Modern tools applied to classic structures: Approaches for mammalian male germ cell RNA granule research

Abstract

First reported in the 1800s, germ cell granules are small nonmembrane bound RNA‐rich regions of the cytoplasm. These sites of critical RNA processing and storage in the male germ cell are essential for proper differentiation and development and are present in a wide range of species from Caenorhabditis elegans through mammals. Initially characterized by light and electron microscopy, more modern techniques such as immunofluorescence and genetic models have played a major role in expanding our understanding of the composition of these structures. While these methods have given light to potential granule functions, much work remains to be done. The current expansion of imaging technologies and omics‐scale analyses to germ cell granule research will drive the field forward considerably. Many of these methods, both current and upcoming, have considerable caveats and limitations that necessitate a holistic approach to the study of germ granules. By combining and balancing different techniques, the field is poised to elucidate the nature of these critical structures.

Two RNA binding proteins, ADAD2 and RNF17, interact to form novel meiotic germ cell granules required for male fertility

“…Most genetic studies of male germ cell granules to date have utilized single gene knockout models [11,31,[41][42][43]. Although powerful for defining the function of the targeted protein, these approaches fail to inform on the function of the granules themselves as they also impact any non-granule functions of the chosen target [22]. Supporting this notion, single gene knockouts of granule-associated proteins often lead to phenotypes prior to formation of the granule [13,31].…”

“…Several previous observations support this hypothesis. First, as discussed above and recently reviewed [22], there are multiple proteins observed across many granules. Second, multiple granules have already been shown, by both EM and immunodetection, to actively share components [3,19,24,49].…”

“…The mammalian spermatocyte contains five distinct germ cell granules [3,24], as identified by electron microscopy (EM). These granules are crucial for proper germ cell development [5,6,31], though their exact protein composition and function are not wholly described [22]. In spite of this, several proteins are observed across four of the five granules, including DDX4 [19] and DDX25 [19,20].…”

“…In even greater contrast to the IMC is the one entirely unstudied meiotic germ cell granule, known only as the “cluster of 30-nm particles”, which has no defined resident proteins and has only been observed via electron microscopy. In addition to relatively poor composition and functional knowledge for the non-IMC granules, the exact functional role of or relationship between them has never been defined [22]. Similarly, whether the cluster of 30-nm particles or any of the other EM-defined granule types represent single, molecularly homogenous populations has been almost entirely unexplored.…”

New horizons in testis biology and men's health: a special issue of the XXVIth North American testis workshop

“…4 New ideas about Sertoli cells are presented in contributions from Dr. Isabelle Mansuy 7 along with a sophisticated approach to the identification of follicle stimulating hormone-responsive genes in immature Sertoli cells from Dr. Raj Kumar. 8 A review by Dr. Elizabeth Snyder identifies new ways in which RNA granules are being studied in male germ cells, 9 providing information about intracellular sites important to developmental regulation of spermatogenesis.…”