Morphological and immunological approach for studying the distribution pattern of mitochondria and germ granules during oogenesis of the guppy (Poecilia reticulata).

A same set of genes is associated to germline determination and differentiation in almost all Metazoa. Previous studies in several animals, also from distantly related taxa, showed a close association between germline determinants in germ granules and mitochondria, with observations at transmission electron microscopy and immunological approaches. However further investigations are needed to document their respective distribution and elucidate the role of mitochondria in the process of germ granule formation. In the present study we used an emerging animal model to study germline differentiation, Poecilia reticulata, also known as guppy, and different experimental approaches: western blot, immunolocalization, and transmission electron microscopy to investigate the distribution of mitochondria and germ granules during oogenesis. We used anti-Vasa, anti-TDRKH, and anti-TDRD7 to label germline markers, anti-TOMM20 to localize mitochondria, and anti-PLD6 to highlight germline mitochondria. Our observations in previtellogenic oocytes support the co-participation of the nucleus and mitochondria in the production of germ plasm-related material. In previtellogenic oocytes, immunodetection revealed the presence of the germline markers and PLD6 staining in the perinuclear area. The most striking evidence is the observation in the same cell type of plume-shaped structures that at electron microscopy appear as formed by mitochondrial aggregates intermingled with electron-dense germ granules distributed around the nuclear envelope. Overall, our results support the close association between germ granule and mitochondria during germline differentiation, strengthening the foundations for further insights.