animals, sexual reproduction is the by far dominating mode of reproduction, although only half of the individuals (i.e. mothers) can produce offspring. The balance of this costly strategy is provided by the evolutionary advantage of the genetic recombination shuffle during meiosis in combination with sexual selection that exerts its pressure predominantly on males, the gender often providing little to the next generation beyond genes. The power of sexual selection is exemplified by extravagant traits for male-male competition in some species. Seemingly wasteful, these traits reflect the male’s competence to recruit resources from the environment and thus involve several genes spread over the genome. Females make use of these indicators of genome quality during mate choice.
The cellular basis of male reproduction are haploid spermatozoa, highly specialized cells functioning as motile genome vectors. Spermatozoa arise from a developmental process known as spermatogenesis. The process is fueled by spermatogonial stem cells and goes through three main phases: the initial mitotic phase, during which the number of spermatogonia increases rapidly, doubling with each round of mitosis; the meiotic phase (spermatocytes), during which genetic information is recombined and reduced to a haploid set; the final spermiogenic phase, during which the haploid spermatids emerging from meiosis differentiate, without further proliferation, into flagellated spermatozoa. Irrespective of the stage of development, germ cells cannot survive unless they receive support from a somatic cell type, known as Sertoli cells in vertebrates, which are in close, physical contact with the germ cells. In vertebrates, the endocrine system has evolved as master control system over spermatogenesis. Since Sertoli cells express the receptors for the two most important hormones regulating spermatogenesis, androgens produced in the testis and FSH produced in the pituitary, the Sertoli cell population is the main target of the endocrine regulation of spermatogenesis. However, also interstitial Leydig cells (producing androgens and growth factors) and peritubular myoid cells (producing growth factors) are testicular somatic cells responding to reproductive hormones and important for regulating spermatogenesis. The dependency on androgens links spermatogenesis to behavioral and morphological traits relevant in sexual selection.
Spermatogonial stem cells (SSCs) can be quiescent or cell-cycling. When cycling, they can either self-renew to produce more SSCs, or differentiate via a number of developmental stages into spermatozoa. The balance between self-renewal and differentiation is regulated. For example, SSC differentiation does not occur in juveniles, but is activated during puberty or the beginning of a reproductive season. Regulating the balance between self-renwal and differentiation is important also to avoid stem cell tumours or exhaustion of the SSC population. Paracrine factors derived from the stem cell’s environment determine this balance. Cell types contributing to this environment are Sertoli cells, myoid and Leydig and endothelial cells, that all are close to SSCs and contribute to the stem cell niche. The production and release of these factors is modulated by reproductive hormones, such as FSH and androgens. Information on the identity of these paracrine factors and their mode of action is largely missing in vertebrates. Hence, our main research question is, “How do hormones and growth factors regulate the proliferation and differentiation behavior of germ cells, in particular of spermatogonial stem cells?” Moreover, we are interested in what modulates production/release of hormones regulating spermatogenesis, in particular FSH.
Our experimental models are the zebrafish (Danio rerio), and in collaboration with other research groups (e.g., at the Institute of Marine Research in Bergen, Norway), also economically relevant species, such as the Atlantic salmon (Salmo salar).
The basic research concentrates on three aspects: (i) Identify candidate growth factors relevant for spermatogenesis via gene expression profiling (e.g. microarray; RNAseq [mRNA, miRNA, lncRNA]). (ii) Characterize the biological activity of identified candidate factors by expression profiling (RNAseq), loss-of-function as well as gain-of-function approaches. The functional approaches often use a primary testis tissue culture system for pharmacological approaches, or for testing recombinant hormones/growth factors; we also use genetic models, such as CRISPR/Cas-mediated gene knock out. (iii) Study the endocrine regulation of expression and/or release of identified candidate factors. And (iv) examine the endocrine regulation of pituitary FSH production and release. Points i to iii are mainly approached with the zebrafish model, points ii (in part) and iv (mainly) with the Atlantic salmon model. Finally, we start examining the use of nanobodies (salmon) but also GMO approaches (zebrafish) to induce sterility, a topic of great relevance for aquaculture biotechnology.
The equilibrium between stem cell self-renewal and differentiation is relevant also for applied research fields, such as aquaculture biotechnology and ecotoxicology. Considering aquaculture, sexual maturation or puberty poses significant economic, ecological, and animal welfare problems, for example for Atlantic salmon aquaculture. Puberty is associated with a switch from self-renewal to differentiation of stem cells. Understanding the physiological mechanisms controlling the switch in stem cell activity is the basis for developing approaches to delay the start of pubertal testis maturation. Moreover, sexually competent salmon escaping from aquaculture facilities (e.g. after storm-induced damage) bring about the risk of genetic introgression into native populations. Precocious male puberty and genetic introgression are two major problems limiting the further development of salmon aquaculture. The foreseeable decrease in resources versus increase in human population numbers, and several parameters of environmental relevance (e.g. feed conversion rate, carbon footprint, use of agricultural surface, protection of wild populations) place salmon aquaculture ahead of terrestrial animal protein production. However, the further improvement of this already rather efficient system requires attention. Considering ecotoxicology or endocrine disruption, it is in particular the expression profiling that provides several candidate genes as indicators for hormones or hormone-like substances that are found in surface freshwater bodies and affect the reproductive biology of freshwater species.