Identification and dynamics of Cajal bodies in relation to karyosphere formation in scorpionfly oocytes

Abstract

In oocyte nuclei of the scorpionfly, Panorpa communis, we have recently defined a population of nuclear bodies (NBs) that contain some components of Cajal bodies (CBs). In the present study, we used several criteria [presence of coilin, U7 snRNA, RNA polymerase II (pol II) and specific ultrastructure] to identify these NBs as CBs. The essential evidence for CB identification came from experiments with microinjection of fluorescein-tagged U7 snRNA. Consistent with the U7 data, we found pol II and pre-mRNA splicing factor, SC35, in Panorpa oocyte CBs. We show here that the dynamics of CBs differs from that in somatic cells and correlates with the level of oocyte chromosome condensation. We also found that the significant increase of CB size is accompanied by condensation of the chromosomes in the karyosphere, which is indicative of a decline in transcription. Using immunogold microscopy we determined that pol II and coilin are shared by CBs and the granular material associated with condensed chromosomes in the Panorpa karyosphere. The colocalization of pol II, U7 snRNA and splicing factors with CBs at the inactive stage of late oogenesis suggests that the latter may serve as storage domains for components that were earlier engaged in RNA transcription and processing.

Factors related to RNA polymerase II transcription are localized in interchromatin granule clusters of Panorpa communis oocytes.

“…The nuclear morphology of P. communis oocytes presented here corresponds to that described earlier [11]. The general appearance of diplotene oocyte nucleus at the stage of late previtellogenesis is shown in Fig.…”

“…Ovarioles were prepared for immunogold labeling electron microscopy as described [11]. Primary antibodies included the following monoclones: αSC35 against the non-snRNP splicing factor SC35 [12], V22 against the hyperphosphorylated C-terminal domain (CTD) of the Pol II, and the following rabbit poly- Secondary antibodies were goat anti-mouse or goat anti-rabbit antibodies conjugated with colloidal gold particles of 10 nm (BBInternational).…”

“…Insect oocyte IGCs have been described only in a beetle [6,7], the house cricket [8], a fleshfly [9] and some bugs [10]. In our previous study, we showed some features of IGC structure and molecular composition in oocyte nuclei of the scorpionfly, Panorpa communis [11]. Importantly, we found that along with splicing factor SC35, a marker protein of IGCs [12,13], P. communis oocyte IGCs contain two (unphosphorylated and hyperphosphorylated) forms of RNA polymerase II (Pol II) [11].…”

“…In our previous study, we showed some features of IGC structure and molecular composition in oocyte nuclei of the scorpionfly, Panorpa communis [11]. Importantly, we found that along with splicing factor SC35, a marker protein of IGCs [12,13], P. communis oocyte IGCs contain two (unphosphorylated and hyperphosphorylated) forms of RNA polymerase II (Pol II) [11]. In the present study, we extend these data attempting to localize in the IGCs of P. communis oocytes additionally to Pol II some other components of Pol II holoenzyme and factors involved in activation of Pol II transcription.…”

“…Therefore, it would be interesting to trace a behaviour of Pol II holoenzyme components in association with the IGCs in transcriptionally inert oocytes. It is the reason for choosing of P. communis diplotene oocytes as an experimental model, because they possess a fully inactivated nuclei at this stage of oogenesis when chromosome condensation and karyosphere formation occur [11,17,18]. Thus, using immunogold labeling/electron microscopy we have mapped inside the diplotene oocyte IGCs of P. communis the following component of Pol II holoenzyme: Pol II itself and basal transcription factors, TFIID and TFIIH.…”